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UFGI publication round-up weeks 4/24, 5/1, 5/8/17

Versatile and precise gene-targeting strategies for functional studies in mammalian cell lines.

Author information: Wassef M1, Luscan A2, Battistella A3, Le Corre S3, Li H4, Wallace MR5, Vidaud M2, Margueron R3.

1Institut Curie, PSL research university, 75005 Paris, France; INSERM U934, Paris, France; CNRS UMR3215, Paris, France. Electronic address: michel.wassef@curie.fr.
2INSERM UMR_S745 et EA7331, Université Paris Descartes, Sorbonne Paris Cité, Facultée des Sciences Pharmaceutiques et Biologiques, 75006 Paris, France; Service de Biochimie et Génétique Moléculaire, Hôpital Cochin, Assistance Publique-Hôpitaux de Paris, 75014 Paris, France.
3Institut Curie, PSL research university, 75005 Paris, France; INSERM U934, Paris, France; CNRS UMR3215, Paris, France.
4Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville, FL, USA.
5Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville, FL, USA; University of Florida Health Cancer Center, University of Florida, Gainesville, FL, USA; University of Florida Genetics Institute, University of Florida, Gainesville, FL, USA.
Journal: Methods (San Diego, Calif.)

Date of e-pub: May 2017

Abstract: The advent of programmable nucleases such as ZFNs, TALENs and CRISPR/Cas9 has brought the power of genetic manipulation to widely used model systems such as model cell lines. In mammalian cells, nuclease-mediated DNA double strand break is mainly repaired through the error-prone non-homologous end-joining (NHEJ) repair pathway, eventually leading to accumulation of small deletions or insertions (indels) that can inactivate gene function. However, due to the variable size of the indels and the polyploid status of many cell lines (e.g., cancer-derived cells), obtaining a knockout usually requires lengthy screening and characterization procedures. Given the more precise type of modifications that can be introduced upon homology-directed repair (HDR), we have developed HDR-based gene-targeting strategies that greatly facilitate the process of knockout generation in cell lines. To generate reversible knockouts (R-KO), a selectable promoter-less STOP cassette is inserted in an intron, interrupting transcription. Loss-of-function can be validated by RT-qPCR and is removable, enabling subsequent restoration of gene function. A variant of the R-KO procedure can be used to introduce point mutations. To generate constitutive knockouts (C-KO), an exon is targeted, which makes use of HDR-based gene disruption together with NHEJ-induced indels on non-HDR targeted allele(s). Hence the C-KO procedure greatly facilitates simultaneous inactivation of multiple alleles. Overall these genome-editing tools offer superior precision and efficiency for functional genetic approaches. We provide detailed protocols guiding in the design of targeting vectors and in the analysis and validation of gene targeting experiments.

 

 

Consequences of MEGF10 deficiency on myoblast function and Notch1 interactions.

Author information: Saha M1, Mitsuhashi S2, Jones MD1, Manko K1, Reddy HM1, Bruels C1, Cho KA1,2, Pacak CA3, Draper I4, Kang PB1,2,5,6.

1Division of Pediatric Neurology, Department of Pediatrics, University of Florida College of Medicine, Gainesville, Florida.
2Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, Massachusetts.
3Child Health Research Institute, Department of Pediatrics, University of Florida College of Medicine, Gainesville, Florida.
4Molecular Cardiology Research Institute, Department of Medicine, Tufts Medical Center, Boston, Massachusetts.
5Department of Neurology and Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville, Florida.
6 Genetics Institute, University of Florida, Gainesville, Florida.
Journal: Human Molecular Genetics

Date of e-pub: May 2017

Abstract: Mutations in MEGF10 cause early onset myopathy, areflexia, respiratory distress, and dysphagia (EMARDD), a rare congenital muscle disease, but the pathogenic mechanisms remain largely unknown. We demonstrate that short hairpin RNA (shRNA)-mediated knockdown of Megf10, as well as overexpression of the pathogenic human p.C774R mutation, leads to impaired proliferation and migration of C2C12 cells. Myoblasts from Megf10-/- mice and Megf10-/-/mdx double knockout (dko) mice also show impaired proliferation and migration compared to myoblasts from wild type and mdx mice, whereas the dko mice show histological abnormalities that are not observed in either single mutant mouse. Cell proliferation and migration are known to be regulated by the Notch receptor, which plays an essential role in myogenesis. Reciprocal co-immunoprecipitation studies show that Megf10 and Notch1 interact via their respective intracellular domains. These interactions are impaired by the pathogenic p.C774R mutation. Megf10 regulation of myoblast function appears to be mediated at least in part via interactions with key components of the Notch signaling pathway, and defects in these interactions may contribute to the pathogenesis of EMARDD.

 

 

Authors’ Response to Jesse D. Riordan, Hum Gene Ther 2017;28:375-376; DOI: 10.1089/hum.2017.045.

Author information: Srivastava A1, Carter BJ2.

11 Division of Cellular and Molecular Therapy, Departments of Pediatrics and Molecular Genetics and Microbiology, Powell Gene Therapy Center, Genetics Institute, University of Florida College of Medicine , Gainesville, Florida.
22 BioMarin Pharmaceutical, Inc. , Novato, California.
Journal: Human Gene Therapy

Date of e-pub: May 2017

Abstract: N/A

 

 

Target enrichment sequencing in cultivated peanut (Arachis hypogaea L.) using probes designed from transcript sequences.

Author information: Peng Z1, Fan W1, Wang L1, Paudel D1, Leventini D1, Tillman BL1, Wang J2,3,4.

1Agronomy Department, University of Florida, Gainesville, FL, 32610, USA.
2Agronomy Department, University of Florida, Gainesville, FL, 32610, USA. wangjp@ufl.edu.
3Genetics Institute, Plant Molecular and Cellular Biology Program, University of Florida, Gainesville, FL, 32610, USA. wangjp@ufl.edu.
4Center for Genomics and Biotechnology, Key Laboratory of Genetics, Breeding and Multiple Utilization of Crops, Ministry of Education, Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Fujian Agriculture and Forestry University, Fuzhou, 350002, Fujian, China. wangjp@ufl.edu.
Journal: Molecular Genetics and Genomics : MGG

Date of e-pub: May 2017

Abstract: Enabled by the next generation sequencing, target enrichment sequencing (TES) is a powerful method to enrich genomic regions of interest and to identify sequence variations. The objective of this study was to explore the feasibility of probe design from transcript sequences for TES application in calling sequence variants in peanut, an important allotetraploid crop with a large genome size. In this study, we applied an in-solution hybridization method to enrich DNA sequences of seven peanut genotypes. Our results showed that it is feasible to apply TES with probes designed from transcript sequences in polyploid peanut. Using a set of 31,123 probes, a total of 5131 and 7521 genes were targeted in peanut A and B genomes, respectively. For each genotype used in this study, the probe target capture regions were efficiently covered with high depth. The average on-target rate of sequencing reads was 42.47%, with a significant amount of off-target reads coming from genomic regions homologous to target regions. In this study, when given predefined genomic regions of interest and the same amount of sequencing data, TES provided the highest coverage of target regions when compared to whole genome sequencing, RNA sequencing, and genotyping by sequencing. Single nucleotide polymorphism (SNP) calling and subsequent validation revealed a high validation rate (85.71%) of homozygous SNPs, providing valuable markers for peanut genotyping. This study demonstrated the success of applying TES for SNP identification in peanut, which shall provide valuable suggestions for TES application in other non-model species without a genome reference available.

 

 

Biopsychosocial influence on shoulder pain: Rationale and protocol for a pre-clinical trial.

Author information: George SZ1, Staud R2, Borsa PA3, Wu SS4, Wallace MR5, Greenfield WH6, Mackie LN7, Fillingim RB8.

1Duke Clinical Research Institute, Department of Orthopaedic Surgery, Duke University, Durham, NC 27715, USA. Electronic address: steven.george@duke.edu.
2Department of Medicine, Univ. of Florida, Gainesville, FL 32610, USA. Electronic address: staudr@ufl.edu.
3Applied Physiology and Kinesiology, Univ. of Florida, Gainesville, FL 32611, USA. Electronic address: pborsa@hhp.ufl.edu.
4Biostatistics, Univ. of Florida, Gainesville, FL 32610, USA. Electronic address: samwu@biostat.ufl.edu.
5Molecular Genetics and Microbiology, UF Genetics Institute, Univ. of Florida, Gainesville, FL 32610, USA. Electronic address: PhDpeggyw@mgm.ufl.edu.
6Department of Physical Therapy, Univ. of Florida, Gainesville, FL 32610, USA. Electronic address: whgiii@phhp.ufl.edu.
7School of Medicine, Saint Louis University, St. Louis, MO 63104, USA. Electronic address: mackieln@slu.edu.
8Pain Research & Intervention Center of Excellence, Univ. of Florida, Gainesville, FL 32610, USA. Electronic address: rfillingim@dental.ufl.edu.
Journal: Contemporary Clinical Trials

Date of e-pub: May 2017

Abstract: Chronic musculoskeletal pain conditions are a prevalent and disabling problem. Preventing chronic musculoskeletal pain requires multifactorial treatment approaches that address its complex etiology. Prior cohort studies identified a high risk subgroup comprised of variation in COMT genotype and pain catastrophizing. This subgroup had increased chance of heightened pain responses (in a pre-clinical model) and higher 12month post-operatives pain intensity ratings (in a clinical model). This pre-clinical trial will test mechanisms and efficacy of personalized pain interventions matched to the genetic and psychological characteristics of the high-risk subgroup.

Potential participants will be screened for high risk subgroup membership, appropriateness for exercise-induced muscle injury protocol, and appropriateness for propranolol administration. Eligible participants that consent to the study will then be randomized into one of four treatment groups; 1) personalized pharmaceutical and psychological education; 2) personalized pharmaceutical and general education; 3) placebo pharmaceutical and psychological education; 4) placebo pharmaceutical and psychological education. Over the 5-day study period participants will complete an exercise-induced muscle injury protocol and receive study interventions. Pain and disability assessments will be completed daily, with primary outcomes being duration of shoulder pain (number of days until recovery), peak shoulder pain intensity, and peak shoulder disability. Secondary outcomes include inflammatory markers, psychological mediators, and measures of pain sensitivity regulation.

This pre-clinical trial builds on prior cohort studies and its completion will provide foundational data supporting efficacy and mechanisms of personalized interventions for individuals that may be at increased risk for developing chronic shoulder pain.

ClinicalTrials.gov registry, NCT02620579 (Registered on November 13, 2015).

 

 

Neonatal sepsis in rural India: timing, microbiology and antibiotic resistance in a population-based prospective study in the community setting.

Author information: Panigrahi P1, Chandel DS2, Hansen NI3, Sharma N4, Kandefer S5, Parida S6, Satpathy R7, Pradhan L8, Mohapatra A9, Mohapatra SS7, Misra PR10, Banaji N11, Johnson JA12, Morris JG Jr13, Gewolb IH14, Chaudhry R4.

1Department of Epidemiology and Pediatrics, Center for Global Health and Development, College of Public Health, University of Nebraska Medical Center, Omaha, NE, USA.
2Department of Environmental, Agricultural, & Occupational Health, Center for Global Health and Development, College of Public Health, University of Nebraska Medical Center, Omaha, NE, USA.
3Division of Biostatistics & Epidemiology, RTI International, Research Triangle Park, NC, USA.
4Department of Microbiology, All India Institute of Medical Sciences, New Delhi, India.
5Division of Social Policy, Health, & Economics Research, RTI International, Research Triangle Park, NC, USA.
6Department of Pediatrics, SCB Medical College, Cuttack, Odisha, India.
7Social Pediatrics and Minority Health, Asian Institute of Public Health, Bhubaneswar, Odisha, India.
8Neonatal Unit, Department of Pediatrics, Capital Hospital, Bhubaneswar, Odisha, India.
9Center for Translational Research, Asian Institute of Public Health, Bhubaneswar, Odisha, India.
10Disease Surveillance Laboratories, Asian Institute of Public Health, Bhubaneswar, Odisha, India.
11Department of Microbiology, Indira Gandhi Medical College & Research Institute, Puducherry, India.
12Department of Pathology, Immunology and Laboratory Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, FL, USA.
13Department of Medicine, College of Medicine, Emerging Pathogens Institute, University of Florida, Gainesville FL, USA.
14Department of Pediatrics, Division of Neonatology, Sparrow Hospital – Neonatology, Lansing, MI, USA.
Journal: Journal of Perinatology : Official Journal of the California Perinatal Association

Date of e-pub: May 2017

Abstract: To examine the timing and microbiology of neonatal sepsis in a population-based surveillance in the Indian community setting.

All live born infants in 223 villages of Odisha state were followed at home for 60 days. Suspect sepsis cases were referred to study hospitals for further evaluation including blood culture.

Of 12 622 births, 842 were admitted with suspected sepsis of whom 95% were 4 to 60 days old. Culture-confirmed incidence of sepsis was 6.7/1000 births with 51% Gram negatives (Klebsiella predominating) and 26% Gram positives (mostly Staphylococcus aureus). A very high level of resistance to penicillin and ampicillin, moderate resistance to cephalosporins and extremely low resistance to Gentamicin and Amikacin was observed.

The bacterial burden of sepsis in the Indian community is not high. Judicious choice of empiric antibiotics, antibiotic stewardship and alternate modalities should be considered for the management or prevention of neonatal sepsis in India.Journal of Perinatology advance online publication, 11 May 2017; doi:10.1038/jp.2017.67.

 

 

Benzotriazole ultraviolet stabilizers alter the expression of the thyroid hormone pathway in zebrafish (Danio rerio) embryos.

Author information: Liang X1, Li J2, Martyniuk CJ3, Wang J2, Mao Y2, Lu H2, Zha J4.

1School of Ecology and Environment, Inner Mongolia University, Hohhot, 010021, China. Electronic address: liangxf@imu.edu.cn.
2School of Ecology and Environment, Inner Mongolia University, Hohhot, 010021, China.
3Department of Physiological Sciences and Center for Environmental and Human Toxicology, College of Veterinary Medicine, UF Genetics Institute, University of Florida, Gainesville, FL, 32611, USA.
4Key Laboratory of Drinking Water Science and Technology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China; Beijing Key Laboratory of Industrial Wastewater Treatment and Reuse, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing, 100085, China.
Journal: Chemosphere

Date of e-pub: May 2017

Abstract: Benzotriazole ultraviolet stabilizers (BUVSs) are widely used in industrial products as well as personal-hygiene products to protect the material or skin from harmful UV-radiation. Due to their persistence and bioaccumulation, BUVSs have been ubiquitously detected in aquatic environments. Although the toxicological effects of BUVSs in aquatic organisms have been previously examined, the effects of BUVSs on the thyroid system have not been adequately addressed. In this study, we assessed putative thyroid disrupting effects of BUVSs (UV-234, UV-326, UV-329 and UV-P) in zebrafish embryos at 1, 10 and 100 μg/L for 96 h. The heart rate was assessed in zebrafish and was observed to be decreased by 6.9%-21.4% in exposure of tested BUVSs. We also observed that the transcript levels of HPT axis-related genes were affected by the 4 BUVSs tested in different ways. Specifically, mRNA levels of thyroid hormone receptors (thraa and thrb) in zebrafish embryos were differentially expressed and the direction of change in these transcripts was isoform and BUVSs dependent. Pathway analysis of the targeted genes measured indicated that cellular processes putatively affected by BUVSs included response to organic substance, regulation of transcription from RNA polymerase II promoter, intracellular receptor signaling pathway, and hypothyroidism. Upon expansion of the network, novel genes involved in this predicted gene network may provide insight into the mechanisms of thyroid disrupting mechanisms of BUVSs. Taken together, our results indicate that BUVSs can potentially impact the thyroid system, and that this is dependent upon the type or structure of BUVSs.

 

 

Salinity Response in Chloroplasts: Insights from Gene Characterization.

Author information: Suo J1, Zhao Q2, David L3, Chen S4, Dai S5,6.

1Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline-alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040, China. suojinwei@nefu.edu.cn.
2Development Center of Plant Germplasm Resources, College of Life and Environmental Sciences, Shanghai Normal University, Shanghai 200234, China. zhaoqizq@yeah.net.
3Department of Biology, Genetics Institute, Plant Molecular and Cellular Biology Program, Interdisciplinary Center for Biotechnology Research, University of Florida, Gainesville, FL 32610, USA. lisaidavid@ufl.edu.
4Department of Biology, Genetics Institute, Plant Molecular and Cellular Biology Program, Interdisciplinary Center for Biotechnology Research, University of Florida, Gainesville, FL 32610, USA. schen@ufl.edu.
5Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline-alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040, China. daishaojun@hotmail.com.
6Development Center of Plant Germplasm Resources, College of Life and Environmental Sciences, Shanghai Normal University, Shanghai 200234, China. daishaojun@hotmail.com.
Journal: International Journal of Molecular Sciences

Date of e-pub: May 2017

Abstract: Salinity is a severe abiotic stress limiting agricultural yield and productivity. Plants have evolved various strategies to cope with salt stress. Chloroplasts are important photosynthesis organelles, which are sensitive to salinity. An understanding of molecular mechanisms in chloroplast tolerance to salinity is of great importance for genetic modification and plant breeding. Previous studies have characterized more than 53 salt-responsive genes encoding important chloroplast-localized proteins, which imply multiple vital pathways in chloroplasts in response to salt stress, such as thylakoid membrane organization, the modulation of photosystem II (PS II) activity, carbon dioxide (CO₂) assimilation, photorespiration, reactive oxygen species (ROS) scavenging, osmotic and ion homeostasis, abscisic acid (ABA) biosynthesis and signaling, and gene expression regulation, as well as protein synthesis and turnover. This review presents an overview of salt response in chloroplasts revealed by gene characterization efforts.

 

 

Professor Yuichi Sugiyama: A Brilliant, Creative, Amicable, Charming and Humorous Pharmaceutical Scientist.

Author information: Kusuhara H1, Obach RS2, Rostami-Hodjegan A3, Pang KS4, Hammarlund-Udenaes M5, Derendorf H6, Artursson P7, Huang SM8, Suzuki H9, Terasaki T10.

1Laboratory of Molecular Pharmacokinetics, Graduate School of Pharmaceutical Sciences, the University of Tokyo, Bunkyo-ku, Tokyo 113-0033, Japan. Electronic address: kusuhara@mol.f.u-tokyo.ac.jp.
2Department of Pharmacokinetics, Dynamics, and Metabolism, Pfizer Global Research and Development, Groton, Connecticut, USA.
3Centre for Applied Pharmacokinetic Research, University of Manchester, Manchester, UK; Simcyp Limited (A Certara Company), Sheffield, UK.
4Leslie Dan Faculty of Pharmacy, University of Toronto, Toronto, Canada.
5Translational Pharmacokinetics-Pharmacodynamics Group, Department of Pharmaceutical Biosciences, Uppsala University, Box 591, 75124 Uppsala, Sweden.
6Dept. of Pharmaceutics, University of Florida, Gainesville, FL 32608-0494, USA.
7Department of Pharmacy, Uppsala University, BMC, Box 580, Uppsala SE-751 23, Sweden.
8Office of Clinical Pharmacology, Office of Translational Sciences, Center for Drug Evaluation and Research, Food and Drug Administration, Silver Spring, MD, USA.
9Department of Pharmacy, the University of Tokyo Hospital, Faculty of Medicine, the University of Tokyo.
10Division of Membrane Transport and Drug Targeting, Graduate School of Pharmaceutical Sciences, Tohoku University, 6-3 Aoba, Aramaki, Aoba-ku, Sendai 980-8578, Japan.
Journal: Journal of Pharmaceutical Sciences

Date of e-pub: May 2017

Abstract: N/A

 

 

Structural Insight into Ubiquitin-Like Protein Recognition and Oligomeric States of JAMM/MPN+ Proteases.

