Fluorescence Lifetime Imaging Microscopy Reveals Rerouting Of
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VAMP3 and VAMP8 Regulate the Development and Functionality of 5 Parasitophorous Vacuoles Housing Leishmania Amazonensis
bioRxiv preprint doi: https://doi.org/10.1101/2020.07.09.195032; this version posted July 9, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 2 3 4 VAMP3 and VAMP8 regulate the development and functionality of 5 parasitophorous vacuoles housing Leishmania amazonensis 6 7 8 Olivier Séguin1, Linh Thuy Mai1, Sidney W. Whiteheart2, Simona Stäger1, Albert Descoteaux1* 9 10 11 12 1Institut national de la recherche scientifique, Centre Armand-Frappier Santé Biotechnologie, 13 Laval, Québec, Canada 14 15 2Department of Molecular and Cellular Biochemistry, University of Kentucky College of 16 Medicine, Lexington, Kentucky, United States of America 17 18 19 *Corresponding author: 20 E-mail: [email protected] 21 22 23 24 Short title: SNAREs and Leishmania-harboring communal parasitophorous vacuoles 25 bioRxiv preprint doi: https://doi.org/10.1101/2020.07.09.195032; this version posted July 9, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 26 ABSTRACT 27 28 To colonize mammalian phagocytic cells, the parasite Leishmania remodels phagosomes into 29 parasitophorous vacuoles that can be either tight-fitting individual or communal. The molecular 30 and cellular bases underlying the biogenesis and functionality of these two types of vacuoles are 31 poorly understood. -
Differential Requirement of NPHP1 for Compartmentalized Protein Localization During
bioRxiv preprint doi: https://doi.org/10.1101/2021.01.20.427412; this version posted January 20, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY 4.0 International license. 1 Differential requirement of NPHP1 for compartmentalized protein localization during 2 photoreceptor outer segment development and maintenance 3 4 Poppy Dattaa,b,#, J. Thomas Cribbsa,b,#, and Seongjin Seoa,b,* 5 6 aDepartment of Ophthalmology and Visual Sciences, The University of Iowa Carver College of Medicine, Iowa 7 City, IA 52242, U.S.A. 8 bInstitute for Vision Research, The University of Iowa, Iowa City, IA 52242, U.S.A. 9 10 11 #Equally contributed 12 *To whom correspondence should be addressed: 13 Seongjin Seo, PhD 14 Department of Ophthalmology and Visual Sciences 15 University of Iowa College of Medicine 16 375 Newton Rd 17 3181D MERF 18 Iowa City, IA 52242, U.S.A. 19 Phone: 1-319-353-4477 20 Email: [email protected] 21 22 Running title: Roles of NPHP1 in photoreceptors 23 Keywords: cilia; ciliary gate; compartmentalization; inherited retinal degeneration; nonsense-associated altered 24 splicing; protein confinement; transition zone 1 bioRxiv preprint doi: https://doi.org/10.1101/2021.01.20.427412; this version posted January 20, 2021. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. -
Directed Proximity-Dependent Biotin Labelling in Zebrafish
TOOLS AND RESOURCES In vivo proteomic mapping through GFP- directed proximity-dependent biotin labelling in zebrafish Zherui Xiong1, Harriet P Lo1, Kerrie-Ann McMahon1, Nick Martel1, Alun Jones1, Michelle M Hill2, Robert G Parton1,3*, Thomas E Hall1* 1Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia; 2QIMR Berghofer Medical Research Institute, Herston, Australia; 3Centre for Microscopy and Microanalysis, The University of Queensland, Brisbane, Australia Abstract Protein interaction networks are crucial for complex cellular processes. However, the elucidation of protein interactions occurring within highly specialised cells and tissues is challenging. Here, we describe the development, and application, of a new method for proximity- dependent biotin labelling in whole zebrafish. Using a conditionally stabilised GFP-binding nanobody to target a biotin ligase to GFP-labelled proteins of interest, we show tissue-specific proteomic profiling using existing GFP-tagged transgenic zebrafish lines. We demonstrate the applicability of this approach, termed BLITZ (Biotin Labelling In Tagged Zebrafish), in diverse cell types such as neurons and vascular endothelial cells. We applied this methodology to identify interactors of caveolar coat protein, cavins, in skeletal muscle. Using this system, we defined specific interaction networks within in vivo muscle cells for the closely related but functionally distinct Cavin4 and Cavin1 proteins. *For correspondence: Introduction [email protected] (RGP); [email protected] (TEH) The understanding of the biological functions of a protein requires detailed knowledge of the mole- cules with which it interacts. However, robust elucidation of interacting proteins, including not only Competing interests: The strong direct protein-protein interactions, but also weak, transient or indirect interactions is challeng- authors declare that no ing. -
