DOCSLIB.ORG

Global-Scale Analysis of the Dynamic Transcriptional Adaptations Within Skeletal Muscle During Hypertrophic Growth

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Global-Scale Analysis of the Dynamic Transcriptional Adaptations Within Skeletal Muscle During Hypertrophic Growth University of Kentucky UKnowledge Theses and Dissertations--Physiology Physiology 2015 GLOBAL-SCALE ANALYSIS OF THE DYNAMIC TRANSCRIPTIONAL ADAPTATIONS WITHIN SKELETAL MUSCLE DURING HYPERTROPHIC GROWTH Tyler Kirby University of Kentucky, [email protected] Right click to open a feedback form in a new tab to let us know how this document benefits ou.y Recommended Citation Kirby, Tyler, "GLOBAL-SCALE ANALYSIS OF THE DYNAMIC TRANSCRIPTIONAL ADAPTATIONS WITHIN SKELETAL MUSCLE DURING HYPERTROPHIC GROWTH" (2015). Theses and Dissertations--Physiology. 22. https://uknowledge.uky.edu/physiology_etds/22 This Doctoral Dissertation is brought to you for free and open access by the Physiology at UKnowledge. It has been accepted for inclusion in Theses and Dissertations--Physiology by an authorized administrator of UKnowledge. For more information, please contact [email protected]. STUDENT AGREEMENT: I represent that my thesis or dissertation and abstract are my original work. Proper attribution has been given to all outside sources. I understand that I am solely responsible for obtaining any needed copyright permissions. I have obtained needed written permission statement(s) from the owner(s) of each third-party copyrighted matter to be included in my work, allowing electronic distribution (if such use is not permitted by the fair use doctrine) which will be submitted to UKnowledge as Additional File. I hereby grant to The University of Kentucky and its agents the irrevocable, non-exclusive, and royalty-free license to archive and make accessible my work in whole or in part in all forms of media, now or hereafter known. I agree that the document mentioned above may be made available immediately for worldwide access unless an embargo applies. I retain all other ownership rights to the copyright of my work. I also retain the right to use in future works (such as articles or books) all or part of my work. I understand that I am free to register the copyright to my work. REVIEW, APPROVAL AND ACCEPTANCE The document mentioned above has been reviewed and accepted by the student’s advisor, on behalf of the advisory committee, and by the Director of Graduate Studies (DGS), on behalf of the program; we verify that this is the final, approved version of the student’s thesis including all changes required by the advisory committee. The undersigned agree to abide by the statements above. Tyler Kirby, Student Dr. John J. McCarthy, Major Professor Dr. Bret N. Smith, Director of Graduate Studies GLOBAL-SCALE ANALYSIS OF THE DYNAMIC TRANSCRIPTIONAL ADAPTATIONS WITHIN SKELETAL MUSCLE DURING HYPERTROPHIC GROWTH ___________________________ ___________________________ A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the College of Medicine at the University of Kentucky By Tyler John Kirby Lexington, Kentucky Co-Directors: Dr. John J. McCarthy, Associate Professor of Physiology and Dr. Timothy S. McClintock, Professor of Physiology Lexington, Kentucky Copyright © Tyler John Kirby 2015 ABSTRACT OF DISSERTATION GLOBAL-SCALE ANALYSIS OF THE DYNAMIC TRANSCRIPTIONAL ADAPTATIONS WITHIN SKELETAL MUSCLE DURING HYPERTROPHIC GROWTH Skeletal muscle possesses remarkable plasticity in responses to altered mechanical load. An established murine model used to increase mechanical load on a muscle is the surgical removal of the gastrocnemius and soleus muscles, thereby placing a functional overload on the plantaris muscle. As a consequence, there is hypertrophic growth of the plantaris muscle. We used this model to study the molecular mechanisms regulating skeletal muscle hypertrophy. Aged skeletal muscle demonstrates blunted hypertrophic growth in response to functional overload. We hypothesized that an alteration in gene expression would contribute to the blunted hypertrophic response observed with aging. However, the difference in gene expression was modest, with cluster analysis showing a similar pattern of expression between the two groups. Despite ribosomal protein gene expression being higher in the aged group, ribosome biogenesis was significantly lower in aged compared with young skeletal muscle in response to the hypertrophic stimulus (50% versus 2.5-fold, respectively). The failure to fully up-regulate pre-47S ribosomal RNA (rRNA) expression in old skeletal muscle undergoing hypertrophy indicated ribosomal DNA transcription by RNA polymerase I was impaired. Contrary to our hypothesis, the findings of the study suggest that impaired ribosome biogenesis was a primary factor underlying the blunted hypertrophic response observed in old skeletal muscle rather than dramatic differences in gene expression. As