Author information: Cao S1, Engilberge S2, Girard E2, Gabel F2, Franzetti B3, Maupin-Furlow JA4.

1Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL 32611, USA.
2Institut de Biologie Structurale (IBS), University Grenoble Alpes, CEA, CNRS, 38044 Grenoble, France.
3Institut de Biologie Structurale (IBS), University Grenoble Alpes, CEA, CNRS, 38044 Grenoble, France. Electronic address: franzetti@ibs.fr.
4Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL 32611, USA; Genetics Institute, University of Florida, Gainesville, FL 32611, USA. Electronic address: jmaupin@ufl.edu.
Journal: Structure (London, England : 1993)

Date of e-pub: May 2017

Abstract: JAMM/MPN+ metalloproteases cleave (iso)peptide bonds C-terminal to ubiquitin (Ub) and ubiquitin-like protein (Ubl) domains and typically require association with protein partners for activity, which has limited a molecular understanding of enzyme function. To provide an insight, we solved the X-ray crystal structures of a catalytically active Pyrococcus furiosus JAMM/MPN+ metalloprotease (PfJAMM1) alone and in complex with a Ubl (PfSAMP2) to 1.7- to 1.9-Å resolution. PfJAMM1 was found to have a redox sensitive dimer interface. In the PfJAMM1-bound state of the SAMP2, a Ubl-to-Ub conformational change was detected. Surprisingly, distant homologs of PfJAMM1 were found to be closely related in 3D structure, including the interface for Ubl/Ub binding. From this work, we infer the molecular basis of how JAMM/MPN+ proteases recognize and cleave Ubl/Ub tags from diverse proteins and highlight an α2-helix structural element that is conserved and crucial for binding and removing the Ubl SAMP2 tag.

 

 

De Novo Coding Variants Are Strongly Associated with Tourette Disorder.

Author information: Willsey AJ1, Fernandez TV2, Yu D3, King RA2, Dietrich A4, Xing J5, Sanders SJ6, Mandell JD1, Huang AY7, Richer P8, Smith L6, Dong S6, Samocha KE3; Tourette International Collaborative Genetics (TIC Genetics); Tourette Syndrome Association International Consortium for Genetics (TSAICG), Neale BM3, Coppola G7, Mathews CA9, Tischfield JA5, Scharf JM10, State MW11, Heiman GA12.

1Department of Psychiatry, UCSF Weill Institute for Neurosciences, University of California San Francisco, San Francisco, CA 94143, USA; Institute for Neurodegenerative Diseases, UCSF Weill Institute for Neurosciences, University of California San Francisco, San Francisco, CA 94143, USA.
2Yale Child Study Center and Department of Psychiatry, Yale University School of Medicine, New Haven, CT 06520, USA.
3Center for Genomic Medicine, Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA; Psychiatric and Neurodevelopmental Genetics Unit, Department of Psychiatry, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA.
4University of Groningen, University Medical Center Groningen, Department of Child and Adolescent Psychiatry, 9713GZ Groningen, the Netherlands.
5Rutgers, the State University of New Jersey, Department of Genetics and the Human Genetics Institute of New Jersey, Piscataway, NJ 08854, USA.
6Department of Psychiatry, UCSF Weill Institute for Neurosciences, University of California San Francisco, San Francisco, CA 94143, USA.
7Department of Neurology, University of California Los Angeles, Los Angeles, California, CA 90095, USA; Department of Psychiatry and Biobehavioral Sciences, University of California Los Angeles, Los Angeles, CA 90095, USA.
8Yale Child Study Center and Department of Psychiatry, Yale University School of Medicine, New Haven, CT 06520, USA; Sewanee: The University of the South, Sewanee, TN 37383, USA.
9Department of Psychiatry, University of Florida School of Medicine, Gainesville, FL 32611, USA.
10Center for Genomic Medicine, Department of Neurology, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA; Psychiatric and Neurodevelopmental Genetics Unit, Department of Psychiatry, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114, USA. Electronic address: jscharf@partners.org.
11Department of Psychiatry, UCSF Weill Institute for Neurosciences, University of California San Francisco, San Francisco, CA 94143, USA. Electronic address: matthew.state@ucsf.edu.
12Rutgers, the State University of New Jersey, Department of Genetics and the Human Genetics Institute of New Jersey, Piscataway, NJ 08854, USA. Electronic address: heiman@dls.rutgers.edu.
Journal: Neuron

Date of e-pub: May 2017

Abstract: Whole-exome sequencing (WES) and de novo variant detection have proven a powerful approach to gene discovery in complex neurodevelopmental disorders. We have completed WES of 325 Tourette disorder trios from the Tourette International Collaborative Genetics cohort and a replication sample of 186 trios from the Tourette Syndrome Association International Consortium on Genetics (511 total). We observe strong and consistent evidence for the contribution of de novo likely gene-disrupting (LGD) variants (rate ratio [RR] 2.32, p = 0.002). Additionally, de novo damaging variants (LGD and probably damaging missense) are overrepresented in probands (RR 1.37, p = 0.003). We identify four likely risk genes with multiple de novo damaging variants in unrelated probands: WWC1 (WW and C2 domain containing 1), CELSR3 (Cadherin EGF LAG seven-pass G-type receptor 3), NIPBL (Nipped-B-like), and FN1 (fibronectin 1). Overall, we estimate that de novo damaging variants in approximately 400 genes contribute risk in 12% of clinical cases.

 

 

Resolving relationships among the megadiverse butterflies and moths with a novel pipeline for Anchored Phylogenomics.

Author information: Breinholt JW1,2, Earl C1, Lemmon AR3, Moriarty Lemmon E3, Xiao L1, Kawahara AY1.

1Florida Museum of Natural History, University of Florida, Gainesville, FL 32611, USA.
2RAPiD Genomics, Gainesville, FL 32601, USA.
3Dept. of Biological Science, Florida State University, Tallahassee, FL 32306, USA.
Journal: Systematic biology

Date of e-pub: May 2017

Abstract: The advent of next-generation sequencing technology has allowed for the collection of large portions of the genome for phylogenetic analysis. Hybrid enrichment and transcriptomics are two techniques that leverage next-generation sequencing and have shown much promise. However, methods for processing hybrid enrichment data are still limited. We developed a pipeline for anchored hybrid enrichment (AHE) read assembly, orthology determination, contamination screening, and data processing for sequences flanking the target “probe” region. We apply this approach to study the phylogeny of butterflies and moths (Lepidoptera), a megadiverse group of more than 157,000 described species with poorly understood deep-level phylogenetic relationships. We introduce a new, 855 locus anchored hybrid enrichment kit for Lepidoptera phylogenetics and compare resulting trees to those from transcriptomes. The enrichment kit was designed from existing genomes, transcriptomes and expressed sequence tag (EST) data and was used to capture sequence data from 54 species from 23 lepidopteran families. Phylogenies estimated from AHE data were largely congruent with trees generated from transcriptomes, with strong support for relationships at all but the deepest taxonomic levels. We combine AHE and transcriptomic data to generate a new Lepidoptera phylogeny, representing 76 exemplar species in 42 families. The tree provides robust support for many relationships, including those among the seven butterfly families. The addition of AHE data to an existing transcriptomic dataset lowers node support along the Lepidoptera backbone, but firmly places taxa with AHE data on the phylogeny. To examine the efficacy of AHE at different taxonomic levels, phylogenetic analyses were also conducted on a sister group representing a more recent divergence, the Saturniidae and Sphingidae. These analyses utilized sequences from the probe region and data flanking it, nearly doubled the size of the dataset; all resulting trees were well supported. We hope that our data processing pipeline, hybrid enrichment gene set, and approach of combining AHE data with transcriptomes will be useful for the broader systematics community.

 

 

Phenotypic characterisation of cell populations in the brains of horses experimentally infected with West Nile Virus.

Author information: Delcambre GH1, Liu J2, Streit WJ3, Shaw GPJ3, Vallario K2, Herrington J2, Wenzlow N2, Barr KL2, Long MT2.

1Department of Biomedical Sciences, College of Veterinary Medicine and Biomedical Sciences, Colorado State University, Fort Collins, Colorado, 80523-1617, USA.
2Department of Infectious Diseases and Pathology, College of Veterinary Medicine, University of Florida, P.O. Box 110880, Gainesville, Florida, 32611-0880, USA.
3Department of Neuroscience, College of Medicine, University of Florida, 1149 Newell Drive, Gainesville, FL, 32611-0244, USA.
Journal: Equine Veterinary Journal

Date of e-pub: May 2017

Abstract: West Nile Virus (WNV), a mosquito borne member of the Flaviviridae, is one of the most commonly diagnosed agents of viral encephalitis in horses and people worldwide.

A cassette of markers for formalin-fixed paraffin-embedded (FFPE) tissue and an archive of tissues from experimental infections in the horse were used to investigate the equine neuroimmune response to WNV meningoencephalomyelitis to phenotype the early response to WNV infection in the horse.

Quantitative analysis using archived tissue from experimentally infected horses.

The thalamus and hindbrain from two groups of six horses were compared and consisted of a culture positive tissues from WNV experimentally horses, in the other, normal horses. FFPE from the thalamus and hindbrain were immunolabeled for microglia, astrocytes, B cells, macrophages/neutrophils, CD3+ T cells. Fresh frozen tissues were immunolabeled for CD4+ and CD8+ T lymphocyte cell markers. Cell counts were obtained using a computer software program. Differences, after meeting assumptions of abnormality, were computed using a general linear model with a Tukey test (p<0.05) for pairwise comparisons.

In WNV-challenged horses, Iba-1+ microglia, CD3+ T lymphocyte, and MAC387+ macrophage staining were significantly increased. The T cell response for the WNV-challenged horses was mixed, composed of CD4+ and CD8+ T lymphocytes. A limited astrocyte response was also observed in WNV-challenged horses, and MAC387+ and B cells were the least abundant cell populations.

The results of this study were limited by a single collection time post-infection. Furthermore, a comprehensive analysis of cellular phenotypes is needed for naturally infected horses. Unfortunately, in clinical horses, there is high variability of sampling in terms of days post infection and tissue handling.

The data show that WNV-challenged horses recruit a mixed T cell population at the onset of neurologic disease. This article is protected by copyright. All rights reserved.

 

 

Plasmacytoid and conventional dendritic cells cooperate in cross-priming AAV capsid-specific CD8+ T cells.

Author information: Rogers GL1, Shirley JL1, Zolotukhin I1, Kumar SRP1, Sherman A1, Perrin GQ1, Hoffman BE1, Srivastava A1, Basner-Tschakarjan E2, Wallet MA3, Terhorst C4, Biswas M1, Herzog RW5.

1University of Florida, Department of Pediatrics, Division of Cellular and Molecular Therapy, Gainesville, FL, United States.
2Center of Molecular and Cellular Therapeutics, The Children’s Hospital of Philadelphia, Philadelphia, PA, United States.
3University of Florida, Department of Pathology, Immunology and Laboratory Medicine, Gainesville, FL, United States.
4Division of Immunology, BIDMC, Harvard Medical School, Boston, MA, United States.
5University of Florida, Department of Pediatrics, Division of Cellular and Molecular Therapy, Gainesville, FL, United States; rherzog@ufl.edu.
Journal: Blood

Date of e-pub: May 2017

Abstract: Adeno-associated virus (AAV) is a replication-deficient parvovirus that is extensively used as a gene therapy vector. CD8+ T cell responses against the AAV capsid protein can, however, affect therapeutic efficacy. Little is known about the in vivo mechanism that leads to the cross-priming of CD8+ T cells against the input viral capsid antigen. Here we report that the TLR9-MyD88 pattern-recognition receptor pathway is uniquely capable of initiating this response. By contrast, the absence of TLR2, STING, or addition of TLR4 agonist has no effect. Surprisingly, both conventional (cDCs) and plasmacytoid dendritic cells (pDCs) are required for the cross-priming of capsid-specific CD8+ T cells, while other antigen presenting cells are not involved. TLR9 signaling is specifically essential in pDCs but not in cDCs, indicating that sensing of the viral genome by pDCs activates cDCs in trans to cross-present capsid antigen during CD8+ T cell activation. Cross-presentation and cross-priming depend not only on TLR9, but also on IFN I signaling, and both mechanisms can be inhibited by administering specific molecules to prevent induction of capsid-specific CD8+ T cells. Thus, these outcomes directly point to therapeutic interventions and demonstrate that innate immune blockade can eliminate unwanted immune responses in gene therapy.

 

 

Redox signaling in neurotransmission and cognition during aging.

Author information: Kumar A1,2, Yegla B3, Foster TC4.

1University of Florida, Neuroscience , 1049 Newell Drive , Brain Institute, Blg 59, Room # L2-132 , FLORIDA , Gainesville, Florida, United States , 32611-7011.
21049 Newell DriveUnited States ; kash@ufl.edu.
3University of Florida McKnight Brain Institute, 137847, Gainesville, Florida, United States ; britney.yegla@ufl.edu.
4University of Florida, Department of Neuroscience, McKnight Brain Institute, Gainesville, Florida, United States ; foster1@ufl.edu.
Journal: Antioxidants & Redox Signaling

Date of e-pub: May 2017

Abstract: Oxidative stress increases in the brain with aging and neurodegenerative diseases. Previous work emphasized irreversible oxidative damage in relation to cognitive impairment. This research has evolved to consider a continuum of alterations, from redox signaling to oxidative damage, which provides a basis for understanding the onset and progression of cognitive impairment. This review provides an update on research linking redox signaling to altered function of neural circuits involved in information processing and memory.

Starting in middle-age, redox signaling triggers changes in nervous system physiology described as senescent physiology. Recent studies indicates N-methyl-D-aspartate and ryanodine receptors, and Ca2+ signaling molecules as molecular substrates of redox-mediated senescent physiology.

We review redox homeostasis mechanisms and consider the chemical character of reactive oxygen and nitrogen species, and their role in regulating different transmitter systems. In this regard, senescent physiology may represent the co-opting of pathways normally responsible for feedback regulation of synaptic transmission.

It will be important to identify the intrinsic mechanisms for the shift in oxidative/reductive processes. Intrinsic mechanism will depend on the transmitter system, the oxidative stressors, and the expression/activity of antioxidant enzymes. In addition, it will be important to identify how intrinsic processes interact with other aging factors including changes in inflammatory or hormonal signals.

Results suggest that physiological rather than pathological mechanisms underlie for the initial diagnosis of cognitive impairment. Because redox signaling is reversible, it provides hope for early identification and treatment of cognitive decline.

 

 

A KCNC3 mutation causes a neurodevelopmental, non-progressive SCA13 subtype associated with dominant negative effects and aberrant EGFR trafficking.

Author information: Khare S1,2,3, Nick JA1,2, Zhang Y4, Galeano K1,2, Butler B2,5, Khoshbouei H2,5, Rayaprolu S5, Hathorn T6, Ranum LPW6, Smithson L2,5, Golde TE2,5, Paucar M7,8, Morse R9, Raff M10, Simon J10, Nordenskjöld M11,12, Wirdefeldt K8,13, Rincon-Limas DE1,2, Lewis J2,5, Kaczmarek LK4, Fernandez-Funez P1,2, Nick HS2,5, Waters MF1,2,3,5.

1Department of Neurology, University of Florida, Gainesville, FL, United States of America.
2McKnight Brain Institute, University of Florida, Gainesville, FL, United States of America.
3Department of Biomedical Engineering, University of Florida, Gainesville, FL, United States of America.
4Department of Pharmacology, Yale University, New Haven, CT, United States of America.
5 Department of Neuroscience, University of Florida, Gainesville, FL, United States of America.
6Department of Molecular Genetics and Microbiology, University of Florida, Gainesville, FL, United States of America.
7Department of Neurology, Karolinska University Hospital, Stockholm, Sweden.
8Department of Clinical Neuroscience, Karolinska Institute, Stockholm, Sweden.
9Department of Neurology, Dartmouth-Hitchcock Medical Center, Lebanon, NH, United States of America.
10Genomics Institute, Multicare Health System, Tacoma, WA, United States of America.
11Department of Genetics, Karolinska University Hospital, Stockholm, Sweden.
12Department of Molecular Medicine and Surgery, Karolinska Institute, Center for Molecular Medicine, Stockholm, Sweden.
13Department of Medical Epidemiology and Biostatistics, Karolinska Institute, Stockholm, Sweden.
Journal: PLoS One

Date of e-pub: May 2017

Abstract: The autosomal dominant spinocerebellar ataxias (SCAs) are a diverse group of neurological disorders anchored by the phenotypes of motor incoordination and cerebellar atrophy. Disease heterogeneity is appreciated through varying comorbidities: dysarthria, dysphagia, oculomotor and/or retinal abnormalities, motor neuron pathology, epilepsy, cognitive impairment, autonomic dysfunction, and psychiatric manifestations. Our study focuses on SCA13, which is caused by several allelic variants in the voltage-gated potassium channel KCNC3 (Kv3.3). We detail the clinical phenotype of four SCA13 kindreds that confirm causation of the KCNC3R423H allele. The heralding features demonstrate congenital onset with non-progressive, neurodevelopmental cerebellar hypoplasia and lifetime improvement in motor and cognitive function that implicate compensatory neural mechanisms. Targeted expression of human KCNC3R423H in Drosophila triggers aberrant wing veins, maldeveloped eyes, and fused ommatidia consistent with the neurodevelopmental presentation of patients. Furthermore, human KCNC3R423H expression in mammalian cells results in altered glycosylation and aberrant retention of the channel in anterograde and/or endosomal vesicles. Confirmation of the absence of plasma membrane targeting was based on the loss of current conductance in cells expressing the mutant channel. Mechanistically, genetic studies in Drosophila, along with cellular and biophysical studies in mammalian systems, demonstrate the dominant negative effect exerted by the mutant on the wild-type (WT) protein, which explains dominant inheritance. We demonstrate that ocular co-expression of KCNC3R423H with Drosophila epidermal growth factor receptor (dEgfr) results in striking rescue of the eye phenotype, whereas KCNC3R423H expression in mammalian cells results in aberrant intracellular retention of human epidermal growth factor receptor (EGFR). Together, these results indicate that the neurodevelopmental consequences of KCNC3R423H may be mediated through indirect effects on EGFR signaling in the developing cerebellum. Our results therefore confirm the KCNC3R423H allele as causative for SCA13, through a dominant negative effect on KCNC3WT and links with EGFR that account for dominant inheritance, congenital onset, and disease pathology.

