UFGI Publications Round-Up Week 10/26/2016

The Skp1 Homologs SKR-1/2 Are Required for the Caenorhabditis elegans SKN-1 Antioxidant/Detoxification Response Independently of p38 MAPK.

Author information: Wu CW1, Deonarine A2, Przybysz A3, Strange K4, Choe KP1.

1Department of Biology and Genetics Institute, University of Florida, Gainesville, FL 32611, USA.
2Department of Cell Biology, Microbiology, and Molecular Biology, University of South Florida, Tampa, FL, 33620.
3Department of Anesthesiology, University of Michigan, Ann Arbor, MI 48109.
4The MDI Biological Laboratory, Salisbury Cove, ME 04672.

Journal: PLoS Genetics

Date of e-pub: October 2016

Abstract: SKN-1/Nrf are the primary antioxidant/detoxification response transcription factors in animals and they promote health and longevity in many contexts. SKN-1/Nrf are activated by a remarkably broad-range of natural and synthetic compounds and physiological conditions. Defining the signaling mechanisms that regulate SKN-1/Nrf activation provides insights into how cells coordinate responses to stress. Nrf2 in mammals is regulated in part by the redox sensor repressor protein named Keap1. In C. elegans, the p38 MAPK cascade in the intestine activates SKN-1 during oxidative stress by promoting its nuclear accumulation. Interestingly, we find variation in the kinetics of p38 MAPK activation and tissues with SKN-1 nuclear accumulation among different pro-oxidants that all trigger strong induction of SKN-1 target genes. Using genome-wide RNAi screening, we identify new genes that are required for activation of the core SKN-1 target gene gst-4 during exposure to the natural pro-oxidant juglone. Among 10 putative activators identified in this screen was skr-1/2, highly conserved homologs of yeast and mammalian Skp1, which function to assemble protein complexes. Silencing of skr-1/2 inhibits induction of SKN-1 dependent detoxification genes and reduces resistance to pro-oxidants without decreasing p38 MAPK activation. Global transcriptomics revealed strong correlation between genes that are regulated by SKR-1/2 and SKN-1 indicating a high degree of specificity. We also show that SKR-1/2 functions upstream of the WD40 repeat protein WDR-23, which binds to and inhibits SKN-1. Together, these results identify a novel p38 MAPK independent signaling mechanism that activates SKN-1 via SKR-1/2 and involves WDR-23.



Loss and Re-emergence of Legs in Snakes by Modular Evolution of Sonic hedgehog and HOXD Enhancers.

Author information: Leal F1, Cohn MJ2.

1Howard Hughes Medical Institute, UF Genetics Institute, University of Florida, P.O. Box 103610, University of Florida, Gainesville, FL 32610, USA; Department of Biology, UF Genetics Institute, University of Florida, P.O. Box 103610, University of Florida, Gainesville, FL 32610, USA.
2Howard Hughes Medical Institute, UF Genetics Institute, University of Florida, P.O. Box 103610, University of Florida, Gainesville, FL 32610, USA; Department of Biology, UF Genetics Institute, University of Florida, P.O. Box 103610, University of Florida, Gainesville, FL 32610, USA; Department of Molecular Genetics and Microbiology, UF Genetics Institute, University of Florida, P.O. Box 103610, University of Florida,Gainesville, FL 32610, USA. Electronic address: mjcohn@ufl.edu.

Journal: Current Biology

Date of e-pub: October 2016

Abstract: Limb reduction and loss are hallmarks of snake evolution. Although advanced snakes are completely limbless, basal and intermediate snakes retain pelvic girdles and small rudiments of the femur. Moreover, legs may have re-emerged in extinct snake lineages [1-5], suggesting that the mechanisms of limb development were not completely lost in snakes. Here we report that hindlimb development arrests in python embryos as a result of mutations that abolish essential transcription factor binding sites in the limb-specific enhancer of Sonic hedgehog (SHH). Consequently, SHH transcription is weak and transient in python hindlimb buds, leading to early termination of a genetic circuit that drives limb outgrowth. Our results suggest that degenerate evolution of the SHH limb enhancer played a role in reduction of hindlimbs during snake evolution. By contrast, HOXD digit enhancers are conserved in pythons, and HOXD gene expression in the hindlimb buds progresses to the distal phase, forming an autopodial (digit) domain. Python hindlimb buds then develop transitory pre-chondrogenic condensations of the tibia, fibula, and footplate, raising the possibility that re-emergence of hindlimbs during snake evolution did not require de novo re-evolution of lost structures but instead could have resulted from persistence of embryonic legs. VIDEO ABSTRACT.


Long-Term Clinical Response to Treatment and Maintenance of Localized Aggressive Periodontitis: A Cohort Study.