it appears ribosomal biogenesis may limit muscle hypertrophy, we assessed the dynamic changes in global transcriptional output during muscle hypertrophy, as the majority of global transcription is dedicated to ribosome biogenesis during periods of rapid growth. Metabolic labeling of nascent RNA using 5-ethynyl uridine permitted the assessment of cell type specific changes in global transcription and how this transcription is distributed within the myofiber. Using this approach, we demonstrate that myofibers are the most transcriptionally active cell-type in skeletal muscle, and furthermore, myonuclei are able to dramatically upregulate global transcription during muscle hypertrophy. Interestingly, the myonuclear accretion that occurs with hypertrophy actually results in lower transcriptional output across nuclei within the muscle fiber relative to sham conditions. These findings argue against the notion that nuclear accretion in skeletal muscle is necessary to increase the transcriptional capacity of the cell in order to support a growth response. KEYWORDS: Skeletal muscle, transcription, hypertrophy, microarray, ribosomal biogenesis ______Tyler J. Kirby______ Student’s Signature ____August 23rd, 2015____ Date GLOBAL-SCALE ANALYSIS OF THE DYNAMIC TRANSCRIPTIONAL ADAPTATIONS WITHIN SKELETAL MUSCLE DURING HYPERTROPHIC GROWTH By Tyler John Kirby _____John J. McCarthy____ Co-Director of Dissertation ___Timothy S. McClintock__ Co-Director of Dissertation ______Bret N. Smith______ Director of Graduate Studies ____August 23rd, 2015_____ ACKNOWLEDGMENTS In my time at the University of Kentucky, I had the opportunity to learn from some remarkable people. First and foremost, I would like thank Dr. John McCarthy for all of the guidance that he has provided over the last 4 years. I truly could not have asked for a better mentor. His passion and work ethic is infectious, and I hope that I still approach science with his same enthusiasm when I reach a similar stage in my career. He has taught me to dream big and never to think that any question is too challenging. John treated me as a colleague rather than his student, and I’ll always consider him my mentor and friend. I was essentially co-mentored by John and Dr. Charlotte Peterson, for which I am eternally grateful. Where John is the dreamer, Char, an amazing person and scientist, taught me how to stay focused and fully develop thoughts in order to stay productive. She taught me how to effectively communicate ideas, which is a necessary aspect of being an exceptional scientist. I thank Dr. Esther Dupont- Versteegden for all of her insightful comments into my projects. She continually made me consider things from novel perspective and helped enhance my critical thinking skills. Finally, I thank Dr. Tim McClintock for providing a different outlook on my research questions and his suggestions absolutely enhanced the quality of my research. Coming from an exercise physiology background, I had a limited number of bench skills when I got to UK. Once here, I was fortunate enough to learn from extremely talented post-docs and scientists. I want to thank Dr. Chris Fry, Dr. Thomas Chaillou, Dr. Janna Jackson, Dr. Grace Walton, Dr. Sarah White and Dr. iii Kate Kosmac. Your help and guidance has been invaluable and I would not be where I am without all of your support. Additionally, Jyothi Mula taught me everything that I know about immunohistochemistry and I appreciate all of her patience while I figured it out. Sometimes our research has some tedious and monotonous aspects, so I have to thank the research staff and undergraduates who are in the trench and grinding through image analysis. Sami Michaelis, Roosil Patel, and Angel Ho, your help is vital to the success of our projects. Finally, I want to thank my amazing family for all their love and support through this process. Words cannot describe how much you mean to me. And I swear that this is my last degree! No more school, I promise. iv TABLE OF CONTENTS Acknowledgments ................................................................................................ iii List of Tables ...................................................................................................... viii List of Figures .................................................................................................... viiii Chapter One: Background Skeletal Muscle Plasticity .................................................................................. 1 Satellite Cells .................................................................................................... 1 Satellite Cell Lineage and Molecular Identifiers ................................................. 2 Satellite cells during post-natal growth .............................................................. 3 Satellite cells in adult muscle hypertrophy ........................................................