 

 

Preemptive Panel-Based Pharmacogenetic Testing: The Time is Now.

Author information: Weitzel KW1,2, Cavallari LH3,4, Lesko LJ5.

1Department of Pharmacotherapy and Translational Research, and Center for Pharmacogenomics, University of Florida College of Pharmacy, PO Box 100486, Gainesville, Florida, 32610-0486, USA. kweitzel@cop.ufl.edu.
2UF Health Personalized Medicine Program,, Gainesville, Florida, USA. kweitzel@cop.ufl.edu.
3Department of Pharmacotherapy and Translational Research, and Center for Pharmacogenomics, University of Florida College of Pharmacy, PO Box 100486, Gainesville, Florida, 32610-0486, USA.
4UF Health Personalized Medicine Program,, Gainesville, Florida, USA.
5Department of Pharmaceutics, University of Florida College of Pharmacy, Orlando, Florida, USA.
Journal: Pharmaceutical Research

Date of e-pub: May 2017

Abstract: While recent discoveries have paved the way for the use of genotype-guided prescribing in some clinical environments, significant debate persists among clinicians and researchers about the optimal approach to pharmacogenetic testing in clinical practice. One crucial factor in this debate surrounds the timing and methodology of genotyping, specifically whether genotyping should be performed reactively for targeted genes when a single drug is prescribed, or preemptively using a panel-based approach prior to drug prescribing. While early clinical models that employed a preemptive approach were largely developed in academic health centers through multidisciplinary efforts, increasing examples of pharmacogenetic testing are emerging in community-based and primary care practice environments. However, educational and practice-based resources for these clinicians remain largely nonexistent. As such, there is a need for the health care system to shift its focus from debating about preemptive genotyping to developing and disseminating needed resources to equip frontline clinicians for clinical implementation of pharmacogenetics. Providing tools and guidance to support these emerging models of care will be essential to support the thoughtful, evidence-based use of pharmacogenetic information in diverse clinical practice environments. Specifically, the creation of efficient and accurate point-of-care resources, practice-based tools, and clinical models is needed, along with identification and dissemination of sustainable avenues for pharmacogenetic test reimbursement.

 

 

The bovine embryo hatches from the zona pellucida through either the embryonic or abembryonic pole.

Author information: Negrón-Pérez VM1, Hansen PJ2.

1Department of Animal Sciences, D. H. Barron Reproductive and Perinatal Biology Research Program and Genetics Institute, University of Florida, Gainesville, Florida, USA.
2Department of Animal Sciences, D. H. Barron Reproductive and Perinatal Biology Research Program and Genetics Institute, University of Florida, Gainesville, Florida, USA. pjhansen@ufl.edu.
Journal: Journal of Assisted Reproduction and Genetics

Date of e-pub: May 2017

Abstract: Implantation of the mammalian embryo in the uterus is preceded by escape from the zona pellucida. In some species, hatching from the zona occurs preferentially from one or the other poles of the embryo. The situation for the bovine embryo, in which hatching precedes attachment to the uterus by more than a week, is unclear. The purpose was to describe whether hatching of the bovine embryo from the zona pellucida occurs preferentially from the embryonic or abembryonic pole.

Bovine blastocysts undergoing hatching were examined by light microscopy (n = 84) and epifluorescence imaging using antibodies for markers of epiblast, hypoblast, and trophectoderm (TE) (n = 26). The location of hatching was classified as being at the embryonic pole, if hatching occurred ipsilateral to the inner cell mass (ICM), or abembryonic, if hatching occurred contralateral to the ICM.

A total of 55% of blastocysts exited the zona pellucida through an opening at the embryonic pole. In these cases, 68% of the cells emerging through the zona pellucida were derived from the ICM. The remainder of blastocysts hatched from an opening either contralateral or to the side of the ICM. In these cases, 87% of hatched cells were TE.

For the bovine embryo, there is nearly equal probability of hatching from the embryonic or abembryonic poles. Given that the surface area of the zona pellucida in contact with the TE overlying the ICM is less than for the remainder of the blastocyst, there is some preference for hatching through the embryonic pole. Thus, the bovine embryo is distinct from the mouse and human, where hatching occurs preferentially at the abembryonic pole.

 

 

Old Plants, New Tricks: Phenological Research Using Herbarium Specimens.

Author information: Willis CG1, Ellwood ER2, Primack RB3, Davis CC4, Pearson KD5, Gallinat AS3, Yost JM6, Nelson G5, Mazer SJ7, Rossington NL7, Sparks TH8, Soltis PS9.

1Department of Organismic and Evolutionary Biology and Harvard University Herbaria, Harvard University, Cambridge, MA 02138, USA. Electronic address: charleswillis@fas.harvard.edu.
2Department of Biological Science, Florida State University, Tallahassee, FL 32306, USA. Electronic address: eellwood@bio.fsu.edu.
3Biology Department, Boston University, Boston, MA 02215, USA.
4Department of Organismic and Evolutionary Biology and Harvard University Herbaria, Harvard University, Cambridge, MA 02138, USA.
5Department of Biological Science, Florida State University, Tallahassee, FL 32306, USA.
6Biological Science Department and Robert F. Hoover Herbarium, California Polytechnic State University, San Luis Obispo, CA 93407, USA.
7Department of Ecology, Evolution, and Marine Biology, University of California, Santa Barbara, Santa Barbara, CA 93106, USA.
8Coventry University, Coventry CV1 5FB, UK; Poznań University of Life Sciences, 60-625 Poznań, Poland.
9Florida Museum of Natural History, University of Florida, Gainesville, FL 32611, USA.
Journal: Trends in Ecology & Evolution

Date of e-pub: April 2017

Abstract: The timing of phenological events, such as leaf-out and flowering, strongly influence plant success and their study is vital to understanding how plants will respond to climate change. Phenological research, however, is often limited by the temporal, geographic, or phylogenetic scope of available data. Hundreds of millions of plant specimens in herbaria worldwide offer a potential solution to this problem, especially as digitization efforts drastically improve access to collections. Herbarium specimens represent snapshots of phenological events and have been reliably used to characterize phenological responses to climate. We review the current state of herbarium-based phenological research, identify potential biases and limitations in the collection, digitization, and interpretation of specimen data, and discuss future opportunities for phenological investigations using herbarium specimens.

 

 

Fatal Systemic Salmonellosis in a Florida Manatee ( Trichechus manatus latirostris).

Author information: Vorbach BS1,2, Rotstein DS3, Stacy NI2, Mavian C4, Salemi M4, Waltzek TB5, de Wit M1.

1Marine Mammal Pathobiology Lab, Florida Fish and Wildlife Conservation Commission, St. Petersburg, Florida 33711, USA.
2Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Florida, Gainesville, Florida 32611, USA.
3Marine Mammal Pathology Services, Olney, Maryland 20832, USA.
4Department of Pathology, Immunology and Laboratory Medicine, College of Medicine, Emerging Pathogens Institute, University of Florida, Gainesville, Florida 32610, USA.
5Department of Infectious Diseases and Pathology, College of Veterinary Medicine, University of Florida, Gainesville, Florida 32611, USA.
Journal: Journal of Wildlife Diseases

Date of e-pub: May 2017

Abstract: A subadult male Florida manatee ( Trichechus manatus latirostris) stranded dead on Florida’s Atlantic coast in January 2015. Necropsy and histopathologic findings confirmed chronic systemic bacterial infection caused by Salmonella enterica serotype IV 50:z4,z23,:- involving renal, respiratory, lymphatic, and skeletal systems. This was a unique case of systemic salmonellosis in a Florida manatee.

 

 

Genetic and Small Molecule Disruption of the AID/RAD51 Axis Similarly Protects Nonobese Diabetic Mice from Type 1 Diabetes through Expansion of Regulatory B Lymphocytes.

Author information: Ratiu JJ1, Racine JJ1, Hasham MG1, Wang Q1,2, Branca JA1, Chapman HD1, Zhu J3, Donghia N1, Philip V1, Schott WH1, Wasserfall C4, Atkinson MA4, Mills KD5, Leeth CM6, Serreze DV7.

1The Jackson Laboratory, Bar Harbor, ME 04609.
2Graduate Program in Genetics, Sackler School of Graduate Biomedical Sciences, Tufts University, Boston, MA 02111.
3Department of Animal and Poultry Sciences, Virginia Polytechnic and State University, Blacksburg, VA 24061.
4Department of Pathology, University of Florida, Gainesville, FL 32610; and.
5Cyteir Therapeutics, Cambridge, MA 02138.
6Department of Animal and Poultry Sciences, Virginia Polytechnic and State University, Blacksburg, VA 24061; dave.serreze@jax.org cmcphee@vt.edu.
7The Jackson Laboratory, Bar Harbor, ME 04609; dave.serreze@jax.org cmcphee@vt.edu.
Journal: Journal of Immunology (Baltimore, Md. : 1950)

Date of e-pub: May 2017

Abstract: B lymphocytes play a key role in type 1 diabetes (T1D) development by serving as a subset of APCs preferentially supporting the expansion of autoreactive pathogenic T cells. As a result of their pathogenic importance, B lymphocyte-targeted therapies have received considerable interest as potential T1D interventions. Unfortunately, the B lymphocyte-directed T1D interventions tested to date failed to halt β cell demise. IgG autoantibodies marking humans at future risk for T1D indicate that B lymphocytes producing them have undergone the affinity-maturation processes of class switch recombination and, possibly, somatic hypermutation. This study found that CRISPR/Cas9-mediated ablation of the activation-induced cytidine deaminase gene required for class switch recombination/somatic hypermutation induction inhibits T1D development in the NOD mouse model. The activation-induced cytidine deaminase protein induces genome-wide DNA breaks that, if not repaired through RAD51-mediated homologous recombination, result in B lymphocyte death. Treatment with the RAD51 inhibitor 4,4′-diisothiocyanatostilbene-2, 2′-disulfonic acid also strongly inhibited T1D development in NOD mice. The genetic and small molecule-targeting approaches expanded CD73+ B lymphocytes that exert regulatory activity suppressing diabetogenic T cell responses. Hence, an initial CRISPR/Cas9-mediated genetic modification approach has identified the AID/RAD51 axis as a target for a potentially clinically translatable pharmacological approach that can block T1D development by converting B lymphocytes to a disease-inhibitory CD73+ regulatory state.

 

 

Evaluation of engineered AAV capsids for hepatic factor IX gene transfer in murine and canine models.

Author information: Markusic DM1, Nichols TC2, Merricks EP2, Palaschak B3, Zolotukhin I3, Marsic D3, Zolotukhin S3, Srivastava A3, Herzog RW4.

1Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA. dmarkusic@ufl.edu.
2Department of Pathology and Laboratory Medicine, University of North Carolina-Chapel Hill, Chapel Hill, NC, 27599, USA.
3Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA.
4Department of Pediatrics, University of Florida, Gainesville, FL, 32610, USA. rherzog@ufl.edu.
Journal: Journal of Translational Medicine

Date of e-pub: May 2017

Abstract: Adeno-associated virus (AAV) gene therapy vectors have shown the best outcomes in human clinical studies for the treatment of genetic diseases such as hemophilia. However, these pivotal investigations have also identified several challenges. For example, high vector doses are often used for hepatic gene transfer, and cytotoxic T lymphocyte responses against viral capsid may occur. Therefore, achieving therapy at reduced vector doses and other strategies to reduce capsid antigen presentation are desirable.

We tested several engineered AAV capsids for factor IX (FIX) expression for the treatment of hemophilia B by hepatic gene transfer. These capsids lack potential phosphorylation or ubiquitination sites, or had been generated through molecular evolution.

AAV2 capsids lacking either a single lysine residue or 3 tyrosine residues directed substantially higher coagulation FIX expression in mice compared to wild-type sequence or other mutations. In hemophilia B dogs, however, expression from the tyrosine-mutant vector was merely comparable to historical data on AAV2. Evolved AAV2-LiC capsid was highly efficient in hemophilia B mice but lacked efficacy in a hemophilia B dog.

Several alternative strategies for capsid modification improve the in vivo performance of AAV vectors in hepatic gene transfer for correction of hemophilia. However, capsid optimization solely in mouse liver may not predict efficacy in other species and thus is of limited translational utility.

 

 

Comparison between an exclusive in vitro-produced embryo transfer system and artificial insemination for genetic, technical, and financial herd performance.

Author information: Kaniyamattam K1, Block J1, Hansen PJ1, De Vries A2.

1Department of Animal Sciences, University of Florida, Gainesville 32611.
2Department of Animal Sciences, University of Florida, Gainesville 32611. Electronic address: devries@ufl.edu.
Journal: Journal of Dairy Science

Date of e-pub: April 2017

Abstract: The objective of this study was to implement an in vitro produced embryo transfer (IVP-ET) system in an existing stochastic dynamic dairy simulation model with multitrait genetics to evaluate the genetic, technical, and financial performance of a dairy herd implementing an exclusive IVP-ET or artificial insemination (AI) system. In the AI system, sexed semen was used on the genetically best heifers only. In the IVP-ET system, all of the animals in the herd were impregnated with female sexed embryos created through in vitro fertilization of oocytes collected from animals of superior genetics for different traits of interest. Each donor was assumed to yield on average 4.25 transferable embryos per collection. The remaining animals in the herd were used as recipients and received either a fresh embryo or a frozen embryo when fresh embryos were not available. Selection of donors was random or based on the greatest estimated breeding value (EBV) of lifetime net merit (NM$), milk yield, or daughter pregnancy rate. For both the IVP-ET and AI systems, culling of surplus heifer calves not needed to replace culled cows was based on the lowest EBV for the same traits. A herd of 1,000 milking cows was simulated 15 yr over time after the start of the IVP-ET system. The default cost to produce and transfer 1 embryo was set at $165. Prices of fresh embryos at which an exclusive IVP-ET system financially breaks even with the comparable AI system in yr 15 and for an investment period of 15 yr were also estimated. More surplus heifer calves were sold from the IVP-ET systems than from the comparable AI systems. The surplus calves from the IVP-ET systems were also genetically superior to the surplus calves from the comparable AI systems, which might be reflected in their market value as a premium price. The most profitable scenario among the 4 IVP-ET scenarios in yr 15 was the one in which NM$ was maximized in the herd. This scenario had an additional profit of $8/cow compared with a similar AI scenario that maximized NM$, provided that surplus heifer calves could be sold at a premium price based on the superiority of the EBV of NM$. For the IVP-ET system to be at least as profitable as the comparable AI system during a 15-yr investment period, the surplus calves from the IVP-ET system needed to be sold at the premium prices. The break-even price of fresh embryos was estimated to be $84 for the exclusive IVP-ET system. This resulted in the same profit as the AI system, which maximized NM$ for a 15-yr investment period and in which heifer calves were sold at a premium price.

 

 

Postnatal phenotype of dairy cows is altered by in vitro embryo production using reverse X-sorted semen.

Author information: Siqueira LGB1, Dikmen S2, Ortega MS3, Hansen PJ4.

1Department of Animal Sciences, D.H. Barron Reproductive and Perinatal Biology Research Program, and Genetics Institute, University of Florida, Gainesville 32611-0910; Embrapa Gado de Leite, Juiz de Fora, MG, Brazil 36038-330.
2Department of Animal Sciences, Faculty of Veterinary Medicine, Uludag University, Bursa, 16059, Turkey.
3Department of Animal Sciences, D.H. Barron Reproductive and Perinatal Biology Research Program, and Genetics Institute, University of Florida, Gainesville 32611-0910.
4Department of Animal Sciences, D.H. Barron Reproductive and Perinatal Biology Research Program, and Genetics Institute, University of Florida, Gainesville 32611-0910. Electronic address: hansen@animal.ufl.edu.
Journal: Journal of Dairy Science

Date of e-pub: April 2017

Abstract: Abnormal fetuses, neonates, and adult offspring derived by assisted reproductive technologies have been reported in humans and mice and have been associated with increased likelihood of certain adult diseases. To test the hypothesis that bovine females derived by assisted reproductive technologies have altered postnatal growth and adult function, a retrospective cohort study evaluated survival, growth, and production traits of offspring derived by in vitro embryo production (IVP) with conventional (IVP-conv) or reverse X-sorted semen (IVP-sexed), multiple ovulation and embryo transfer, and artificial insemination (AI) in a large dairy herd. Live calves produced by IVP were born slightly heavier compared with AI calves. In addition, IVP-sexed calves had a higher cumulative mortality from 90 to 180 d of age compared with AI offspring. Mortality of IVP-conv and multiple ovulation and embryo transfer offspring was intermediate and not different from AI or IVP-sexed offspring. The altered phenotype of offspring from IVP-sexed extended to adult milk production. Cows derived by IVP-sexed produced less milk, fat, and protein in their first lactation compared with dairy cows derived by AI. Additionally, females born to nulliparous dams had a distinct postnatal phenotype compared with offspring from parous dams even when data were restricted to offspring of surrogate females. In conclusion, procedures associated with in vitro production of embryos involving use of reverse-sorted spermatozoa for fertilization result in an alteration of embryonic programming that persists postnatally and causes an effect on milk production in adulthood. Thus, some benefits of reverse-sorted semen for genetic improvement may be offset by adverse programming events.

 

 

Recent Advances in Nanomaterials for Gene Delivery-A Review.

Author information: Riley MK1,2, Vermerris W3,4,5.

1Graduate Program in Plant Cellular and Molecular Biology, University of Florida, Gainesville, FL 32611, USA. mike.riley2@ufl.edu.
2UF Genetics Institute, University of Florida, Gainesville, FL 32611, USA. mike.riley2@ufl.edu.
3Graduate Program in Plant Cellular and Molecular Biology, University of Florida, Gainesville, FL 32611, USA. wev@ufl.edu.
4UF Genetics Institute, University of Florida, Gainesville, FL 32611, USA. wev@ufl.edu.
5Department of Microbiology & Cell Science, University of Florida, Cancer/Genetics Research Complex 302, 2033 Mowry Road, Gainesville, FL 32610, USA. wev@ufl.edu.
Journal: Nanomaterials (Basel, Switzerland)

Date of e-pub: April 2017

Abstract: With the rapid development of nanotechnology in the recent decade, novel DNA and RNA delivery systems for gene therapy have become available that can be used instead of viral vectors. These non-viral vectors can be made of a variety of materials, including inorganic nanoparticles, carbon nanotubes, liposomes, protein and peptide-based nanoparticles, as well as nanoscale polymeric materials. They have as advantages over viral vectors a decreased immune response, and additionally offer flexibility in design, allowing them to be functionalized and targeted to specific sites in a biological system with low cytotoxicity. The focus of this review is to provide an overview of novel nanotechnology-based methods to deliver DNA and small interfering RNAs into biological systems.

 

 

Complete Genome Sequences of Identical Zika virus Isolates in a Nursing Mother and Her Infant.