A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated. -
Defining the Kv2.1–Syntaxin Molecular Interaction Identifies a First-In-Class Small Molecule Neuroprotectant
Defining the Kv2.1–syntaxin molecular interaction identifies a first-in-class small molecule neuroprotectant Chung-Yang Yeha,b,1, Zhaofeng Yec,d,1, Aubin Moutale, Shivani Gaura,b, Amanda M. Hentonf,g, Stylianos Kouvarosf,g, Jami L. Salomana, Karen A. Hartnett-Scotta,b, Thanos Tzounopoulosa,f,g, Rajesh Khannae, Elias Aizenmana,b,g,2, and Carlos J. Camachoc,2 aDepartment of Neurobiology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; bPittsburgh Institute for Neurodegenerative Diseases, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; cDepartment of Computational and Systems Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; dSchool of Medicine, Tsinghua University, Beijing 100871, China; eDepartment of Pharmacology, College of Medicine, University of Arizona, Tucson, AZ 85724; fDepartment of Otolaryngology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; and gPittsburgh Hearing Research Center, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261 Edited by Lily Yeh Jan, University of California, San Francisco, CA, and approved June 19, 2019 (received for review February 27, 2019) + The neuronal cell death-promoting loss of cytoplasmic K follow- (13). The Kv2.1-dependent cell death pathway is normally initiated ing injury is mediated by an increase in Kv2.1 potassium channels in by the oxidative liberation of zinc from intracellular metal-binding the plasma membrane. This phenomenon relies on Kv2.1 binding to proteins (14), leading to the sequential phosphorylation of syntaxin 1A via 9 amino acids within the channel intrinsically disor- Kv2.1 residues Y124 and S800 by Src and p38 kinases, respectively dered C terminus. Preventing this interaction with a cell and blood- (15–17). -
Supplementary Figures 1-14 and Supplementary References
SUPPORTING INFORMATION Spatial Cross-Talk Between Oxidative Stress and DNA Replication in Human Fibroblasts Marko Radulovic,1,2 Noor O Baqader,1 Kai Stoeber,3† and Jasminka Godovac-Zimmermann1* 1Division of Medicine, University College London, Center for Nephrology, Royal Free Campus, Rowland Hill Street, London, NW3 2PF, UK. 2Insitute of Oncology and Radiology, Pasterova 14, 11000 Belgrade, Serbia 3Research Department of Pathology and UCL Cancer Institute, Rockefeller Building, University College London, University Street, London WC1E 6JJ, UK †Present Address: Shionogi Europe, 33 Kingsway, Holborn, London WC2B 6UF, UK TABLE OF CONTENTS 1. Supplementary Figures 1-14 and Supplementary References. Figure S-1. Network and joint spatial razor plot for 18 enzymes of glycolysis and the pentose phosphate shunt. Figure S-2. Correlation of SILAC ratios between OXS and OAC for proteins assigned to the SAME class. Figure S-3. Overlap matrix (r = 1) for groups of CORUM complexes containing 19 proteins of the 49-set. Figure S-4. Joint spatial razor plots for the Nop56p complex and FIB-associated complex involved in ribosome biogenesis. Figure S-5. Analysis of the response of emerin nuclear envelope complexes to OXS and OAC. Figure S-6. Joint spatial razor plots for the CCT protein folding complex, ATP synthase and V-Type ATPase. Figure S-7. Joint spatial razor plots showing changes in subcellular abundance and compartmental distribution for proteins annotated by GO to nucleocytoplasmic transport (GO:0006913). Figure S-8. Joint spatial razor plots showing changes in subcellular abundance and compartmental distribution for proteins annotated to endocytosis (GO:0006897). Figure S-9. Joint spatial razor plots for 401-set proteins annotated by GO to small GTPase mediated signal transduction (GO:0007264) and/or GTPase activity (GO:0003924). -
Uncoupling the Functions of CALM in VAMP Sorting and Clathrin-Coated Pit Formation