Load more

Recommended publications
  • CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding Mandy Muller, Caroline Demeret
    CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding Mandy Muller, Caroline Demeret To cite this version: Mandy Muller, Caroline Demeret. CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding. PLoS ONE, Public Library of Science, 2014, 9 (3), pp.e92581. 10.1371/journal.pone.0092581. pasteur-01971541 HAL Id: pasteur-01971541 https://hal-pasteur.archives-ouvertes.fr/pasteur-01971541 Submitted on 7 Jan 2019 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. Distributed under a Creative Commons Attribution| 4.0 International License CCHCR1 Interacts Specifically with the E2 Protein of Human Papillomavirus Type 16 on a Surface Overlapping BRD4 Binding Mandy Muller1,2¤b, Caroline Demeret1*¤a 1 Unite´ Ge´ne´tique Papillomavirus et Cancer Humain, Institut Pasteur, Paris, France, 2 Universite´ Paris Diderot, Sorbonne Paris Cite´, Cellule Pasteur, Paris, France Abstract The Human Papillomavirus E2 proteins are key regulators of the viral life cycle, whose functions are commonly mediated through protein-protein interactions with the host cell proteome. We identified an interaction between E2 and a cellular protein called CCHCR1, which proved highly specific for the HPV16 genotype, the most prevalent in HPV-associated cancers.
    [Show full text]
  • 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.
    [Show full text]
  • 4-6 Weeks Old Female C57BL/6 Mice Obtained from Jackson Labs Were Used for Cell Isolation
    Methods Mice: 4-6 weeks old female C57BL/6 mice obtained from Jackson labs were used for cell isolation. Female Foxp3-IRES-GFP reporter mice (1), backcrossed to B6/C57 background for 10 generations, were used for the isolation of naïve CD4 and naïve CD8 cells for the RNAseq experiments. The mice were housed in pathogen-free animal facility in the La Jolla Institute for Allergy and Immunology and were used according to protocols approved by the Institutional Animal Care and use Committee. Preparation of cells: Subsets of thymocytes were isolated by cell sorting as previously described (2), after cell surface staining using CD4 (GK1.5), CD8 (53-6.7), CD3ε (145- 2C11), CD24 (M1/69) (all from Biolegend). DP cells: CD4+CD8 int/hi; CD4 SP cells: CD4CD3 hi, CD24 int/lo; CD8 SP cells: CD8 int/hi CD4 CD3 hi, CD24 int/lo (Fig S2). Peripheral subsets were isolated after pooling spleen and lymph nodes. T cells were enriched by negative isolation using Dynabeads (Dynabeads untouched mouse T cells, 11413D, Invitrogen). After surface staining for CD4 (GK1.5), CD8 (53-6.7), CD62L (MEL-14), CD25 (PC61) and CD44 (IM7), naïve CD4+CD62L hiCD25-CD44lo and naïve CD8+CD62L hiCD25-CD44lo were obtained by sorting (BD FACS Aria). Additionally, for the RNAseq experiments, CD4 and CD8 naïve cells were isolated by sorting T cells from the Foxp3- IRES-GFP mice: CD4+CD62LhiCD25–CD44lo GFP(FOXP3)– and CD8+CD62LhiCD25– CD44lo GFP(FOXP3)– (antibodies were from Biolegend). In some cases, naïve CD4 cells were cultured in vitro under Th1 or Th2 polarizing conditions (3, 4).