Author information: Blohm GM1,2, Lednicky JA3,4, Márquez M2,5, White SK3,4, Loeb JC4, Pacheco CA6, Nolan DJ3,7, Paisie T3,7, Salemi M3,7, Rodríguez-Morales AJ8, Morris JG Jr3,9, Pulliam JRC1,3,10, Carrillo AS11, Plaza JD11, Paniz-Mondolfi AE12.

1Department of Biology, College of Liberal Arts and Sciences, University of Florida, Gainesville, Florida, USA.
2Department of Pathology and Laboratory Medicine, Hospital Internacional Barquisimeto, Lara, Barquisimeto, Venezuela.
3Emerging Pathogens Institute, University of Florida, Gainesville, Florida, USA.
4Department of Environmental and Global Health, College of Public Health and Health Professions, University of Florida, Gainesville, Florida, USA.
5Health Sciences Department, College of Medicine, Universidad Centroccidental Lisandro Alvarado, Barquisimeto, Lara, Venezuela.
6Policlínica Barquisimeto, Barquisimeto, Lara, Venezuela.
7Department of Pathology, Immunology, and Laboratory Medicine, College of Medicine, University of Florida, Gainesville, Florida, USA.
8Public Health and Infection Research Group, Faculty of Health Sciences, Universidad Tecnológica de Pereira, Pereira, Colombia.
9Division of Infectious Diseases and Global Health, Department of Medicine, College of Medicine, University of Florida, Gainesville, Florida, USA.
10DST-NRF Centre of Excellence in Epidemiological Modelling and Analysis (SACEMA), Matieland, South Africa.
11Health Sciences Department, College of Medicine, Universidad Nacional Experimental Francisco de Miranda, Punto Fijo, Falcon, Venezuela.
12Department of Pathology and Laboratory Medicine, Hospital Internacional Barquisimeto, Lara, Barquisimeto, Venezuela albertopaniz@yahoo.com.
Journal: Genome Announcements

Date of e-pub: April 2017

Abstract: Complete genome sequences were obtained for Zika viruses isolated from the breast milk of a Venezuelan patient and her child, who was exclusively breastfeeding at the time. These sequences are the first to be reported from a presumptive autochthonous postnatal transmission case from mother to child in Venezuela.

 

 

Quercetin, a natural product supplement, impairs mitochondrial bioenergetics and locomotor behavior in larval zebrafish (Danio rerio).

Author information: Zhang JL1, Laurence Souders C 2nd2, Denslow ND2, Martyniuk CJ3.

1Henan Open Laboratory of Key Subjects of Environmental and Animal Products Safety, College of Animal Science and Technology, Henan University of Science and Technology, Henan, China; Center for Environmental and Human Toxicology, Department of Physiological Sciences, College of Veterinary Medicine, UF Genetics Institute, University of Florida, Gainesville, FL 32611, USA. Electronic address: jlzhang@haust.edu.cn.
2Center for Environmental and Human Toxicology, Department of Physiological Sciences, College of Veterinary Medicine, UF Genetics Institute, University of Florida, Gainesville, FL 32611, USA.
3Center for Environmental and Human Toxicology, Department of Physiological Sciences, College of Veterinary Medicine, UF Genetics Institute, University of Florida, Gainesville, FL 32611, USA. Electronic address: cmartyn@ufl.edu.
Journal: Toxicology and Applied Pharmacology

Date of e-pub: April 2017

Abstract: Quercetin is a natural product that is sold as a supplement in health food stores. While there are reported benefits for this flavonoid as a dietary supplement due to antioxidant properties, the full scope of its biological interactions has not been fully addressed. To learn more about the mechanisms of action related to quercetin, we exposed zebrafish (Danio rerio) embryos to 1 and 10μg/L quercetin for 96h starting at 3h post fertilization. Quercetin up to 10μg/L did not induce significant mortality in developing fish, but did increase prevalence of an upward-curved dorsal plane in hatched larvae. To determine whether this developmental defect was potentially related to mitochondrial bioenergetics during development, we measured oxygen consumption rate in whole embryos following a 24-hour exposure to quercetin. Basal mitochondrial and ATP-linked respiration were decreased at 1 and 10μg/L quercetin, and maximal respiration was decreased at 10μg/L quercetin, suggesting that quercetin impairs mitochondrial bioenergetics. This is proposed to be related to the deformities observed during development. Due to the fact that ATP production was affected by quercetin, larval behaviors related to locomotion were investigated, as well as transcriptional responses of six myogenesis transcripts. Quercetin at 10μg/L significantly reduced the swimming velocity of zebrafish larvae. The expression levels of both myostatin A (mstna) and myogenic differentiation (myoD) were also altered by quercetin. Mstna, an inhibitory factor for myogenesis, was significantly increased at 1μg/L quercetin exposure, while myoD, a stimulatory factor for myogenesis, was significantly increased at 10μg/L quercetin exposure. There were no changes in transcripts related to apoptosis (bcl2, bax, casp3, casp7), but we did observe a decrease in mRNA levels for catalase (cat) in fish exposed to each dose, supporting an oxidative stress response. Our data support the hypothesis that quercetin may affect locomotion and induce deformities in zebrafish larvae by diminishing ATP production and by altering the expression of transcripts related to muscle formation and activity.

 

 

Role of CC Cytokines in Spatial Arrangement of the Inner Cell Mass of the Bovine Embryo.

Author information: Negrón-Pérez VM1, Vargas-Franco D2, Hansen PJ1.

1Department of Animal Sciences, D. H. Barron Reproductive and Perinatal Biology Research Program and Genetics Institute, University of Florida, Gainesville, Florida, USA.
2Department of Molecular Genetics and Microbiology, Center for Epigenetics and Genetics Institute, University of Florida, Gainesville, Florida, USA.
Journal: Biology of Reproduction

Date of e-pub: April 2017

Abstract: The process of spatial rearrangement of cells of the inner cell mass (ICM) that are destined to become hypoblast is not well understood. The observation that the chemokine CCL24 and several other genes involved in chemokine signaling are expressed more in the ICM than the trophectoderm of the bovine embryo resulted in the hypothesis that CCL24 participates in spatial organization of the ICM. Temporally, expression of CCL24 in the bovine embryo occurs coincident with blastocyst formation: transcript abundance was low until the late morula stage, peaked in the blastocyst at Day 7 of development and declined by Day 9. Treatment of embryos with two separate antagonists of CCR3 (the prototypical receptor for CCL24) decreased the percent of GATA6+ cells (hypoblast precursors) that were located in the outside of the ICM. Similarly, injection of zygotes with a CCL24-specific morpholino decreased the percent of GATA6+ cells in the outside of the ICM. In conclusion, CCL24 assists in spatial arrangement of the ICM in the bovine embryo. This experiment points to new functions of chemokine signaling in the bovine embryo and is consistent with the idea that cell migration is involved in the spatial organization of hypoblast cells in the blastocyst.

 

 

Insights into the evolution of hydroxyproline rich glycoproteins from 1000 plant transcriptomes.

Author information: Johnson KL1, Cassin AM2, Lonsdale A3, Wong GK4, Soltis D5, Miles NW6, Melkonian M7, Melkonian B7, Deyholos MK8, Leebens-Mack J9, Rothfels CJ10, Stevenson DW11, Graham SW12, Wang X13, Wu S13, Pires JC14, Edger PP15, Carpenter EJ16, Bacic A17, Doblin MS18, Schultz CJ19.

1University of Melbourne CITY: Melbourne STATE: Vic POSTAL_CODE: 3010 Australia [AU].
2University of Melbourne CITY: Melbourne STATE: VIC Australia [AU].
3University of Melbourne CITY: Melbourne STATE: Vic Australia [AU].
4University of Alberta University of Alberta, CW405 Biological Sciences CITY: Edmonton STATE: AB POSTAL_CODE: T6G 2E9 Canada [CA].
5University of Florida Dept Biology Univ Florida CITY: Gainesville POSTAL_CODE: 32605 United States Of America [US].
6University of Florida CITY: Gainesville United States Of America [US].
7Universität zu Köln CITY: Köln Germany [DE].
8University of British Columbia, Okanagan SCI 316, 1177 Research Road, UBCO CITY: Kelowna STATE: BC POSTAL_CODE: V1V 1V7 Canada [CA].
9Univ Georgia CITY: N/A POSTAL_CODE: N/A United States Of America [US].
10University of California, Berkeley CITY: Berkeley STATE: California United States Of America [US].
11New York Botanical Garden CITY: New York STATE: NY United States Of America [US].
12University of British Columbia CITY: Vancouver Canada [CA].
13Chinese Academy of Sciences CITY: Beijing China [CN].
14University of Missouri CITY: Columbia STATE: Missouri POSTAL_CODE: 65211 United States Of America [US].
15Michigan State University CITY: East Lansing STATE: MI United States Of America [US].
16University of Alberta CITY: Edmonton STATE: Alberta Canada [CA].
17The University of Melbourne CITY: Victoria POSTAL_CODE: 3010 Australia [AU].
18University of Melbourne CITY: Parkville STATE: Victoria POSTAL_CODE: 3010 Australia [AU].
19The University of Adelaide Waite Agricultural Research Institute, RMB1 CITY: Glen Osmond STATE: SA POSTAL_CODE: 5064 Australia [AU] carolyn.schultz@adelaide.edu.au.
Journal: Plant Physiology

Date of e-pub: April 2017

Abstract: The carbohydrate-rich cell walls of land plants and algae have been the focus of much interest given the value of cell wall based products to our current and future economies. Hydroxyproline-rich glycoproteins (HRGPs), a major group of wall glycoproteins, play important roles in plant growth and development, yet little is known about how they have evolved in parallel with the polysaccharide components of walls. We investigate the origins and evolution of the HRGP superfamily, which is commonly divided into three major multigene families: the arabinogalactan-proteins (AGPs), extensins (EXTs) and proline-rich proteins (PRPs). Using MAAB, a newly developed bioinformatics pipeline, we identified HRGPs in sequences from the 1000 plants (1KP) transcriptome project (www.onekp.com). Our analyses provide new insights into the evolution of HRGPs across major evolutionary milestones, including the transition to land and early radiation of angiosperms. Significantly, data mining reveals the origin of GPI-anchored AGPs in green algae and a 3-4 fold increase in GPI-AGPs in liverworts and mosses. The first detection of cross-linking (CL)-EXTs is observed in bryophytes, which suggest that CL-EXTs arose though the juxtaposition of pre-existing SPn EXT glycomotifs with refined Y-based motifs. We also detected the loss of CL-EXT in a few lineages, including the grass family (Poaceae), that have a cell wall composition distinct from other monocots and eudicots. A key challenge in HRGP research is tracking individual HRGPs throughout evolution. Using the 1KP output we were able to find putative orthologs of Arabidopsis pollen-specific GPI-AGPs in basal eudicots.

 

 

Whole genome sequencing reveals a novel deletion variant in the KIT gene in horses with white spotted coat colour phenotypes.

Author information: Dürig N1,2, Jude R1,2,3, Holl H4,5, Brooks SA4, Lafayette C5, Jagannathan V1,2, Leeb T1,2.

1Vetsuisse Faculty, Institute of Genetics, University of Bern, 3001, Bern, Switzerland.
2DermFocus, University of Bern, 3001, Bern, Switzerland.
3RJC, 53919, Weilerswist, Germany.
4Department of Animal Sciences, University of Florida, Gainesville, FL, 32611-0910, USA.
5Etalon Inc., Menlo Park, CA, 94025, USA.
Journal: Animal Genetics

Date of e-pub: April 2017

Abstract: White spotting phenotypes in horses can range in severity from the common white markings up to completely white horses. EDNRB, KIT, MITF, PAX3 and TRPM1 represent known candidate genes for such phenotypes in horses. For the present study, we re-investigated a large horse family segregating a variable white spotting phenotype, for which conventional Sanger sequencing of the candidate genes’ individual exons had failed to reveal the causative variant. We obtained whole genome sequence data from an affected horse and specifically searched for structural variants in the known candidate genes. This analysis revealed a heterozygous ~1.9-kb deletion spanning exons 10-13 of the KIT gene (chr3:77,740,239_77,742,136del1898insTATAT). In continuity with previously named equine KIT variants we propose to designate the newly identified deletion variant W22. We had access to 21 horses carrying the W22 allele. Four of them were compound heterozygous W20/W22 and had a completely white phenotype. Our data suggest that W22 represents a true null allele of the KIT gene, whereas the previously identified W20 leads to a partial loss of function. These findings will enable more precise genetic testing for depigmentation phenotypes in horses.

 

 

Tet2 loss leads to hypermutagenicity in haematopoietic stem/progenitor cells.

Author information: Pan F1,2,3, Wingo TS4,5,6, Zhao Z7, Gao R1, Makishima H8, Qu G1,2, Lin L4, Yu M9, Ortega JR10, Wang J2, Nazha A8, Chen L4, Yao B4, Liu C1, Chen S1, Weeks O3, Ni H11, Phillips BL12, Huang S13, Wang J14, He C9, Li GM10, Radivoyevitch T8, Aifantis I15,16, Maciejewski JP8, Yang FC1,2, Jin P4, Xu M1,2.

1Department of Biochemistry and Molecular Biology, Sylvester Comprehensive Cancer Center, University of Miami Miller School of Medicine, 1011 NW 15th Street, Room 411, Gautier Building, MC R629, Miami, Florida 33136, USA.
2Department of Pediatrics, Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, Indiana 46202, USA.
3Department of Biological Sciences, Florida International University, Miami, Florida 33199, USA.
4Departments of Human Genetics, Emory University School of Medicine, Atlanta, Georgia 30307, USA.
5Neurology, Emory University School of Medicine, Atlanta, Georgia 30307, USA.
6Division of Neurology, Department of Veterans Affairs Medical Center, Atlanta, Georgia 30033, USA.
7Department of Hematology and Oncology, Tianjin Medical University Cancer Institute and Hospital, National Clinical Research Center for Cancer, Key Laboratory of Cancer Prevention and Therapy, Tianjin 300060, China.
8Department of Translational Hematology and Oncology Research, Taussig Cancer Institute, Cleveland Clinic, Cleveland, Ohio 44195, USA.
9Department of Chemistry and Institute for Biophysical Dynamics, University of Chicago, Chicago, Illinois 60637, USA.
10Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, California 90033, USA.
11Department of Pathology, University of Illinois at Chicago, Chicago, Illinois 60607, USA.
12Department of Biochemistry and Pharmacology, Emory University School of Medicine, Atlanta, Georgia 30307, USA.
13Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, Florida 32611, USA.
14Department of Developmental and Regenerative Biology, Black Family Stem Cell Institute, Icahn School of Medicine at Mount Sinai, New York, New York 10029, USA.
7School of Medicine, Saint Louis University, St. Louis, MO 63104, USA. Electronic address: mackieln@slu.edu.
15Howard Hughes Medical Institute and Department of Pathology, NYU School of Medicine, New York, New York 10016, USA.
16NYU Cancer Institute and Helen L. and Martin S. Kimmel Center for Stem Cell Biology, NYU School of Medicine, New York, New York 10016, USA.
Journal: Nature Communications

Date of e-pub: April 2017

Abstract: TET2 is a dioxygenase that catalyses multiple steps of 5-methylcytosine oxidation. Although TET2 mutations frequently occur in various types of haematological malignancies, the mechanism by which they increase risk for these cancers remains poorly understood. Here we show that Tet2-/- mice develop spontaneous myeloid, T- and B-cell malignancies after long latencies. Exome sequencing of Tet2-/- tumours reveals accumulation of numerous mutations, including Apc, Nf1, Flt3, Cbl, Notch1 and Mll2, which are recurrently deleted/mutated in human haematological malignancies. Single-cell-targeted sequencing of wild-type and premalignant Tet2-/- Lin-c-Kit+ cells shows higher mutation frequencies in Tet2-/- cells. We further show that the increased mutational burden is particularly high at genomic sites that gained 5-hydroxymethylcytosine, where TET2 normally binds. Furthermore, TET2-mutated myeloid malignancy patients have significantly more mutational events than patients with wild-type TET2. Thus, Tet2 loss leads to hypermutagenicity in haematopoietic stem/progenitor cells, suggesting a novel TET2 loss-mediated mechanism of haematological malignancy pathogenesis.

 

 

ESPRIT-Forest: Parallel clustering of massive amplicon sequence data in subquadratic time.

Author information: Cai Y1, Zheng W2, Yao J3, Yang Y1, Mai V4, Mao Q3, Sun Y2,3,5.

1Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China.
2Department of Computer Science and Engineering, The State University of New York at Buffalo, Buffalo, New York, United States of America.
3Department of Microbiology and Immunology, The State University of New York at Buffalo, Buffalo, New York, United States of America.
4Department of Epidemiology, University of Florida, Gainesville, Florida, United States of America.
5Department of Biostatistics, The State University of New York at Buffalo, Buffalo, New York, United States of America.
Journal: PLoS Computational Biology

Date of e-pub: April 2017

Abstract: The rapid development of sequencing technology has led to an explosive accumulation of genomic sequence data. Clustering is often the first step to perform in sequence analysis, and hierarchical clustering is one of the most commonly used approaches for this purpose. However, it is currently computationally expensive to perform hierarchical clustering of extremely large sequence datasets due to its quadratic time and space complexities. In this paper we developed a new algorithm called ESPRIT-Forest for parallel hierarchical clustering of sequences. The algorithm achieves subquadratic time and space complexity and maintains a high clustering accuracy comparable to the standard method. The basic idea is to organize sequences into a pseudo-metric based partitioning tree for sub-linear time searching of nearest neighbors, and then use a new multiple-pair merging criterion to construct clusters in parallel using multiple threads. The new algorithm was tested on the human microbiome project (HMP) dataset, currently one of the largest published microbial 16S rRNA sequence dataset. Our experiment demonstrated that with the power of parallel computing it is now compu- tationally feasible to perform hierarchical clustering analysis of tens of millions of sequences. The software is available at http://www.acsu.buffalo.edu/∼yijunsun/lab/ESPRIT-Forest.html.

 

 

Insulitis in Autoantibody-Positive Pancreatic Donor With History of Gestational Diabetes Mellitus.

Author information: Jackson J1,2, Posgai A1, Campbell-Thompson M1, Kusmartseva I3.

1Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, FL.
2Division of Maternal & Fetal Medicine, Department of Obstetrics & Gynecology, University of Florida, Gainesville, FL.
3Department of Pathology, Immunology and Laboratory Medicine, University of Florida, Gainesville, FL inkusmartseva@ufl.edu.
Journal: Diabetes Care

Date of e-pub: May 2017

Abstract: N/A

 

 

Disulfide bond disrupting agents activate the unfolded protein response in EGFR- and HER2-positive breast tumor cells.

Author information: Ferreira RB1, Wang M2,3, Law ME2,3, Davis BJ2,3, Bartley AN1, Higgins PJ4, Kilberg MS5, Santostefano KE6, Terada N6, Heldermon CD7, Castellano RK1, Law BK2,3.