Author information: Miller KA1, Branco-de-Almeida LS1,2, Wolf S1, Hovencamp N1, Treloar T1, Harrison P1,3, Aukhil I1, Gong Y4, Shaddox LM5.

1Department of Periodontology, College of Dentistry, University of Florida, Gainesville, FL, USA.
2Department of Dentistry II, School of Dentistry, Federal University of Maranhão, São Luís, MA, Brazil.
3Division of Restorative Dentistry & Periodontology, Dublin Dental University Hospital, Trinity College Dublin, Dublin, Ireland.
4Department of Pharmacotherapy and Translational Research and Center for Pharmacogenomics, College of Pharmacy, University of Florida,Gainesville, FL, USA.
5Department of Periodontology, College of Dentistry, University of Florida, Gainesville, FL, USA. lshaddox@dental.ufl.edu.

Journal: Journal of Clinical Periodontology

Date of e-pub: October 2016

Abstract: To evaluate long-term clinical response to periodontal therapy and maintenance in localized aggressive periodontitis (LAP).

One hundred forty-one African-Americans diagnosed with LAP, aged 5-25 years, were enrolled. Patients underwent periodontal mechanical debridement plus one week of amoxicillin/metronidazole. Mechanical therapy was repeated as needed and clinical parameters were recorded at baseline, 3, 6, 12, 18, and 24 months, and 2 additional annual follow-up visits after treatment. Radiographs from primary dentition of patients with LAP in permanent dentition, and additional healthy siblings (HS) were analyzed retrospectively.

Periodontal therapy significantly improved probing depth and clinical attachment level up to 4 years (mean reductions: 2.18 ± 1.03 and 2.80 ± 1.43 mm, respectively). Percentage of affected sites was reduced at all timepoints and maintained up to 4 years. Noncompliance with antibiotics/appointments negatively affected the treatment response. Ninety percent of LAP patients in permanent dentition and 32% of HS presented radiographic bone loss in primary dentition.

Mechanical debridement with one-week of systemic antibiotics along with proper periodontal maintenance was effective in the treatment and successful maintenance of LAP for up to 4 years. LAP in permanent dentition may be preceded in the primary dentition. Clinicaltrials. gov #NCT01330719. This article is protected by copyright. All rights reserved.



Tissue, developmental, and caste-specific expression of odorant binding proteins in a eusocial insect, the red imported fire ant, Solenopsis invicta.

Author information: Zhang W1,2, Wanchoo A2, Ortiz-Urquiza A2, Xia Y1, Keyhani NO1,2.

1Genetic Engineering Research Center, School of Life Sciences, Chongqing University, Chongqing 400045, PR China.
2Department of Microbiology and Cell Science, Institute of Food and Agricultural Sciences, University of Florida, Gainesville, FL, 32611, USA.

Journal: Science Reports

Date of e-pub: October 2016

Abstract: Insects interact with the surrounding environment via chemoreception, and in social insects such as ants, chemoreception functions to mediate diverse behaviors including food acquisition, self/non-self recognition, and intraspecific communication. The invasive red imported fire ant, Solenopsis invicta, has spread worldwide, displaying a remarkable environmental adaptability. Odorant binding proteins (OBPs) are chemical compound carriers, involved in diverse physiological processes including odor detection and chemical transport. S. invicta contains a highly divergent 17-member OBP gene family, that includes an ant-specific expansion and the social organization implicated Gp-9 (OBP3) gene. A systematic gene expression analysis of the SiOBP repertoire was performed across social caste (workers, male and female alates), tissues (antennae, head, thorax, and abdomen), and developmental stages (egg, larvae, and pupae), revealing that although SiOBPs were expressed in the antennae, the major regions of expression were in the head and thorax across all castes, and the abdomen in male and female alates. SiOBPs were very highly expressed in female alates and at somewhat lower levels in male alates and workers. SiOBPs were differentially expressed, with unique signatures in various castes and tissues, suggesting functionality of SiOBPs beyond olfaction Expression patterns of SiOBP subgroups also showed relationships with their evolutionary relatedness.



Metabolomic Responses of Arabidopsis Suspension Cells to Bicarbonate under Light and Dark Conditions.

Author information: Misra BB1, Yin Z1,2, Geng S1, de Armas E3, Chen S1,4.

1Department of Biology, Genetics Institute, Plant Molecular and Cellular Biology Program, University of Florida, Gainesville, FL 32610, USA.
2Alkali 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.
3Training Institute, Thermo Fisher Scientific, 1400 North point Parkway, Ste 10., West Palm Beach, FL 33407, USA.
4Interdisciplinary Center for Biotechnology Research, University of Florida, Gainesville, FL 32610, USA.