Uncoupling the Functions of CALM in VAMP Sorting and Clathrin-Coated Pit Formation Daniela A. Sahlender¤a, Patrycja Kozik¤b, Sharon E. Miller, Andrew A. Peden¤c, Margaret S. Robinson* Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom Abstract CALM (clathrin assembly lymphoid myeloid leukemia protein) is a cargo-selective adaptor for the post-Golgi R-SNAREs VAMPs 2, 3, and 8, and it also regulates the size of clathrin-coated pits and vesicles at the plasma membrane. The present study has two objectives: to determine whether CALM can sort additional VAMPs, and to investigate whether VAMP sorting contributes to CALM-dependent vesicle size regulation. Using a flow cytometry-based endocytosis efficiency assay, we demonstrate that CALM is also able to sort VAMPs 4 and 7, even though they have sorting signals for other clathrin adaptors. CALM homologues are present in nearly every eukaryote, suggesting that the CALM family may have evolved as adaptors for retrieving all post-Golgi VAMPs from the plasma membrane. Using a knockdown/rescue system, we show that wild-type CALM restores normal VAMP sorting in CALM-depleted cells, but that two non-VAMP-binding mutants do not. However, when we assayed the effect of CALM depletion on coated pit morphology, using a fluorescence microscopy- based assay, we found that the two mutants were as effective as wild-type CALM. Thus, we can uncouple the sorting function of CALM from its structural role. Citation: Sahlender DA, Kozik P, Miller SE, Peden AA, Robinson MS (2013) Uncoupling the Functions of CALM in VAMP Sorting and Clathrin-Coated Pit Formation. -
STX Stainless Steel Boxes Characteristics Enclosure and Door Manufactured from AISI 304 Stainless Steel (AISI 316 on Request)
STX stainless steel boxes characteristics Enclosure and door manufactured from AISI 304 stainless steel (AISI 316 on request). Mounting plate manufactured from 2.5mm sendzimir sheet steel. Hinge in stainless steel. composition Box complete with: • mounting plate • locking system body in zinc alloy and lever in stainless steel with Ø 3mm double bar key • package with hardware for earth connection and screws to mounting plate. conformity and approval protection degree • IP 65 complying with EN50298; EN60529 for box with single blank door • IP 55 complying with EN50298; EN60529 for box with double blank door • type 12, 4, 4X complying with UL508A; UL50 • impact resistance IK10 complying with EN50298; EN50102. box with single blank door code B A P C D E F weight kg mod. art. STX2 315 200 300 150 150 250 * 219 6 STX3 415 300 400 150 250 350 215 319 9,5 STX3 420 300 400 200 250 350 215 319 11 STX4 315 400 300 150 350 250 315 219 9,5 STX4 420 400 400 200 350 350 315 319 13,5 STX4 520 400 500 200 350 450 315 419 15,5 STX4 620 400 600 200 350 550 315 519 18 STX5 520 500 500 200 450 450 415 419 18 STX5 725 500 700 250 450 650 415 619 27 STX6 420 600 400 200 550 350 315 519 17,3 STX6 620 600 600 200 550 550 515 519 24,5 STX6 625 600 600 250 550 550 515 519 27 STX6 630 600 600 300 550 550 515 519 30 STX6 820 600 800 200 550 750 515 719 31 STX6 825 600 800 250 550 750 515 719 34 STX6 830 600 800 300 550 750 515 719 37 STX6 1230 600 1200 300 550 1150 515 1119 54 STX8 830 800 800 300 750 750 715 719 48 STX8 1030 800 1000 300 750 950 715 919 58 STX8 1230 800 1200 300 750 1150 715 1119 67 * B=200 M6 studs welded only on the hinge side. -
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Human VAMP3 Suppresses Or Negatively Regulates Bax Induced Apoptosis in Yeast
biomedicines Article Human VAMP3 Suppresses or Negatively Regulates Bax Induced Apoptosis in Yeast Damilare D. Akintade 1,2,* and Bhabatosh Chaudhuri 2 1 School of Life Sciences, Medical School, University of Nottingham, Nottingham NG7 2UH, UK 2 Leicester School of Pharmacy, De Montfort University, Leicester LE1 9BH, UK; [email protected] * Correspondence: [email protected] Abstract: Apoptosis is an essential process that is regulated genetically and could lead to a serious disease condition if not well controlled. Bax is one of the main proapoptotic proteins and actively involved in programmed cell death. It has been suggested that Bax induced apoptosis in yeast could be obstructed by enhancing vesicular membrane trafficking. Plasma membrane proteins and lipid oxidation were reduced by a vesicle-associated membrane protein (VAMP) when expressed in yeast, suggesting its potential role in repairing membranes. Membrane integrity is crucial, as the loss of membrane integrity will result in the leakage of ions from mitochondria, and ultimately