1Department of Chemistry, University of Florida, Gainesville, FL 32611, USA.
2Department of Pharmacology & Therapeutics, University of Florida, Gainesville, FL 32610, USA.
3UF-Health Cancer Center, University of Florida, Gainesville, FL 32610, USA.
4Center for Cell Biology and Cancer Research, Albany Medical College, Albany, NY 12208, USA.
5Department of Biochemistry, University of Florida, Gainesville, FL, 32610, USA.
6Department of Pathology, Immunology, and Laboratory Medicine, Center for Cellular Reprogramming, University of Florida College of Medicine, Gainesville, FL 32610, USA.
7Department of Medicine, University of Florida, Gainesville, FL, 32610, USA.
Journal: Oncotarget

Date of e-pub: April 2017

Abstract: Many breast cancer deaths result from tumors acquiring resistance to available therapies. Thus, new therapeutic agents are needed for targeting drug-resistant breast cancers. Drug-refractory breast cancers include HER2+ tumors that have acquired resistance to HER2-targeted antibodies and kinase inhibitors, and “Triple-Negative” Breast Cancers (TNBCs) that lack the therapeutic targets Estrogen Receptor, Progesterone Receptor, and HER2. A significant fraction of TNBCs overexpress the HER2 family member Epidermal Growth Factor Receptor (EGFR). Thus agents that selectively kill EGFR+ and HER2+ tumors would provide new options for breast cancer therapy. We previously identified a class of compounds we termed Disulfide bond Disrupting Agents (DDAs) that selectively kill EGFR+ and HER2+ breast cancer cells in vitro and blocked the growth of HER2+ breast tumors in an animal model. DDA-dependent cytotoxicity was found to correlate with downregulation of HER1-3 and Akt dephosphorylation. Here we demonstrate that DDAs activate the Unfolded Protein Response (UPR) and that this plays a role in their ability to kill EGFR+ and HER2+ cancer cells. The use of breast cancer cell lines ectopically expressing EGFR or HER2 and pharmacological probes of UPR revealed all three DDA responses: HER1-3 downregulation, Akt dephosphorylation, and UPR activation, contribute to DDA-mediated cytotoxicity. Significantly, EGFR overexpression potentiates each of these responses. Combination studies with DDAs suggest that they may be complementary with EGFR/HER2-specific receptor tyrosine kinase inhibitors and mTORC1 inhibitors to overcome drug resistance.

 

 

LPS Stimulation of Cord Blood Reveals a Newborn-Specific Neutrophil Transcriptomic Response and Cytokine Production.

Author information: Mathias B1, Mira JC, Rehfuss JP, Rincon JC, Ungaro R, Nacionales DC, Lopez MC, Baker HV, Moldawer LL, Larson SD.

1Department of Surgery, University of Florida College of Medicine, Gainesville, Florida†Department of Molecular Genetics and Microbiology, University of Florida College of Medicine, Gainesville, Florida.
Journal: Shock (Augusta, Ga.)

Date of e-pub: May 2017

Abstract: The neonatal innate immune system differs to microbial infection both quantitatively and qualitatively when compared with adults. Here, we provide the first genome-wide ex-vivo expression profile of umbilical cord blood (UCB) neutrophils from full-term infants prior to and in response to whole-blood lipopolysaccharide (LPS) stimulation. Additionally, we provide cytokine expression prior to and following LPS stimulation. The genomic expression and cytokine profile are compared with LPS-stimulated whole blood from healthy adult subjects (HC).

Whole blood from UCB (n = 6) and HC (n = 6) was studied at baseline or was stimulated for 24 h with 100 ngs/mL of LPS. CD66b neutrophils were subsequently isolated with microfluidic techniques and genome-wide expression analyses were performed. Ingenuity Pathway Analysis (IPA) software was utilized to predict downstream functional effects. Additionally, cytokine concentrations in whole blood prior to and after 24 h of LPS incubation were determined.

LPS stimulated whole blood from UCB demonstrated significant differences in both ex-vivo cytokine production and PMN gene expression. Mixed-effect modeling identified 1,153 genes whose expression changed significantly in UCB and HC after exposure to LPS (P < 0.001 with a minimum 1.5-fold change). IPA downstream predictions suggest that PMNs from UCB fail to effectively upregulate genes associated with activation, phagocytosis, and chemotaxis in response to LPS stimulation. Furthermore, whole blood from UCB showed increased interleukin (IL)-10 production to LPS, but failed to significantly increase several pro-inflammatory cytokines.

LPS-stimulated whole blood from UCB exhibited a markedly suppressed inflammatory cytokine production and PMN innate immune genome response. These differences in gene expression and cytokine production may be an adaptive response to a prior fetal environment, but may also explain their increased susceptibility to infections. Characterization of these deficits is the first step toward developing prophylactic and therapeutic interventions.

 

 

Semipermeable Functional DNA-Encapsulated Nanocapsules as Protective Bioreactors for Biosensing in Living Cells.

Author information: Hong CY1,2, Wu SX1, Li SH1, Liang H1, Chen S1, Li J1,3, Yang HH1, Tan W3,2.

1MOE Key Laboratory for Analytical Science of Food Safety and Biology, Fujian Provincial Key Laboratory of Analysis and Detection Technology for Food Safety, State Key Laboratory of Photocatalysis on Energy and Environment, College of Chemistry, Fuzhou University , Fuzhou 350002, China.
2Department of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, UF Health Cancer Center, University of Florida , Gainesville, Florida 32611-7200, United States.
3Molecular Sciences and Biomedicine Laboratory, State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering and College of Biology, Collaborative Innovation Center for Molecular Engineering and Theranostics, Hunan University , Changsha 410082, China.
Journal: Analytical Chemistry

Date of e-pub: May 2017

Abstract: The development of functional DNA-based nanosensors in living cells has experienced some design challenges, including, for example, poor cellular uptake, rapid nuclease degradation, and high false positives. Herein, we designed selectively permeable poly(methacrylic acid) (PMA) nanocapsules to encapsulate functional DNAs for metal ions and small-molecules sensing in living cells. Since functional DNAs are concentrated in the nanocapsules, an increasing reaction rate is obtained in vitro. During endocytosis, polymeric capsules simultaneously improve cellular uptake of functional DNAs and preserve their structural integrity inside the confined capsule space. More importantly, selective shell permeability allows for the free diffusion of small molecular targets through capsule shells but limits the diffusion of large biomolecules, such as nuclease and nonspecific protein. Compared to the free DNAzyme, PMA nanocapsules could reduce false positives and enhance detection accuracy. Furthermore, PMA nanocapsules are biocompatible and biodegradable. Through the controllability of wall thickness, permeability, and size distribution, these nanocapsules could be expanded easily to other targets, such as microRNAs, small peptides, and metabolites. These nanocapsules will pave the way for in situ monitoring of various biological processes in living cells and in vivo.

 

 

Functional characterization of LotP from Liberibacter asiaticus.

Author information: Loto F1,2, Coyle JF1, Padgett KA1,3, Pagliai FA1, Gardner CL1, Lorca GL1, Gonzalez CF1.

1Department of Microbiology and Cell Science, Genetics Institute, Institute of Food and Agricultural Sciences, University of Florida, 2033 Mowry road, PO Box 103610, Gainesville, FL 32610-3610, USA.
2PROIMI Planta Piloto de Procesos Industriales Microbiológicos, CONICET, Tucumán, Argentina.
3Department of Microbiology and Cell Science, Undergraduate Research Program, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL, USA.
Journal: Microbial Biotechnology

Date of e-pub: May 2017

Abstract: Liberibacter asiaticus is an unculturable parasitic bacterium of the alphaproteobacteria group hosted by both citrus plants and a psyllid insect vector (Diaphorina citri). In the citrus tree, the bacteria thrive only inside the phloem, causing a systemically incurable and deadly plant disease named citrus greening or Huanglongbing. Currently, all commercial citrus cultivars in production are susceptible to L. asiaticus, representing a serious threat to the citrus industry worldwide. The technical inability to isolate and culture L. asiaticus has hindered progress in understanding the biology of this bacterium directly. Consequently, a deep understanding of the biological pathways involved in the regulation of host-pathogen interactions becomes critical to rationally design future and necessary strategies of control. In this work, we used surrogate strains to evaluate the biochemical characteristics and biological significance of CLIBASIA_03135. This gene, highly induced during early stages of plant infection, encodes a 23 kDa protein and was renamed in this work as LotP. This protein belongs to an uncharacterized family of proteins with an overall structure resembling the LON protease N-terminus. Co-immunoprecipitation assays allowed us to identify the Liberibacter chaperonin GroEL as the main LotP-interacting protein. The specific interaction between LotP and GroEL was reconstructed and confirmed using a two-hybrid system in Escherichia coli. Furthermore, it was demonstrated that LotP has a native molecular weight of 44 kDa, corresponding to a dimer in solution with ATPase activity in vitro. In Liberibacter crescens, LotP is strongly induced in response to conditions with high osmolarity but repressed at high temperatures. Electrophoretic mobility shift assay (EMSA) results suggest that LotP is a member of the LdtR regulon and could play an important role in tolerance to osmotic stress.

 

 

Association of the HLA-B*53:01 Allele With Drug Reaction With Eosinophilia and Systemic Symptoms (DRESS) Syndrome During Treatment of HIV Infection With Raltegravir.

Author information: Thomas M1,2, Hopkins C2, Duffy E2, Lee D3, Loulergue P4, Ripamonti D5, Ostrov DA6, Phillips E7,8,9.

1Department of Molecular Medicine and Pathology, Faculty of Medical and Health Sciences, University of Auckland.
2Infectious Diseases Department, Auckland City Hospital, New Zealand.
3Department of Internal Medicine, University of California San Diego.
4Centre d’Investigation Clinique Cochin-Pasteur de Vaccinologie Cochin-Pasteur, Hôpital Cochin, Paris, France.
5Department of Infectious Diseases, Ospedale Papa Giovanni XXIII, Bergamo, Italy.
6Department of Pathology, Immunology and Laboratory Medicine, University of Florida College of Medicine, Gainesville.
7Oates Institute for Experimental Therapeutics.
8Department of Medicine, Vanderbilt University, Nashville Tennessee.
9Institute for Immunology and Infectious Diseases, Murdoch University, Murdoch, Western Australia.
Journal: Clinical Infectious Diseases : An Official Publication of the Infectious Diseases Society of America

Date of e-pub: May 2017

Abstract: Drug reaction with eosinophilia and systemic symptoms (DRESS) syndrome is a rare, severe adverse event during treatment with raltegravir. The occurrence of DRESS syndrome during treatment with other drugs is strongly associated with particular HLA alleles.

We performed HLA testing in 3 of the 5 patients previously reported to have developed raltegravir-induced DRESS syndrome and in 1 previously unreported patient. We then used virtual modeling to visualize interactions between raltegravir and the imputed HLA molecule.

Five of the 6 patients who developed raltegravir-induced DRESS syndrome were African, and 1 was Hispanic. HLA typing was performed in 4 patients, all of whom carried both the HLA-B*53 allele and the HLA-C*04 allele to which it is commonly haplotypic. No other HLA alleles were shared by all of the tested patients. Given the approximate prevalence of HLA-B*53 carriage in African (20%) and Hispanic (6%) populations, the probability of all 4 patients being HLA-B*53 carriers, and 2 of 3 African patients being homozygous for HLA-B*53:01, is approximately 0.00002.

These data implicate the prevalent African allele HLA-B*53:01 in the immunopathogenesis of raltegravir-induced DRESS syndrome. Although the immunopathogenic mechanisms are currently unknown, virtual modeling suggests that raltegravir may bind within the antigen binding cleft of the HLA-B*53:01 molecule, but not within the closely related HLA-B*35:01 molecule. Further studies are necessary to confirm the strength of the association between carriage of the HLA-B*53:01 allele and raltegravir-induced DRESS syndrome, and the potential utility of HLA screening before raltegravir treatment.

 

 

Dynamic Gene Regulatory Networks of Human Myeloid Differentiation.

Author information: Ramirez RN1, El-Ali NC1, Mager MA1, Wyman D2, Conesa A3, Mortazavi A4.

1Developmental and Cell Biology, University of California Irvine, Irvine, CA 92697-2300, USA; Center for Complex Biological Systems, University of California Irvine, Irvine, CA 92697-2300, USA.
2Center for Complex Biological Systems, University of California Irvine, Irvine, CA 92697-2300, USA.
3Centro de Investigacion Principe Felipe, 46012 Valencia, Spain; Microbiology and Cell Science, University of Florida, Gainesville, FL 32603, USA.
4Developmental and Cell Biology, University of California Irvine, Irvine, CA 92697-2300, USA; Center for Complex Biological Systems, University of California Irvine, Irvine, CA 92697-2300, USA. Electronic address: ali.mortazavi@uci.edu.
Journal: Cell Systems

Date of e-pub: April 2017

Abstract: The reconstruction of gene regulatory networks underlying cell differentiation from high-throughput gene expression and chromatin data remains a challenge. Here, we derive dynamic gene regulatory networks for human myeloid differentiation using a 5-day time series of RNA-seq and ATAC-seq data. We profile HL-60 promyelocytes differentiating into macrophages, neutrophils, monocytes, and monocyte-derived macrophages. We find a rapid response in the expression of key transcription factors and lineage markers that only regulate a subset of their targets at a given time, which is followed by chromatin accessibility changes that occur later along with further gene expression changes. We observe differences between promyelocyte- and monocyte-derived macrophages at both the transcriptional and chromatin landscape level, despite using the same differentiation stimulus, which suggest that the path taken by cells in the differentiation landscape defines their end cell state. More generally, our approach of combining neighboring time points and replicates to achieve greater sequencing depth can efficiently infer footprint-based regulatory networks from long series data.

 

 

Pharmacogenetic Associations of β1-Adrenergic Receptor Polymorphisms With Cardiovascular Outcomes in the SPS3 Trial (Secondary Prevention of Small Subcortical Strokes).

Author information: Magvanjav O1, McDonough CW1, Gong Y1, McClure LA1, Talbert RL1, Horenstein RB1, Shuldiner AR1, Benavente OR1, Mitchell BD1, Johnson JA2; NINDS SiGN (Stroke Genetics Network).

1From the Department of Pharmacotherapy and Translational Research, Center for Pharmacogenomics, College of Pharmacy, University of Florida, Gainesville (O.M., C.W.M., Y.G., J.A.J.); Department of Epidemiology and Biostatistics, Dornsife School of Public Health, Drexel University, Philadelphia, PA (L.A.M.); College of Pharmacy, University of Texas, Austin (R.L.T.); Division of Endocrinology, Diabetes and Nutrition and Program for Personalized and Genomic Medicine, University of Maryland School of Medicine, Baltimore (R.B.H., A.R.S., B.D.M.); Department of Neurology, University of British Columbia, Vancouver, Canada (O.R.B.); and Geriatrics Research and Education Clinical Center, Baltimore Veterans Administration Medical Center, MD (B.D.M.).
2From the Department of Pharmacotherapy and Translational Research, Center for Pharmacogenomics, College of Pharmacy, University of Florida, Gainesville (O.M., C.W.M., Y.G., J.A.J.); Department of Epidemiology and Biostatistics, Dornsife School of Public Health, Drexel University, Philadelphia, PA (L.A.M.); College of Pharmacy, University of Texas, Austin (R.L.T.); Division of Endocrinology, Diabetes and Nutrition and Program for Personalized and Genomic Medicine, University of Maryland School of Medicine, Baltimore (R.B.H., A.R.S., B.D.M.); Department of Neurology, University of British Columbia, Vancouver, Canada (O.R.B.); and Geriatrics Research and Education Clinical Center, Baltimore Veterans Administration Medical Center, MD (B.D.M.). julie.johnson@ufl.edu.
Journal: Stroke

Date of e-pub: May 2017

Abstract: Functional polymorphisms (Ser49Gly and Arg389Gly) in ADRB1 have been associated with cardiovascular and β-blocker response outcomes. Herein we examined associations of these polymorphisms with major adverse cardiovascular events (MACE), with and without stratification by β-blocker treatment in patients with a history of stroke.

Nine hundred and twenty-six participants of the SPS3 trial’s (Secondary Prevention of Small Subcortical Strokes) genetic substudy with hypertension were included. MACE included stroke, myocardial infarction, and all-cause death. Kaplan-Meier and multivariable Cox regression analyses were used. Because the primary component of MACE was ischemic stroke, we tested the association of Ser49Gly with ischemic stroke among 41 475 individuals of European and African ancestry in the NINDS (National Institute of Neurological Disorders and Stroke) SiGN (Stroke Genetics Network).

MACE was higher in carriers of the Gly49 allele than in those with the Ser49Ser genotype (10.5% versus 5.4%, log-rank P=0.005). Gly49 carrier status was associated with MACE (hazard ratio, 1.62; 95% confidence interval, 1.00-2.68) and ischemic stroke (hazard ratio, 1.81; 95% confidence interval, 1.01-3.23) in SPS3 and with small artery ischemic stroke (odds ratio, 1.14; 95% confidence interval, 1.03-1.26) in SiGN. In SPS3, β-blocker-treated Gly49 carriers had increased MACE versus non-β-blocker-treated individuals and noncarriers (hazard ratio, 2.03; 95% confidence interval, 1.20-3.45). No associations were observed with the Arg389Gly polymorphism.

Among individuals with previous small artery ischemic stroke, the ADRB1 Gly49 polymorphism was associated with MACE, particularly small artery ischemic stroke, a risk that may be increased among β-blocker-treated individuals. Further research is needed to define β-blocker benefit among ischemic stroke patients by ADRB1 genotype.

 

 

Structure Determination of Mycobacterium tuberculosis Serine Protease Hip1 (Rv2224c).

Author information: Naffin-Olivos JL1, Daab A1,2, White A1, Goldfarb NE3, Milne AC1, Liu D4, Baikovitz J5, Dunn BM6, Rengarajan J7, Petsko GA8, Ringe D1.

1Rosenstiel Basic Medical Sciences Research Center, Brandeis University , Waltham, Massachusetts 02454, United States.
2Department of Biochemistry, Brandeis University , Waltham, Massachusetts 02454, United States.
3Department of Pharmaceutical and Biomedical Sciences, California Health Sciences University , Clovis, California 93612, United States.
4 Department of Chemistry and Biochemistry, Loyola University Chicago , Chicago, Illinois 60660, United States.
5Department of Biology, Brandeis University , Waltham, Massachusetts 02454, United States.
6Department of Biochemistry and Molecular Biology, University of Florida , Gainesville, Florida 32610, United States.
7Division of Infectious Diseases, Department of Medicine, Emory Vaccine Center, Emory University School of Medicine , Atlanta, Georgia 30329, United States.
8Appel Alzheimer’s Disease Research Institute, Weill Cornell Medical College , New York, New York 10021, United States.
Journal: Biochemistry

Date of e-pub: May 2017

Abstract: The Mycobacterium tuberculosis (Mtb) serine protease Hip1 (hydrolase important for pathogenesis; Rv2224c) promotes tuberculosis (TB) pathogenesis by impairing host immune responses through proteolysis of a protein substrate, Mtb GroEL2. The cell surface localization of Hip1 and its immunomodulatory functions make Hip1 a good drug target for new adjunctive immune therapies for TB. Here, we report the crystal structure of Hip1 to a resolution of 2.6 Å and the kinetic studies of the enzyme against model substrates and the protein GroEL2. The structure shows a two-domain protein, one of which contains the catalytic residues that are the signature of a serine protease. Surprisingly, a threonine is located within the active site close enough to hydrogen bond with the catalytic residues Asp463 and His490. Mutation of this residue, Thr466, to alanine established its importance for function. Our studies provide insights into the structure of a member of a novel family of proteases. Knowledge of the Hip1 structure will aid in designing inhibitors that could block Hip1 activity.