Journal: Science Reports

Date of e-pub: October 2016

Abstract: Global CO2 level presently recorded at 400 ppm is expected to reach 550 ppm in 2050, an increment likely to impact plant growth and productivity. Using targeted LC-MS and GC-MS platforms we quantified 229 and 29 metabolites, respectively in a time-course study to reveal short-term responses to different concentrations (1, 3, and 10 mM) of bicarbonate (HCO3) under light and dark conditions. Results indicate that HCO3 treatment responsive metabolomic changes depend on the HCO3 concentration, time of treatment, and light/dark. Interestingly, 3 mM HCO3 concentration treatment induced more significantly changed metabolites than either lower or higher concentrations used. Flavonoid biosynthesis and glutathione metabolism were common to both light and dark-mediated responses in addition to showing concentration-dependent changes. Our metabolomics results provide insights into short-term plant cellular responses to elevated HCO3concentrations as a result of ambient increases in CO2 under light and dark.



High-Efficiency Transduction of Primary Human Hematopoietic Stem/Progenitor Cells by AAV6 Vectors: Strategies for Overcoming Donor-Variation and Implications in Genome Editing.

Author information: Ling C1,2, Bhukhai K3, Yin Z1,2,4, Tan M5, Yoder MC6, Leboulch P3, Payen E3, Srivastava A1,2,7.

1Division of Cellular and Molecular Therapy, Department of Pediatrics, University of Florida College of Medicine, Gainesville, FL, USA.
2Powell Gene Therapy Center; University of Florida College of Medicine, Gainesville, FL, USA.
3CEA/Paris Sud University (UMR-E 007), Institute of Emerging Diseases and Innovative Therapies (iMETI), Fontenay-aux-Roses, France.
4Department of Traditional Chinese Medicine, Second Military Medical University, Shanghai, China.
5Department of Physiology, Xiang-Ya School of Medicine, Central South University, Changsha, China.
6Herman B Well Center for Pediatrics Research, Indiana University School of Medicine, Indianapolis, IN, USA.
7Department of Molecular Genetics &Microbiology; University of Florida College of Medicine, Gainesville, FL, USA.

Journal: Science Reports

Date of e-pub: October 2016

Abstract: We have reported that of the 10 commonly used AAV serotype vectors, AAV6 is the most efficient in transducing primary human hematopoietic stem/progenitor cells (HSPCs). However, the transduction efficiency of the wild-type (WT) AAV6 vector varies greatly in HSPCs from different donors. Here we report two distinct strategies to further increase the transduction efficiency in HSPCs from donors that are transduced less efficiently with the WT AAV6 vectors. The first strategy involved modifications of the viral capsid proteins where specific surface-exposed tyrosine (Y) and threonine (T) residues were mutagenized to generate a triple-mutant (Y705 + Y731F + T492V) AAV6 vector. The second strategy involved the use of ex vivo transduction at high cell density. The combined use of these strategies resulted in transduction efficiency exceeding ~90% in HSPCs at significantly reduced vector doses. Our studies have significant implications in the optimal use of capsid-optimized AAV6 vectors in genome editing in HSPCs.



Activation of Fetal γ-globin Gene Expression via Direct Protein Delivery of Synthetic Zinc-finger DNA-Binding Domains.

Author information: Hossain MA1, Shen Y1, Knudson I1, Thakur S1, Stees JR1, Qiu Y2, Pace BS3, Peterson KR4, Bungert J1.

1Department of Biochemistry and Molecular Biology, College of Medicine, University of Florida, Gainesville, Florida, USA.
2Department of Anatomy and Cell Biology, College of Medicine, UF Health Cancer Center, Genetics Institute, University of Florida, Gainesville, Florida, USA.
3Department of Pediatrics, Augusta University, Augusta, Georgia, USA.
4Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, USA.

Journal: Molecular Therapy. Nucleic Acids.

Date of e-pub: October 2016

Abstract: Reactivation of γ-globin expression has been shown to ameliorate disease phenotypes associated with mutations in the adult β-globin gene, including sickle cell disease. Specific mutations in the promoter of the γ-globin genes are known to prevent repression of the genes in the adult and thus lead to hereditary persistence of fetal hemoglobin. One such hereditary persistence of fetal hemoglobin is associated with a sequence located 567 bp upstream of the Gγ-globin gene which assembles a GATA-containing repressor complex. We generated two synthetic zinc-finger DNA-binding domains (ZF-DBDs) targeting this sequence. The -567Gγ ZF-DBDs associated with high affinity and specificity with the target site in the γ-globin gene promoter. We delivered the -567Gγ ZF-DBDs directly to primary erythroid cells. Exposure of these cells to the recombinant -567Gγ ZF-DBDs led to increased expression of the γ-globin gene. Direct protein delivery of ZF-DBDs that compete with transcription regulatory proteins will have broad implications for modulating gene expression in analytical or therapeutic settings.

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