cell death due to overproduction of reactive oxygen species (ROS). Expression of Arabidopsis’ VAMP has been linked to antiapoptosis activity. Since plant VAMP has been associated with antiapoptotic activities, this study investigates the possible participation of human VAMP3 in blocking human Bax mediated apoptosis. Some novel genes were identified to rescue Bax’s proapoptotic effects, in a yeast-based human hippocampal cDNA library screen. VAMP3 (a gene code for proteins involved in protein secretion) gene was chosen for further study to confirm its role in inhibiting apoptosis. VAMP3 was coexpressed with a chromosomally integrated Bax gene expression cassette driven by the GAL1 promoter. -
Species Groups Distributed Across Elevational Gradients Reveal Convergent and Continuous Genetic Adaptation to High Elevations
Species groups distributed across elevational gradients reveal convergent and continuous genetic adaptation to high elevations Yan-Bo Suna,1, Ting-Ting Fua,b,1, Jie-Qiong Jina, Robert W. Murphya,c, David M. Hillisd,2, Ya-Ping Zhanga,e,f,2, and Jing Chea,e,g,2 aState Key Laboratory of Genetic Resources and Evolution, Kunming Institute of Zoology, Chinese Academy of Sciences, 650223 Kunming, China; bKunming College of Life Science, University of Chinese Academy of Sciences, 650204 Kunming, China; cCentre for Biodiversity and Conservation Biology, Royal Ontario Museum, Toronto, ON M5S 2C6, Canada; dDepartment of Integrative Biology and Biodiversity Center, University of Texas at Austin, Austin, TX 78712; eCenter for Excellence in Animal Evolution and Genetics, Chinese Academy of Sciences, 650223 Kunming, China; fState Key Laboratory for Conservation and Utilization of Bio-Resources in Yunnan, Yunnan University, 650091 Kunming, China; and gSoutheast Asia Biodiversity Research Institute, Chinese Academy of Sciences, Yezin, 05282 Nay Pyi Taw, Myanmar Contributed by David M. Hillis, September 7, 2018 (sent for review August 7, 2018; reviewed by John H. Malone and Fuwen Wei) Although many cases of genetic adaptations to high elevations Most previous studies of the genetic processes of HEA have have been reported, the processes driving these modifications and compared species or populations from high elevations above the pace of their evolution remain unclear. Many high-elevation 3,500 m with those from low elevations to identify sequence adaptations (HEAs) are thought to have arisen in situ as popula- variation and/or expression shifts in the high-elevation group (8– tions rose with growing mountains. -
Identification of a Novel Autophagy-Related Gene Signature
Identication of a novel autophagy-related gene signature for predicting metastasis and survival in patients with osteosarcoma Guangzhi Zhang Lanzhou University Second Hospital https://orcid.org/0000-0003-3193-0297 Yajun Deng Lanzhou University Second Hospital Zuolong Wu Lanzhou University Second Hospital Enhui Ren Lanzhou University Second Hospital Wenhua Yuan Lanzhou University Second Hospital Qiqi Xie ( [email protected] ) Lanzhou University Second Hospital https://orcid.org/0000-0003-4099-5287 Primary research Keywords: osteosarcoma, autophagy-related genes, signature, survival, metastasis Posted Date: March 26th, 2020 DOI: https://doi.org/10.21203/rs.3.rs-19384/v1 License: This work is licensed under a Creative Commons Attribution 4.0 International License. Read Full License Page 1/20 Abstract Background: Osteosarcoma (OS) is a bone malignant tumor that occurs in children and adolescents. Due to a lack of reliable prognostic biomarkers, the prognosis of OS patients is often uncertain. This study aimed to construct an autophagy-related gene signature to predict the prognosis of OS patients. Methods: The gene expression prole data of OS and normal muscle tissue samples were downloaded separately from the Therapeutically Applied Research To Generate Effective Treatments (TARGET) and Genotype-Tissue Expression (GTEx) databases . The differentially expressed autophagy-related genes (DEARGs) in OS and normal muscle tissue samples were screened using R software, before being subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. A protein-protein interaction (PPI) network was constructed and hub autophagy-related genes were screened. Finally, the screened autophagy-related genes were subjected to univariate Cox regression, Lasso Cox regression, survival analysis, and clinical correlation analysis.