 

 

Development of a QTL-environment-based predictive model for node addition rate in common bean.

Author information: Zhang L1, Gezan SA2, Eduardo Vallejos C3, Jones JW1, Boote KJ4, Clavijo-Michelangeli JA4, Bhakta M3, Osorno JM5, Rao I6, Beebe S6, Roman-Paoli E7, Gonzalez A7, Beaver J7, Ricaurte J6, Colbert R5, Correll MJ8.

1Agricultural and Biological Engineering Department, University of Florida, Gainesville, FL, 32611, USA.
2School of Forest Resources and Conservation, University of Florida, Gainesville, FL, 32611, USA.
3Horticultural Sciences Department, University of Florida, Gainesville, FL, 32611, USA.
4Agronomy Department, University of Florida, Gainesville, FL, 32611, USA.
5Department of Plant Sciences, North Dakota State University, Fargo, ND, 58108, USA.
6CIAT, Cali, Colombia.
7University of Puerto Rico, Mayagüez, 00682, Puerto Rico.
8Agricultural and Biological Engineering Department, University of Florida, Gainesville, FL, 32611, USA. correllm@ufl.edu.
Journal: TAG. Theoretical and Applied Genetics. Theoretische und Angewandte Genetik

Date of e-pub: May 2017

Abstract: This work reports the effects of the genetic makeup, the environment and the genotype by environment interactions for node addition rate in an RIL population of common bean. This information was used to build a predictive model for node addition rate. To select a plant genotype that will thrive in targeted environments it is critical to understand the genotype by environment interaction (GEI). In this study, multi-environment QTL analysis was used to characterize node addition rate (NAR, node day- 1) on the main stem of the common bean (Phaseolus vulgaris L). This analysis was carried out with field data of 171 recombinant inbred lines that were grown at five sites (Florida, Puerto Rico, 2 sites in Colombia, and North Dakota). Four QTLs (Nar1, Nar2, Nar3 and Nar4) were identified, one of which had significant QTL by environment interactions (QEI), that is, Nar2 with temperature. Temperature was identified as the main environmental factor affecting NAR while day length and solar radiation played a minor role. Integration of sites as covariates into a QTL mixed site-effect model, and further replacing the site component with explanatory environmental covariates (i.e., temperature, day length and solar radiation) yielded a model that explained 73% of the phenotypic variation for NAR with root mean square error of 16.25% of the mean. The QTL consistency and stability was examined through a tenfold cross validation with different sets of genotypes and these four QTLs were always detected with 50-90% probability. The final model was evaluated using leave-one-site-out method to assess the influence of site on node addition rate. These analyses provided a quantitative measure of the effects on NAR of common beans exerted by the genetic makeup, the environment and their interactions.

 

 

Duvoglustat HCl Increases Systemic and Tissue Exposure of Active Acid α-Glucosidase in Pompe Patients Co-administered with Alglucosidase α.

Author information: Kishnani P1, Tarnopolsky M2, Roberts M3, Sivakumar K4, Dasouki M5, Dimachkie MM5, Finanger E6, Goker-Alpan O7, Guter KA8, Mozaffar T9, Pervaiz MA10, Laforet P11, Levine T12, Adera M13, Lazauskas R14, Sitaraman S14, Khanna R14, Benjamin E14, Feng J14, Flanagan JJ15, Barth J14, Barlow C16, Lockhart DJ17, Valenzano KJ14, Boudes P18, Johnson FK19, Byrne B20.

1Duke University Medical Center, Durham, NC 27710, USA.
2McMaster University Medical Center, Hamilton, ON L8N 3Z5, Canada.
3Salford Royal Hope HNS Trust Hope Hospital, Salford M6 8HD, UK.
4Neuromuscular Research Center, Scottsdale, AZ 85028, USA.
5University of Kansas Medical Center, Kansas City, KS 66160, USA.
6Oregon Health and Science University, Portland, OR 97239, USA.
7LSD Research and Treatment Unit, O&O Alpan, LLC, Fairfax, VA 22030, USA.
8Great Falls Clinic, Great Falls, MT 59405, USA.
9University of California, Irvine, Irvine, CA 92697, USA.
10Emory University, Decatur, GA 30030, USA.
11Hopital la Salpetriere Institut de Myologie, 75013 Paris, France.
12Phoenix Neurological Associates, Phoenix, AZ 85018, USA.
13Insys Therapeutics, Chandler, AZ 85224, USA.
14Amicus Therapeutics, Cranbury, NJ 08512, USA.
15Arvinas, Inc., New Haven, CT 06511, USA.
16The Parkinson’s Institute and Clinical Center, Sunnyvale, CA 94085, USA.
17TranscripTx, Inc., Sunnyvale, CA 94085, USA.
18Cymabay Therapeutics, Newark, CA 94560, USA.
19Amicus Therapeutics, Cranbury, NJ 08512, USA. Electronic address: fjohnson@amicusrx.com.
20University of Florida, Gainesville, FL 32611, USA.
Journal: Molecular Therapy : The Journal of the American Society of Gene Therapy

Date of e-pub: May 2017

Abstract: Duvoglustat HCl (AT2220, 1-deoxynojirimycin) is an investigational pharmacological chaperone for the treatment of acid α-glucosidase (GAA) deficiency, which leads to the lysosomal storage disorder Pompe disease, which is characterized by progressive accumulation of lysosomal glycogen primarily in heart and skeletal muscles. The current standard of care is enzyme replacement therapy with recombinant human GAA (alglucosidase alfa [AA], Genzyme). Based on preclinical data, oral co-administration of duvoglustat HCl with AA increases exposure of active levels in plasma and skeletal muscles, leading to greater substrate reduction in muscle. This phase 2a study consisted of an open-label, fixed-treatment sequence that evaluated the effect of single oral doses of 50 mg, 100 mg, 250 mg, or 600 mg duvoglustat HCl on the pharmacokinetics and tissue levels of intravenously infused AA (20 mg/kg) in Pompe patients. AA alone resulted in increases in total GAA activity and protein in plasma compared to baseline. Following co-administration with duvoglustat HCl, total GAA activity and protein in plasma were further increased 1.2- to 2.8-fold compared to AA alone in all 25 Pompe patients; importantly, muscle GAA activity was increased for all co-administration treatments from day 3 biopsy specimens. No duvoglustat-related adverse events or drug-related tolerability issues were identified.

 

 

Structural Insights into Human Bocaparvoviruses.

Author information: Mietzsch M1, Kailasan S1, Garrison J1, Ilyas M1, Chipman P1, Kantola K2, Janssen ME3, Spear J4, Sousa D4, McKenna R1, Brown K5, Söderlund-Venermo M2, Baker T3, Agbandje-McKenna M6.

1Department of Biochemistry and Molecular Biology, Center for Structural Biology, McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, Florida, USA.
2Department of Virology, University of Helsinki, Helsinki, Finland.
3Department of Chemistry and Biochemistry and Division of Biological Sciences, University of California-San Diego, San Diego, California, USA.
4 Biological Science Imaging Resource, Department of Biological Sciences, The Florida State University, Tallahassee, Florida, USA.
5Virus Reference Department, National Infection Service, Public Health England, London, United Kingdom.
6Department of Biochemistry and Molecular Biology, Center for Structural Biology, McKnight Brain Institute, College of Medicine, University of Florida, Gainesville, Florida, USA mckenna@ufl.edu.
Journal: Journal of Virology

Date of e-pub: May 2017

Abstract: Bocaparvoviruses are emerging pathogens of the Parvoviridae family. Human bocavirus 1 (HBoV1) causes severe respiratory infections and HBoV2 to HBoV4 cause gastrointestinal infections in young children. Recent reports of life-threatening cases, lack of direct treatment or vaccination, and a limited understanding of their disease mechanisms highlight the need to study these pathogens on a molecular and structural level for the development of therapeutics. Toward this end, the capsid structures of HBoV1, HBoV3, and HBoV4 were determined to a resolution of 2.8 to 3.0 Å by cryo-electron microscopy and three-dimensional image reconstruction. The bocaparvovirus capsids, which display different tissue tropisms, have features in common with other parvoviruses, such as depressions at the icosahedral 2-fold symmetry axis and surrounding the 5-fold symmetry axis, protrusions surrounding the 3-fold symmetry axis, and a channel at the 5-fold symmetry axis. However, unlike other parvoviruses, densities extending the 5-fold channel into the capsid interior are conserved among the bocaparvoviruses and are suggestive of a genus-specific function. Additionally, their major viral protein 3 contains loops with variable regions at their apexes conferring capsid surface topologies different from those of other parvoviruses. Structural comparisons at the strain (HBoV) and genus (bovine parvovirus and HBoV) levels identified differences in surface loops that are functionally important in host/tissue tropism, pathogenicity, and antigenicity in other parvoviruses and likely play similar roles in these viruses. This study thus provides a structural framework to characterize determinants of host/tissue tropism, pathogenicity, and antigenicity for the development of antiviral strategies to control human bocavirus infections.IMPORTANCE Human bocaviruses are one of only a few members of the Parvoviridae family pathogenic to humans, especially young children and immunocompromised adults. There are currently no treatments or vaccines for these viruses or the related enteric bocaviruses. This study obtained the first high-resolution structures of three human bocaparvoviruses determined by cryo-reconstruction. HBoV1 infects the respiratory tract, and HBoV3 and HBoV4 infect the gastrointestinal tract, tissues that are likely targeted by the capsid. Comparison of these viruses provides information on conserved bocaparvovirus-specific features and variable regions resulting in unique surface topologies that can serve as guides to characterize HBoV determinants of tissue tropism and antigenicity in future experiments. Based on the comparison to other existing parvovirus capsid structures, this study suggests capsid regions that likely control successful infection, including determinants of receptor attachment, host cell trafficking, and antigenic reactivity. Overall, these observations could impact efforts to design antiviral strategies and vaccines for HBoVs.

 

 

Fluorescence Resonance Energy Transfer-Based DNA Tetrahedron Nanotweezer for Highly Reliable Detection of Tumor-Related mRNA in Living Cells.

Author information: He L1, Lu DQ1, Liang H1, Xie S1, Luo C1, Hu M1, Xu L1, Zhang X1, Tan W1,2.

1Molecular Science and Biomedicine Laboratory, State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Life Sciences, and Collaborative Innovation Center for Chemistry and Molecular Medicine, Hunan University , Changsha 410082, China.
2Department of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, Health Cancer Center, University of Florida , Gainesville, Florida 32611-7200, United States.
Journal: ACS Nano

Date of e-pub: April 2017

Abstract: Accurate detection and imaging of tumor-related mRNA in living cells hold great promise for early cancer detection. However, currently, most probes designed to image intracellular mRNA confront intrinsic interferences arising from complex biological matrices and resulting in inevitable false-positive signals. To circumvent this problem, an intracellular DNA nanoprobe, termed DNA tetrahedron nanotweezer (DTNT), was developed to reliably image tumor-related mRNA in living cells based on the FRET (fluorescence resonance energy transfer) “off” to “on” signal readout mode. DTNT was self-assembled from four single-stranded DNAs. In the absence of target mRNA, the respectively labeled donor and acceptor fluorophores are separated, thus inducing low FRET efficiency. However, in the presence of target mRNA, DTNT alters its structure from the open to closed state, thus bringing the dual fluorophores into close proximity for high FRET efficiency. The DTNT exhibited high cellular permeability, fast response and excellent biocompatibility. Moreover, intracellular imaging experiments showed that DTNT could effectively distinguish cancer cells from normal cells and, moreover, distinguish among changes of mRNA expression levels in living cells. The DTNT nanoprobe also exhibits minimal effect of probe concentration, distribution and laser power as other ratiometric probe. More importantly, as a result of the FRET “off” to “on” signal readout mode, the DTNT nanoprobe almost entirely avoids false-positive signals due to intrinsic interferences, such as nuclease digestion, protein binding and thermodynamic fluctuations in complex biological matrices. This design blueprint can be applied to the development of powerful DNA nanomachines for biomedical research and clinical early diagnosis.

 

 

Electrophysiologic features of ulnar neuropathy in childhood and adolescence.

Author information: Karakis I1, Liew W2, Fournier HS3, Jones HR Jr4, Darras BT3, Kang PB5.

1Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA; Department of Neurology, Lahey Clinic, Burlington, MA, USA; Department of Neurology, Emory University School of Medicine, Atlanta, GA, USA.
2Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA; Department of Neurology, KK Women’s & Children’s Hospital, Singapore.
3Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA.
4Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA; Department of Neurology, Lahey Clinic, Burlington, MA, USA.
5Department of Neurology, Boston Children’s Hospital and Harvard Medical School, Boston, MA, USA; Division of Pediatric Neurology, Department of Pediatrics, University of Florida College of Medicine, Gainesville, FL, USA; Department of Neurology, University of Florida College of Medicine, Gainesville, FL, USA. Electronic address: pbkang@ufl.edu.
Journal: Clinical Neurophysiology : Official Journal of the International Federation of Clinical Neurophysiology

Date of e-pub: May 2017

Abstract: To analyze patterns of nerve injury in pediatric ulnar neuropathy (PUN).

Retrospective analysis of 49 children with PUN.

Sensory loss in digit V was the prevailing complaint (89%). Predominant localization was at the elbow (55%). Diminished ulnar SNAP was the most common abnormality (71%) with median axon loss estimate (MAXE) of 62%. Dorsal ulnar cutaneous (DUC) sensory nerve action potential (SNAP) was reduced in 55% with MAXE of 43%. Abductor digiti minimi (ADM) and first dorsal interosseous (FDI) compound muscle action potential (CMAP) were reduced half of the time, with MAXE of 30% and 28% respectively. There was high correlation between ulnar sensory MAXE and ADM MAXE (r=0.76, p<0.0001), FDI MAXE (r=0.81, p<0.0001) and DUC MAXE (r=0.60, p=0.0048). Neurogenic changes were seen in the ADM, FDI, flexor carpi ulnaris (FCU) and flexor digitorum profundus IV (FDP IV) in 79%, 77%, 25% and 35% respectively. Pathophysiology was demyelinating in 27%, axonal in 59% and mixed in 14%.

In proximal axonal lesions, sensory fibers to digit V and motor fibers to distal muscles are predominantly affected, whereas in demyelinating lesions, slowing occurs twice as frequently as conduction block.

There is frequent axonal and fascicular injury in PUN.

 

 

DNA probes for monitoring dynamic and transient molecular encounters on live cell membranes.

Author information: You M1,2,3,4, Lyu Y1,2, Han D1,2, Qiu L1,2, Liu Q1, Chen T1,2, Sam Wu C1,2, Peng L2, Zhang L1,2, Bao G5, Tan W1,2.

1Molecular Science and Biomedicine Laboratory, State Key Laboratory of Chemo/Bio-sensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Collaborative Innovation Center for Chemistry and Molecular Medicine, Hunan University, Changsha 410082, People’s Republic of China.
2Department of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, Health Cancer Center, UF Genetics Institute, McKnight Brain Institute, University of Florida, Gainesville, Florida 32611, USA.
3Department of Chemistry, University of Massachusetts Amherst, Amherst, Massachusetts 01003, USA.
4Department of Mathematics, Michigan State University, East Lansing, Michigan 48824, USA.
5School of Mathematical Sciences, Zhejiang University, Hangzhou, Zhejiang 310027, People’s Republic of China.
Journal: Nature Nanotechnology

Date of e-pub: May 2017

Abstract: Cells interact with the extracellular environment through molecules expressed on the membrane. Disruption of these membrane-bound interactions (or encounters) can result in disease progression. Advances in super-resolution microscopy have allowed membrane encounters to be examined, however, these methods cannot image entire membranes and cannot provide information on the dynamic interactions between membrane-bound molecules. Here, we show a novel DNA probe that can transduce transient membrane encounter events into readable cumulative fluorescence signals. The probe, which translocates from one anchor site to another, mimicking motor proteins, is realized through a toehold-mediated DNA strand displacement reaction. Using this probe, we successfully monitored rapid encounter events of membrane lipid domains using flow cytometry and fluorescence microscopy. Our results show a preference for encounters within the same lipid domains.

 

 

Impact of the CYP2C19 genotype on voriconazole exposure in adults with invasive fungal infections.

Author information: Hamadeh IS1, Klinker KP, Borgert SJ, Richards AI, Li W, Mangal N, Hiemenz JW, Schmidt S, Langaee TY, Peloquin CA, Johnson JA, Cavallari LH.

1aDepartment of Pharmacotherapy and Translational Research bCenter for Pharmacogenomics, College of Pharmacy, University of Florida cUniversity of Florida Health Shands Hospital, Gainesville dDepartment of Pharmaceutics eCenter for Pharmacometrics and Systems Pharmacology, College of Pharmacy, University of Florida, Orlando fDepartment of Medicine, Division of Hematology and Oncology gDepartment of Medicine, Division of Cardiology, College of Medicine, University of Florida, Gainesville, Florida, USA.
Journal: Pharmacogenetics and Genomics

Date of e-pub: May 2017

Abstract: Voriconazole, a first-line agent for the treatment of invasive fungal infections (IFIs), is metabolized by CYP2C19. A significant proportion of patients fail to achieve therapeutic trough concentrations with standard weight-based voriconazole dosing, placing them at increased risk for treatment failure, which can be life threatening. We sought to test the association between the CYP2C19 genotype and subtherapeutic voriconazole concentrations in adults with IFIs.

Adults receiving weight-based voriconazole dosing for the treatment of IFIs were genotyped for the CYP2C19*2, *3, and *17 polymorphisms, and CYP2C19 metabolizer phenotypes were inferred. Steady-state voriconazole trough plasma concentrations and the prevalence of subtherapeutic troughs (

Of 70 patients included (mean age 52.5±18 years), 39% were RMs or UMs. Compared with patients with the other phenotypes, RMs/UMs had a lower steady-state trough concentration (4.26±2.2 vs. 2.86±2.3, P=0.0093) and a higher prevalence of subtherapeutic troughs (16 vs. 52%, P=0.0028), with an odds ratio of 5.6 (95% confidence interval: 1.64-19.24, P=0.0044).

Our findings indicate that adults with the CYP2C19 RM or UM phenotype are more likely to have subtherapeutic concentrations with weight-based voriconazole dosing. These results corroborate previous findings in children and support the potential clinical utility of CYP2C19 genotype-guided voriconazole dosing to avoid underexposure in RMs and UMs.

 

 

Electroacupuncture Promotes Central Nervous System-Dependent Release of Mesenchymal Stem Cells.

Author information: Salazar TE1,2,3, Richardson MR4, Beli E3, Ripsch MS5,6, George J2, Kim Y5, Duan Y3, Moldovan L3, Yan Y1, Bhatwadekar A3, Jadhav V5, Smith JA5, McGorray S7, Bertone AL8, Traktuev DO9,10, March KL9,10, Colon-Perez LM11, Avin KG12, Sims E13, Mund JA4,13, Case J13,14, Deng X15, Kim MS16, McDavitt B17, Boulton ME3, Thinschmidt J18, Li Calzi S3, Fitz SD19, Fuchs RK12, Warden SJ12, McKinley T20, Shekhar A19, Febo M11, Johnson PL21, Chang LJ22,23, Gao Z24, Kolonin MG24, Lai S25, Ma J25, Dong X26, White FA5,6, Xie H27, Yoder MC4,13, Grant MB3.

1Genetics Institute, Gainesville, Florida, USA.
2College of Medicine, Gainesville, Florida, USA.
3Department of Ophthalmology, Indianapolis, Indiana, USA.
4Wells Center for Pediatric Research, Indianapolis, Indiana, USA.
5Department of Anesthesia, Indianapolis, Indiana, USA.
6Richard L. Roudebush VA Medical Center, Indianapolis, Indiana, USA.
7Department of Biostatistics, Gainesville, Florida, USA.
8Department of Veterinary Clinical Sciences, The Ohio State University, Columbus, Ohio, USA.
9Krannert Institute of Cardiology, Indianapolis, Indiana, USA.
10Indiana Center for Vascular Biology and Medicine, Indianapolis, Indiana, USA.
11Department of Psychiatry, McKnight Brain Institute, Gainesville, Florida, USA.
12Department of Physical Therapy, School of Health and Rehabilitation Sciences, Indianapolis, Indiana, USA.
13Melvin and Bren Simon Cancer Center, Indiana University, Indianapolis, Indiana, USA.
14Scripps Clinic Medical Group, Scripps Center for Organ and Cell Transplantation, La Jolla, California, USA.
15Mainland Acupuncture Center, Gainesville, Florida, USA.
16College of Veterinary Medicine, Chon Buk National University, Jeonju, South Korea.
17McDavitt Veterinary Clinic, Zionsville, Indiana, USA.
18Department of Pharmacology and Therapeutics, Gainesville, Florida, USA.
19Department of Psychiatry, Indianapolis, Indiana, USA.
20Department of Orthopedic Surgery, Indianapolis, Indiana, USA.
21Department of Anatomy and Cell Biology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
22Department of Molecular Genetics and Microbiology, Gainesville, Florida, USA.
23School of Medicine, University of Electronic Science and Technology of China, Sichuan, China.
24The Brown Foundation, Institute of Molecular Medicine, University of Texas Health Science Center, Houston, Texas, USA.
25Department of Radiation Oncology, College of Medicine, Gainesville, Florida, USA.
26Department of Neuroscience, Center of Sensory Biology, the Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
27College of Veterinary Medicine, University of Florida, Gainesville, Florida, USA.
Journal: Stem Cells

Date of e-pub: May 2017

Abstract: Electroacupuncture (EA) performed in rats and humans using limb acupuncture sites, LI-4 and LI-11, and GV-14 and GV-20 (humans) and Bai-hui (rats) increased functional connectivity between the anterior hypothalamus and the amygdala and mobilized mesenchymal stem cells (MSCs) into the systemic circulation. In human subjects, the source of the MSC was found to be primarily adipose tissue, whereas in rodents the tissue sources were considered more heterogeneous. Pharmacological disinhibition of rat hypothalamus enhanced sympathetic nervous system (SNS) activation and similarly resulted in a release of MSC into the circulation. EA-mediated SNS activation was further supported by browning of white adipose tissue in rats. EA treatment of rats undergoing partial rupture of the Achilles tendon resulted in reduced mechanical hyperalgesia, increased serum interleukin-10 levels and tendon remodeling, effects blocked in propranolol-treated rodents. To distinguish the afferent role of the peripheral nervous system, phosphoinositide-interacting regulator of transient receptor potential channels (Pirt)-GCaMP3 (genetically encoded calcium sensor) mice were treated with EA acupuncture points, ST-36 and LIV-3, and GV-14 and Bai-hui and resulted in a rapid activation of primary sensory neurons. EA activated sensory ganglia and SNS centers to mediate the release of MSC that can enhance tissue repair, increase anti-inflammatory cytokine production and provide pronounced analgesic relief. Stem Cells 2017;35:1303-1315.

 

 

The IGNITE Pharmacogenetics Working Group: An Opportunity for Building Evidence with Pharmacogenetic Implementation in a Real-World Setting.

Author information: Cavallari LH1, Beitelshees AL2, Blake KV3, Dressler LG4, Duarte JD1, Elsey A1, Eichmeyer JN5, Empey PE6, Franciosi JP7, Hicks JK8, Holmes AM9, Jeng L10, Lee CR11, Lima JJ3, Limdi NA12, Modlin J5, Obeng AO13, Petry N14, Pratt VM15, Skaar TC16, Tuteja S17, Voora D18, Wagner M5, Weitzel KW1, Wilke RA19, Peterson JF20, Johnson JA1.

1Department of Pharmacotherapy and Translational Research, University of Florida, Gainesville, Florida, USA.
2Department of Medicine, University of Maryland, Baltimore, Maryland, USA.
3Biomedical Research Department, Nemours Children’s Specialty Care, Jacksonville, Florida, USA.
4Personalized Medicine and Pharmacogenetics Program, Mission Health, Asheville, North Carolina, USA.
5Department of Oncology, St. Luke’s Mountain States Tumor Institute, Boise, Idaho, USA.
6Department of Pharmacy and Therapeutics, Center for Clinical Pharmaceutical Sciences, University of Pittsburgh School of Pharmacy, Pittsburgh, Pennsylvania, USA.
7Biomedical Research Department, Nemours Children’s Specialty Care, Orlando, Florida, USA.
8Division of Population Science, DeBartolo Family Personalized Medicine Institute, Moffitt Cancer Center, Tampa, Florida, USA.
9 Department of Health Policy and Management, Richard M. Fairbanks School of Public Health, Indiana University – Purdue University, Indianapolis, Indiana, USA.
10Departments of Medicine, Pathology, and Pediatrics, University of Maryland School of Medicine, Baltimore, Maryland, USA.
11Division of Pharmacotherapy and Experimental Therapeutics, Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.
12Department of Neurology, University of Alabama, Birmingham, Alabama, USA.
13Charles Bronfman Institute for Personalized Medicine, Icahn School of Medicine at Mount Sinai, New York, New York, USA.
14Department of Pharmacy Practice, North Dakota State University, Fargo, North Dakota, USA.
15Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, Indiana, USA.
16Department of Medicine, Division of Clinical Pharmacology, Indiana University School of Medicine, Indianapolis, Indiana, USA.
17Division of Translational Medicine and Human Genetics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania, USA.
18Center for Applied Genomics & Precision Medicine, Department of Medicine, Duke University, Durham, North Carolina, USA.
19Department of Internal Medicine, University of South Dakota, Sioux Falls, South Dakota, USA.
20Departments of Biomedical Informatics and Medicine, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
Journal: Clinical and Translational Science

Date of e-pub: May 2017

Abstract: N/A

 

 

Aptasensor with Expanded Nucleotide Using DNA Nanotetrahedra for Electrochemical Detection of Cancerous Exosomes.

Author information: Wang S1,2, Zhang L3,2, Wan S2, Cansiz S2, Cui C2, Liu Y3,2, Cai R2, Hong C2, Teng IT2, Shi M3,2, Wu Y3,2, Dong Y1, Tan W3,2.

1Beijing Key Laboratory of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology , Beijing 100029, China.
2Department of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, Health Cancer Center, McKnight Brain Institute, UF Genetics Institute, University of Florida , Gainesville, Florida 32611-7200, United States.
3Molecular Science and Biomedicine Laboratory, State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, College of Biology, Collaborative Innovation Center for Chemistry and Molecular Medicine, Hunan University , Changsha 410082, China.
Journal: ACS Nano

Date of e-pub: April 2017

Abstract: Exosomes are extracellular vesicles (50-100 nm) circulating in biofluids as intercellular signal transmitters. Although the potential of cancerous exosomes as tumor biomarkers is promising, sensitive and rapid detection of exosomes remains challenging. Herein, we combined the strengths of advanced aptamer technology, DNA-based nanostructure, and portable electrochemical devices to develop a nanotetrahedron (NTH)-assisted aptasensor for direct capture and detection of hepatocellular exosomes. The oriented immobilization of aptamers significantly improved the accessibility of an artificial nucleobase-containing aptamer to suspended exosomes, and the NTH-assisted aptasensor could detect exosomes with 100-fold higher sensitivity when compared to the single-stranded aptamer-functionalized aptasensor. The present study provides a proof-of-concept for sensitive and efficient quantification of tumor-derived exosomes. We thus expect the NTH-assisted electrochemical aptasensor to become a powerful tool for comprehensive exosome studies.

 

 

tRNA N6-adenosine threonylcarbamoyltransferase defect due to KAE1/TCS3 (OSGEP) mutation manifest by neurodegeneration and renal tubulopathy.

Author information: Edvardson S1,2, Prunetti L3, Arraf A4, Haas D3, Bacusmo JM3, Hu JF5, Ta-Shma A1, Dedon PC5,6, de Crécy-Lagard V3, Elpeleg O1.

1Monique and Jacques Roboh Department of Genetic Research, Hadassah Medical Center, Hebrew University of Jerusalem, Jerusalem, Israel.
2Pediatric Neurology Unit, Hadassah Medical Center, Hebrew University of Jerusalem, Jerusalem, Israel.
3Department of Microbiology and Cell Science, Institute for Food and Agricultural Sciences and Genetic Institute, University of Florida, Gainesville, FL, USA.
4Hebrew University School of Medicine, Jerusalem, Israel.
5Center for Environmental Health Sciences, Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA, USA.
6Infectious Disease Interdisciplinary Research Group, Singapore-MIT Alliance for Research and Technology, Singapore, Singapore.
Journal: European Journal of Human Genetics : EJHG

Date of e-pub: May 2017

Abstract: Post-transcriptional tRNA modifications are numerous and require a large set of highly conserved enzymes in humans and other organisms. In yeast, the loss of many modifications is tolerated under unstressed conditions; one exception is the N6-threonyl-carbamoyl-adenosine (t6A) modification, loss of which causes a severe growth phenotype. Here we aimed at a molecular diagnosis in a brother and sister from a consanguineous family who presented with global developmental delay, failure to thrive and a renal defect manifesting in proteinuria and hypomagnesemia. Using exome sequencing, the patients were found to be homozygous for the c.974G>A (p.(Arg325Gln)) variant of the KAE1 gene. KAE1 is a constituent of the KEOPS complex, a five-subunit complex that catalyzes the second biosynthetic step of t6A in the cytosol. The yeast KAE1 allele carrying the equivalent mutation did not rescue the t6A deficiency of the kae1Δ yeast strain as efficiently as the WT allele; furthermore, t6A levels quantified by LC-MS/MS were lower in the kae1Δ strain which was complemented by the mutation than in the kae1Δ strain, which was complemented by the WT allele. We conclude that homozygosity for c.974G>A (p.(Arg325Gln)) in KAE1 likely exerts its pathogenic effect by perturbing t6A synthesis, thereby interfering with global protein production. This is the first report of t6A biosynthesis defect in human. KAE1 joins the growing list of cytoplasmic tRNA modification enzymes, all associated with severe neurological disorders.

 

 

Association of single nucleotide polymorphisms in candidate genes previously related to genetic variation in fertility with phenotypic measurements of reproductive function in Holstein cows.

Author information: Ortega MS1, Denicol AC1, Cole JB2, Null DJ2, Taylor JF3, Schnabel RD4, Hansen PJ5.

1Department of Animal Sciences, D.H. Barron Reproductive and Perinatal Biology Research Program and Genetics Institute, University of Florida, Gainesville 32611.
2Animal Genomics and Improvement Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705.
3Division of Animal Sciences, University of Missouri, Columbia 65211.
4Division of Animal Sciences, University of Missouri, Columbia 65211; Informatics Institute, University of Missouri, Columbia 65211.
5Department of Animal Sciences, D.H. Barron Reproductive and Perinatal Biology Research Program and Genetics Institute, University of Florida, Gainesville 32611. Electronic address: pjhansen@ufl.edu.
Journal: Journal of Dairy Science

Date of e-pub: May 2017

Abstract: Many genetic markers related to health or production traits are not evaluated in populations independent of the discovery population or related to phenotype. Here we evaluated 68 single nucleotide polymorphisms (SNP) in candidate genes previously associated with genetic merit for fertility and production traits for association with phenotypic measurements of fertility in a population of Holstein cows that was selected based on predicted transmitting ability (PTA) for daughter pregnancy rate (DPR; high, ≥1, n = 989; low, ≤ -1.0, n = 1,285). Cows with a high PTA for DPR had higher pregnancy rate at first service, fewer services per conception, and fewer days open than cows with a low PTA for DPR. Of the 68 SNP, 11 were associated with pregnancy rate at first service, 16 with services per conception, and 19 with days open. Single nucleotide polymorphisms in 12 genes (BDH2, BSP3, CAST, CD2, CD14, FUT1, FYB, GCNT3, HSD17B7, IBSP, OCLN, and PCCB) had significant associations with 2 fertility traits, and SNP in 4 genes (CSPP1, FCER1G, PMM2, and TBC1D24) had significant associations with each of the 3 traits. Results from this experiment were compared with results from 2 earlier studies in which the SNP were associated with genetic estimates of fertility. One study involved the same animals as used here, and the other study was of an independent population of bulls. A total of 13 SNP associated with 1 or more phenotypic estimates of fertility were directionally associated with genetic estimates of fertility in the same cow population. Moreover, 14 SNP associated with reproductive phenotype were directionally associated with genetic estimates of fertility in the bull population. Nine SNP (located in BCAS, BSP3, CAST, FUT1, HSD17B7, OCLN, PCCB, PMM2, and TBC1D24) had a directional association with fertility in all 3 studies. Examination of the function of the genes with SNP associated with reproduction in more than one study indicates the importance of steroid hormones and immune function as determinants of reproductive function. All but 1 of the 68 evaluated SNP were variable in 11 breeds besides Holstein, indicating the potential effects of these SNP on reproductive function across breeds of cattle.

 

 

Effects of supplementation with docosahexaenoic acid on reproduction of dairy cows.

Author information: Sinedino LD1,2, Honda PM1, Souza LR1, Lock AL3, Boland MP4, Staples CR1, Thatcher WW1,2, Santos JE5,2.

1Department of Animal SciencesUniversity of Florida, Gainesville, Florida, USA.
2DH Barron Reproductive and Perinatal Biology Research ProgramUniversity of Florida, Gainesville, Florida, USA.
3Department of Animal ScienceMichigan State University, East Lansing, Michigan, USA.
4Alltech Inc.Nicholasville, Kentucky, USA.
5Department of Animal SciencesUniversity of Florida, Gainesville, Florida, USA jepsantos@ufl.edu.
Journal: Reproduction (Cambridge, England)

Date of e-pub: May 2017

Abstract: The objectives were to determine the effects of supplementing docosahexaenoic acid (DHA)-rich algae on reproduction of dairy cows. Holstein cows were assigned randomly to either a control (n = 373) or the same diet supplemented daily with 100 g/cow of an algae product containing 10% DHA (algae, n = 366) from 27 to 147 days postpartum. Measurements included yields of milk and milk components, fatty acids (FA) profiles in milk fat and plasma phospholipids, resumption of ovulation by 57 days postpartum, pregnancy per artificial insemination (AI) and expression of interferon-stimulated genes in leukocytes. Feeding algae increased resumption of estrous cyclicity (77.6 vs 65.9%) and pregnancy at first AI (47.6 vs 32.8%) in primiparous cows. Algae increased pregnancy per AI in all AI in both primiparous and multiparous cows (41.6 vs 30.7%), which reduced days to pregnancy by 22 days (102 vs 124 days) compared with control cows. Pregnant cows fed algae had greater expression of RTP4 in blood leukocytes compared with those in pregnant control cows. Feeding algae increased the incorporation of DHA, eicosapentaenoic acid, conjugated linoleic acid isomers cis-9 trans-11, trans-10 cis-12 and total n-3 FA in phospholipids in plasma and milk fat. Yields of milk and true protein increased by 1.1 kg/day and 30 g/day respectively, whereas fat yield decreased 40 g/day in algae compared with that in control. Supplementing DHA-rich algae altered the FA composition of lipid fractions and improved reproduction in dairy cows. The benefits on reproduction might be mediated by enhanced embryo development based on changes in interferon-stimulated gene expression.

 

 

Conditional knockout of activin like kinase-1 (ALK-1) leads to heart failure without maladaptive remodeling.

Author information: Morine KJ1, Qiao X1, Paruchuri V1, Aronovitz MJ1, Mackey EE1, Buiten L1, Levine J1, Ughreja K1, Nepali P1, Blanton RM1, Karas RH1, Oh SP2, Kapur NK3.

1Molecular Cardiology Research Institute and Division of Cardiology, Department of Medicine, Tufts Medical Center, 800 Washington Street, Boston, MA, 02111, USA.
2Department of Physiology and Functional Genomics, University of Florida, 1345 Center Drive, Gainesville, FL, 32610-0274, USA.
3Molecular Cardiology Research Institute and Division of Cardiology, Department of Medicine, Tufts Medical Center, 800 Washington Street, Boston, MA, 02111, USA. Nkapur@tuftsmedicalcenter.org.
Journal: Heart and Vessels

Date of e-pub: May 2017

Abstract: Activin like kinase-1 (AlK-1) mediates signaling via the transforming growth factor beta (TGFβ) family of ligands. AlK-1 activity promotes endothelial proliferation and migration. Reduced AlK-1 activity is associated with arteriovenous malformations. No studies have examined the effect of global AlK-1 deletion on indices of cardiac remodeling. We hypothesized that reduced levels of AlK-1 promote maladaptive cardiac remodeling. To test this hypothesis, we employed AlK-1 conditional knockout mice (cKO) harboring the ROSA26-CreER knock-in allele, whereby a single dose of intraperitoneal tamoxifen triggered ubiquitous Cre recombinase-mediated excision of floxed AlK-1 alleles. Tamoxifen treated wild-type (WT-TAM; n = 5) and vehicle treated AlK-1-cKO mice (cKO-CON; n = 5) served as controls for tamoxifen treated AlK-1-cKO mice (cKO-TAM; n = 15). AlK-1 cKO-TAM mice demonstrated reduced 14-day survival compared to cKO-CON controls (13 vs 100%, respectively, p < 0.01). Seven days after treatment, cKO-TAM mice exhibited reduced left ventricular (LV) fractional shortening, progressive LV dilation, and gastrointestinal bleeding. After 14 days total body mass was reduced, but LV and lung mass increased in cKO-TAM not cKO-CON mice. Peak LV systolic pressure, contractility, and arterial elastance were reduced, but LV end-diastolic pressure and stroke volume were increased in cKO-TAM, not cKO-CON mice. LV AlK-1 mRNA levels were reduced in cKO-TAM, not cKO-CON mice. LV levels of other TGFβ-family ligands and receptors (AlK5, TBRII, BMPRII, Endoglin, BMP7, BMP9, and TGFβ1) were unchanged between groups. Cardiomyocyte area and LV levels of BNP were increased in cKO-TAM mice, but LV levels of β-MHC and SERCA were unchanged. No increase in markers of cardiac fibrosis, Type I collagen, CTGF, or PAI-1, were observed between groups. No differences were observed for any variable studied between cKO-CON and WT-TAM mice. Global deletion of AlK-1 is associated with the development of high output heart failure without maladaptive remodeling. Future studies exploring the functional role of AlK-1 in cardiac remodeling independent of systemic AVMs are required.

 

 

Diversification of Rosaceae since the Late Cretaceous based on plastid phylogenomics.

Author information: Zhang SD1, Jin JJ1,2, Chen SY1, Chase MW3,4, Soltis DE5,6,7, Li HT1, Yang JB1, Li DZ1, Yi TS1.

1Germplasm Bank of Wild Species, Kunming Institute of Botany, Chinese Academy of Sciences, Kunming, 650201, China.
2Kunming College of Life Sciences, University of Chinese Academy of Sciences, Kunming, Yunnan 650201, China.
3Science Directorate, Royal Botanic Gardens, Kew, Richmond, Surrey, TW9 3DS, UK.
4School of Plant Biology, University of Western Australia, 35 Stirling Highway, Crawley, WA, 6009, Australia.
5Florida Museum of Natural History, University of Florida, Gainesville, FL, 32611-7800, USA.
6Department of Biology, University of Florida, Gainesville, FL, 32611, USA.
7Genetics Institute, University of Florida, Gainesville, FL, 32608, USA.
Journal: The New Phytologist

Date of e-pub: May 2017

Abstract: Phylogenetic relationships in Rosaceae have long been problematic because of frequent hybridisation, apomixis and presumed rapid radiation, and their historical diversification has not been clarified. With 87 genera representing all subfamilies and tribes of Rosaceae and six of the other eight families of Rosales (outgroups), we analysed 130 newly sequenced plastomes together with 12 from GenBank in an attempt to reconstruct deep relationships and reveal temporal diversification of this family. Our results highlight the importance of improving sequence alignment and the use of appropriate substitution models in plastid phylogenomics. Three subfamilies and 16 tribes (as previously delimited) were strongly supported as monophyletic, and their relationships were fully resolved and strongly supported at most nodes. Rosaceae were estimated to have originated during the Late Cretaceous with evidence for rapid diversification events during several geological periods. The major lineages rapidly diversified in warm and wet habits during the Late Cretaceous, and the rapid diversification of genera from the early Oligocene onwards occurred in colder and drier environments. Plastid phylogenomics offers new and important insights into deep phylogenetic relationships and the diversification history of Rosaceae. The robust phylogenetic backbone and time estimates we provide establish a framework for future comparative studies on rosaceous evolution.

 

 

Seasonal resource value and male size influence male aggressive interactions in the leaf footed cactus bug, Narnia femorata.

Author information: Nolen ZJ1, Allen PE2, Miller CW3.

1Entomology and Nematology Department, University of Florida, 1881 Natural Area Drive, Gainesville, FL, 32611, USA. Electronic address: zjnolen@ufl.edu.
2Entomology and Nematology Department, University of Florida, 1881 Natural Area Drive, Gainesville, FL, 32611, USA. Electronic address: pabloallen@ufl.edu.
3Entomology and Nematology Department, University of Florida, 1881 Natural Area Drive, Gainesville, FL, 32611, USA. Electronic address: cwmiller@ufl.edu.
Journal: Behavioural Processes

Date of e-pub: May 2017

Abstract: In animal contests, resource value (the quality of a given resource) and resource holding potential (a male’s absolute fighting ability) are two important factors determining the level of engagement and outcome of contests. Few studies have tested these factors simultaneously. Here, we investigated whether natural, seasonal differences in cactus phenology (fruit quality) influence interactions between males in the leaf-footed cactus bug, Narnia femorata (Hemiptera: Coreidae). We also considered whether males were more likely to interact when they were similar in size, as predicted by theory. Finally, we examined if male size relative to the size of an opponent predicted competitive success. We found that males have more interactions on cactus with high value ripe fruit, as we predicted. Further, we found that males that were closer in size were more likely to interact, and larger males were more likely to become dominant.

 

 

Structure and Origin of the White Cap Locus and Its Role in Evolution of Grain Color in Maize.

Author information: Tan BC1, Guan JC2, Ding S1, Wu S2, Saunders JW2, Koch KE2, McCarty DR3.

1Key Laboratory of Plant Cell Engineering and Germplasm Innovation, Ministry of Education, School of Life Sciences, Shandong University, Jinan, Shandong 250100, China.
2Plant Molecular Cellular Biology Program and Genetics Institute, Horticultural Sciences Department, University of Florida, Gainesville, Florida 32611.
3Plant Molecular Cellular Biology Program and Genetics Institute, Horticultural Sciences Department, University of Florida, Gainesville, Florida 32611 drm@ufl.edu.
Journal: Genetics

Date of e-pub: May 2017

Abstract: Selection for yellow- and white-grain types has been central to postdomestication improvement of maize. While genetic control of carotenoid biosynthesis in endosperm is attributed primarily to the Yellow1 (Y1) phytoene synthase gene, less is known about the role of the dominant white endosperm factor White Cap (Wc). We show that the Wc locus contains multiple, tandem copies of a Carotenoid cleavage dioxygenase 1 (Ccd1) gene that encodes a carotenoid-degrading enzyme. A survey of 111 maize inbreds and landraces, together with 22 teosinte accessions, reveals that Wc is exclusive to maize, where it is prevalent in white-grain (y1) varieties. Moreover, Ccd1 copy number varies extensively among Wc alleles (from 1 to 23 copies), and confers a proportional range of Ccd1 expression in diverse organs. We propose that this dynamic source of quantitative variation in Ccd1 expression was created in maize shortly after domestication by a two-step, Tam3L transposon-mediated process. First, a chromosome segment containing Ccd1 and several nearby genes duplicated at a position 1.9 Mb proximal to the progenitor Ccd1r locus on chromosome 9. Second, a subsequent interaction of Tam3L transposons at the new locus created a 28-kb tandem duplication, setting up expansion of Ccd1 copy number by unequal crossing over. In this way, transposon-mediated variation in copy number at the Wc locus generated phenotypic variation that provided a foundation for breeding and selection of white-grain color in maize.

 

 

The Arabidopsis Elongator complex is required for nonhost resistance against the bacterial pathogens Xanthomonas citri subsp. citri and Pseudomonas syringae pv. phaseolicola NPS3121.

Author information: An C1, Wang C1, Mou Z1.

1Department of Microbiology and Cell Science, University of Florida, PO Box 110700, Gainesville, FL, 32611, USA.
Journal: The New Phytologist

Date of e-pub: May 2017

Abstract: Although in recent years nonhost resistance has attracted considerable attention for its broad spectrum and durability, the genetic and mechanistic components of nonhost resistance have not been fully understood. We used molecular and histochemical approaches including quantitative PCR, chromatin immunoprecipitation, and 3,3′-diaminobenzidine and aniline blue staining. The evolutionarily conserved histone acetyltransferase complex Elongator was identified as a major component of nonhost resistance against Xanthomonas citri subsp. citri (Xcc) and Pseudomonas syringae pv. phaseolicola (Psp) NPS3121. Mutations in Elongator genes inhibit Xcc-, Psp NPS3121- and/or flg22-induced defense responses including defense gene expression, callose deposition, and reactive oxygen species (ROS) and salicylic acid (SA) accumulation. Mutations in Elongator also attenuate the ROS-SA amplification loop. We show that suppressed ROS and SA accumulation in Elongator mutants is correlated with reduced expression of the Arabidopsis respiratory burst oxidase homologue AtrbohD and the SA biosynthesis gene ISOCHORISMATE SYNTHASE1 (ICS1). Furthermore, we found that the Elongator subunit ELP2 is associated with the chromatin of AtrbohD and ICS1 and is required for maintaining basal histone H3 acetylation levels in these key defense genes. As both AtrbohD and ICS1 contribute to nonhost resistance against Xcc, our results reveal an epigenetic mechanism by which Elongator regulates nonhost resistance in Arabidopsis.

 

 

Anticoagulation endpoints with clinical implementation of warfarin pharmacogenetic dosing in a real-world setting: A proposal for a new pharmacogenetic dosing approach.

Author information: Arwood MJ1,2, Deng J3, Drozda K4, Pugach O5, Nutescu EA6,7,8, Schmidt S3, Duarte JD1,2, Cavallari LH1,2.

1Department of Pharmacotherapy and Translational Research, University of Florida, Gainesville, Florida, USA.
2Center for Pharmacogenomics, University of Florida, Gainesville, Florida, USA.
3Center for Pharmacometrics and Systems Pharmacology, University of Florida at Lake Nona, Orlando, Florida, USA.
4Genomics and Targeted Therapy, Office of Clinical Pharmacology, US Food and Drug Administration, Silver Spring, Maryland, USA.
5Institute for Health Research and Policy, University of Illinois at Chicago, Chicago, Illinois, USA.
6Personalized Medicine Program, University of Illinois at Chicago, Chicago, Illinois, USA.
7Department of Pharmacy Systems, Outcomes and Policy, University of Illinois at Chicago, Chicago, Illinois, USA.
8Center for Pharmacoepidemiology and Pharmacoeconomic Research, University of Illinois at Chicago, Chicago, Illinois, USA.
Journal: Clinical Pharmacology and Therapeutics

Date of e-pub: May 2017

Abstract: Achieving therapeutic anticoagulation efficiently with warfarin is important to reduce thrombotic and bleeding risks and is influenced by genotype. Utilizing data from a diverse population of 257 patients who received VKORC1 and CYP2C9 genotype-guided warfarin dosing, we aimed to examine genotype-associated differences in anticoagulation endpoints and derive a novel pharmacogenetic nomogram to more optimally dose warfarin. We observed significant differences across patients with 0, 1, or ≥2 reduced-function VKORC1 or CYP2C9 alleles, respectively, in time to achieve therapeutic international normalized ratio (INR) (7.8 ± 5.8, 7.2 ± 4.7, and 5.4 ± 4.6 days, P = 0.0004) and mean percentage of time in therapeutic range in the first 28 days (22.2, 27.8, and 32.2%, P = 0.0127) with use of existing pharmacogenetic algorithms. These data suggest that more aggressive dosing is necessary for patients with 0 to 1 VKORC1/CYP2C9 variants to more efficiently achieve therapeutic anticoagulation. Herein, we provide a novel kinetic/pharmacodynamic-derived dosing nomogram optimized for a heterogeneous patient population.

 

 

Prevalence of Escherichia coli O157:H7 From House Flies (Diptera: Muscidae) and Dairy Samples in North Central Florida1.

Author information: Burrus RG1, Hogsette JA2, Kaufman PE3, Maruniak JE3, Simonne AH4, Mai V5,6.

1Department of Viral and Rickettsial Diseases, Infectious Diseases Directorate, Navy Medical Research Center, Silver Spring, MD 20910 (roxanne.g.burrus.mil@mail.mil).
2United States Department of Agriculture, Agricultural Research Service, Center for Medical, Agricultural and Veterinary Entomology, Gainesville, FL 32608 (jerry.hogsette@ars.usda.gov).
3Entomology and Nematology Department, University of Florida, Gainesville, FL 32611 (pkaufman@ufl.edu; marun@ufl.edu).
4Department of Family, Youth, and Community Sciences, University of Florida, Gainesville, FL 32611 (asim@ufl.edu).
5Department of Epidemiology, University of Florida, Gainesville, FL 32611 ( vmai@ufl.edu ).
6Emerging Pathogens Institute, University of Florida, Gainesville, FL 32611.
Journal: Journal of Medical Entomology

Date of e-pub: May 2017

Abstract:  La detección de Escherichia coli O157:H7 en las lecherías es importante para mejorar la seguridad de los productos lácteos, y se ha llevado a cabo principalmente mediante el aislamiento de las bacterias a partir de las muestras de estiércol. Sin embargo, los componentes biliares presentes en el estiércol complica la identificación genética utilizando la técnica del PCR, y el aislamiento microbiológico se dificulta por la presencia de bacterias competidoras que comparten características microbiológicas similares. El aislamiento de E. coli O157:H7 a partir de la mosca doméstica evita las dificultades asociadas con el estiércol del ganado. El aislamiento de patógenos a partir de las moscas domésticas proporciona información adicional sobre el potencial impacto epidemiológico de la dispersión de la mosca doméstica en la distribución de patógenos, ya que las moscas domésticas se dispersan desde las lecherías donde la E. coli O157:H7 existe en forma endémica en el ganado. En este estudio, se encontró que las moscas domésticas son 2,6 veces más sensibles para la detección de E. coli O157:H7 en las lecherías. Las moscas son más fáciles de capturar y manejar que el estiércol, y deberían ser utilizadas en cualquier ensayo para detectar E. coli O157:H7 en las lecherías y otros establecimientos.

 

 

Advances in the development of aptamer drug conjugates for targeted drug delivery.

Author information: Chen K1, Liu B1, Yu B1, Zhong W1, Lu Y1, Zhang J1, Liao J1, Liu J1,2,3,4,5,6, Pu Y1, Qiu L2,3,4,5,6, Zhang L2,3,4,5,6,7, Liu H1, Tan W2,3,4,5,6,7.

1Xiangya Hospital, Central South University, Changsha, China.
2Molecular Science and Biomedicine Laboratory, Hunan University, Changsha, China.
3State Key Laboratory for Chemo/Bio-Sensing and Chemometrics, Hunan University, Changsha, China.
4College of Chemistry and Chemical Engineering, Hunan University, Changsha, China.
5College of Biology, Hunan University, Changsha, China.
6Collaborative Research Center of Molecular Engineering for Theranostics, Hunan University, Changsha, China.
7Department of Chemistry and Department of Physiology and Functional Genomics, Center for Research at the Bio/Nano Interface, UF Health Cancer Center, University of Florida, Gainesville, FL, USA.
Journal: Wiley Interdisciplinary Reviews. Nanomedicine and Nanobiotechnology

Date of e-pub: May 2017

Abstract:  A key goal of modern medicine is target-specific therapeutic intervention. However, most drugs lack selectivity, resulting in ‘off-target’ side effects. To address the requirements of ‘targeted therapy,’ aptamers, which are artificial oligonucleotides, have been used as novel targeting ligands to construct aptamer drug conjugates (ApDC) that can specifically bind to a broad spectrum of targets, including diseased cells. Accordingly, the application of aptamers in targeted drug delivery has attracted broad interest due to their impressive selectivity and affinity, low immunogenicity, easy synthesis with high reproducibility, facile modification, and relatively rapid tissue penetration with no toxicity. Functionally, aptamers themselves can be used as macromolecular drugs, and they are also commonly used in biomarker discovery and targeted drug delivery. In this review, we will highlight the most recent advances in the development of aptamers and aptamer conjugates, and discuss their potential in targeted therapy. WIREs Nanomed Nanobiotechnol 2017, 9:e1438. doi: 10.1002/wnan.1438 For further resources related to this article, please visit the WIREs website.

 

 

Electrophysiologic features of fibular neuropathy in childhood and adolescence.

Author information: Karakis I1,2,3, Khoshnoodi M3,4, Liew W1,5, Nguyen ES1, Jones HR1,2, Darras BT1, Kang PB1,6.

1Department of Neurology, Boston Children’s Hospital, Harvard Medical School, Boston, Massachusetts, USA.
2Department of Neurology, Lahey Clinic, Burlington, Massachusetts, USA.
3Department of Neurology, Emory University School of Medicine, Atlanta, Georgia, USA.
4Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
5Department of Neurology, KK Women’s & Children’s Hospital, Singapore.
6Division of Pediatric Neurology, University of Florida College of Medicine, PO Box 100296, Gainesville, Florida, USA, 32610.
Journal: Muscle & Nerve

Date of e-pub: May 2017

Abstract: We studied patterns of nerve injury in pediatric common fibular (peroneal) neuropathy (CFN).

A retrospective analysis was performed on data from 53 children with CFN at a pediatric electromyography laboratory.

Conduction block at the fibular head was present in 35% of patients. Deep fibular axonal loss was identified in 77%, while superficial fibular axonal loss was identified in 45%. The pathophysiology was predominantly axonal in 72%, mostly demyelinating in 6%, and mixed in 22%. Predominantly demyelinating lesions at the fibular head demonstrated sparing of the superficial fibular sensory nerve (P = 0.01, Fischer exact test). Predominantly axonal lesions had a moderate correlation between superficial and deep fibular axonal loss (Spearman r = 0.52; P = 0.0001).

There is frequent axonal and fascicular injury in pediatric CFN, similar to adults. Deep and superficial fibular nerve involvements correlate in axonal lesions, whereas superficial fibular sensory fibers are often spared in demyelinating lesions. Muscle Nerve, 2016 Muscle Nerve 55: 693-697, 2017.

 

 

Life-extending Dietary Restriction Reduces Oxidative Damage of Proteins in Grasshoppers but Does Not Alter Allocation of Ingested Nitrogen to Somatic Tissues.

Author information: Heck MJ1, Pehlivanovic M1,2, Purcell JU1,3, Hahn DA4, Hatle JD1.

1Department of Biology, University of North Florida, Jacksonville.
2Present address: Stony Brook University, New York.
3Present address: Lake Erie College of Medicine, Florida, Pennsylvania.
4Department of Entomology, University of Florida, Gainesville.
Journal: The Journals of Gerontology. Series A, Biological Sciences and Medical Sciences

Date of e-pub: May 2017

Abstract:  Dietary restriction (DR) extends life span and reduces reproduction in most animals. The disposable soma hypothesis suggests that this longevity is the result of reduced investment in reproduction and increased nutrient allocation to the soma, permitting an increase in cellular maintenance. To investigate the role of nutrient allocation upon life-extending DR, tissue-specific nitrogen allocation was tracked in grasshoppers (Romalea microptera) upon a full or restricted (60% of full) diet. In addition, carbonyl (oxidized protein) assays addressed tissue maintenance. To develop a labeled diet on which grasshoppers could thrive, hydroponically grown Romaine lettuce was enriched with 15N. This allowed quantification of nitrogen allocation upon a normal or restricted diet. There was a 50% decrease in reproductive investment upon DR. At the same time, relative allocation of 15N to the ovary did not change. Most important, relative allocation was similar between restricted and full diet grasshoppers for somatic tissues (ie, mandibular and femur muscle, dried hemolymph, gut, and fat body). Carbonyl assays of muscles, hemolymph, and gut revealed an overall reduction in protein oxidation upon DR. These data suggest that DR does not alter nutrient allocation but does reduce protein oxidation, an observation that is inconsistent with the basic predictions of the disposable soma hypothesis.
NOTE: These abstracts were retrieved from the U.S. National Library of Medicine website managed in collaboration with the U.S. National Library of Medicine

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