    [Show full text]
  • Mechanisms of Synaptic Plasticity Mediated by Clathrin Adaptor-Protein Complexes 1 and 2 in Mice
    Mechanisms of synaptic plasticity mediated by Clathrin Adaptor-protein complexes 1 and 2 in mice Dissertation for the award of the degree “Doctor rerum naturalium” at the Georg-August-University Göttingen within the doctoral program “Molecular Biology of Cells” of the Georg-August University School of Science (GAUSS) Submitted by Ratnakar Mishra Born in Birpur, Bihar, India Göttingen, Germany 2019 1 Members of the Thesis Committee Prof. Dr. Peter Schu Institute for Cellular Biochemistry, (Supervisor and first referee) University Medical Center Göttingen, Germany Dr. Hans Dieter Schmitt Neurobiology, Max Planck Institute (Second referee) for Biophysical Chemistry, Göttingen, Germany Prof. Dr. med. Thomas A. Bayer Division of Molecular Psychiatry, University Medical Center, Göttingen, Germany Additional Members of the Examination Board Prof. Dr. Silvio O. Rizzoli Department of Neuro-and Sensory Physiology, University Medical Center Göttingen, Germany Dr. Roland Dosch Institute of Developmental Biochemistry, University Medical Center Göttingen, Germany Prof. Dr. med. Martin Oppermann Institute of Cellular and Molecular Immunology, University Medical Center, Göttingen, Germany Date of oral examination: 14th may 2019 2 Table of Contents List of abbreviations ................................................................................. 5 Abstract ................................................................................................... 7 Chapter 1: Introduction ............................................................................
    [Show full text]
  • Supplementary Table S4. FGA Co-Expressed Gene List in LUAD
    Supplementary Table S4. FGA co-expressed gene list in LUAD tumors Symbol R Locus Description FGG 0.919 4q28 fibrinogen gamma chain FGL1 0.635 8p22 fibrinogen-like 1 SLC7A2 0.536 8p22 solute carrier family 7 (cationic amino acid transporter, y+ system), member 2 DUSP4 0.521 8p12-p11 dual specificity phosphatase 4 HAL 0.51 12q22-q24.1histidine ammonia-lyase PDE4D 0.499 5q12 phosphodiesterase 4D, cAMP-specific FURIN 0.497 15q26.1 furin (paired basic amino acid cleaving enzyme) CPS1 0.49 2q35 carbamoyl-phosphate synthase 1, mitochondrial TESC 0.478 12q24.22 tescalcin INHA 0.465 2q35 inhibin, alpha S100P 0.461 4p16 S100 calcium binding protein P VPS37A 0.447 8p22 vacuolar protein sorting 37 homolog A (S. cerevisiae) SLC16A14 0.447 2q36.3 solute carrier family 16, member 14 PPARGC1A 0.443 4p15.1 peroxisome proliferator-activated receptor gamma, coactivator 1 alpha SIK1 0.435 21q22.3 salt-inducible kinase 1 IRS2 0.434 13q34 insulin receptor substrate 2 RND1 0.433 12q12 Rho family GTPase 1 HGD 0.433 3q13.33 homogentisate 1,2-dioxygenase PTP4A1 0.432 6q12 protein tyrosine phosphatase type IVA, member 1 C8orf4 0.428 8p11.2 chromosome 8 open reading frame 4 DDC 0.427 7p12.2 dopa decarboxylase (aromatic L-amino acid decarboxylase) TACC2 0.427 10q26 transforming, acidic coiled-coil containing protein 2 MUC13 0.422 3q21.2 mucin 13, cell surface associated C5 0.412 9q33-q34 complement component 5 NR4A2 0.412 2q22-q23 nuclear receptor subfamily 4, group A, member 2 EYS 0.411 6q12 eyes shut homolog (Drosophila) GPX2 0.406 14q24.1 glutathione peroxidase
    [Show full text]
  • Supplementary Material
    BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s) J Neurol Neurosurg Psychiatry Page 1 / 45 SUPPLEMENTARY MATERIAL Appendix A1: Neuropsychological protocol. Appendix A2: Description of the four cases at the transitional stage. Table A1: Clinical status and center proportion in each batch. Table A2: Complete output from EdgeR. Table A3: List of the putative target genes. Table A4: Complete output from DIANA-miRPath v.3. Table A5: Comparison of studies investigating miRNAs from brain samples. Figure A1: Stratified nested cross-validation. Figure A2: Expression heatmap of miRNA signature. Figure A3: Bootstrapped ROC AUC scores. Figure A4: ROC AUC scores with 100 different fold splits. Figure A5: Presymptomatic subjects probability scores. Figure A6: Heatmap of the level of enrichment in KEGG pathways. Kmetzsch V, et al. J Neurol Neurosurg Psychiatry 2021; 92:485–493. doi: 10.1136/jnnp-2020-324647 BMJ Publishing Group Limited (BMJ) disclaims all liability and responsibility arising from any reliance Supplemental material placed on this supplemental material which has been supplied by the author(s) J Neurol Neurosurg Psychiatry Appendix A1. Neuropsychological protocol The PREV-DEMALS cognitive evaluation included standardized neuropsychological tests to investigate all cognitive domains, and in particular frontal lobe functions. The scores were provided previously (Bertrand et al., 2018). Briefly, global cognitive efficiency was evaluated by means of Mini-Mental State Examination (MMSE) and Mattis Dementia Rating Scale (MDRS). Frontal executive functions were assessed with Frontal Assessment Battery (FAB), forward and backward digit spans, Trail Making Test part A and B (TMT-A and TMT-B), Wisconsin Card Sorting Test (WCST), and Symbol-Digit Modalities test.
    [Show full text]
  • Role of Stromal Caveolin-1 (CAV1) Levels in Breast Cancer Angiogenesis
    Universidad Autónoma de Madrid Programa de Doctorado en Biociencias Moleculares Role of stromal Caveolin-1 (CAV1) levels in breast cancer angiogenesis Alberto Díez Sánchez Madrid, 2018 0 1 Departamento de Bioquímica Facultad de Medicina Universidad Autónoma de Madrid Role of stromal Caveolin-1 (CAV1) levels in breast cancer angiogenesis Doctorando: Alberto Díez Sánchez, Licenciado en Biotecnología Director: Miguel Ángel del Pozo Barriuso, MD, PhD. Fundación Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC) Madrid, 2018 1 2 CERTIFICADO DEL DIRECTOR DE TESIS El doctor Miguel Ángel del Pozo Barriuso CERTIFICA que el doctorando Alberto Díez Sánchez ha desarrollado y concluido su trabajo de tesis doctoral “Role of stromal Caveolin-1 (CAV1) levels in breast cancer angiogenesis” bajo su supervisión, en el Centro Nacional de Investigaciones Cardiovasculares (CNIC). Y para que así conste lo firma en Madrid, a 10 de Julio de 2018, Fdo. Dr. Miguel Ángel del Pozo Barriuso Centro Nacional de Investigaciones Cardiovasculares (CNIC) 3 4 ACKNOWLEDGMENTS It is said that scientific knowledge is built on top of the shoulder of giants, in more practical terms, I consider all these people below my personal giants. First ones I encountered, were my parents and grandparents, everything I have achieved has been done on top of their previous efforts, to them I dedicate my most sincere gratitude for teaching this once lazy kid the value of effort. Next, I have to thank all those high-school teachers and university professors that during my education have been able to spark in me the sense of amazement derived from understanding how nature works.
    [Show full text]
  • Genome-Wide Identification of Genes Regulating DNA Methylation Using Genetic Anchors for Causal Inference
    bioRxiv preprint doi: https://doi.org/10.1101/823807; this version posted October 30, 2019. 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. Genome-wide identification of genes regulating DNA methylation using genetic anchors for causal inference Paul J. Hop1,2, René Luijk1, Lucia Daxinger3, Maarten van Iterson1, Koen F. Dekkers1, Rick Jansen4, BIOS Consortium5, Joyce B.J. van Meurs6, Peter A.C. ’t Hoen 7, M. Arfan Ikram8, Marleen M.J. van Greevenbroek9,10, Dorret I. Boomsma11, P. Eline Slagboom1, Jan H. Veldink2, Erik W. van Zwet12, Bastiaan T. Heijmans1* 1 Molecular Epidemiology, Department of Biomedical Data Sciences, Leiden University Medical Center, Leiden, 2333 ZC, The Netherlands 2 Department of Neurology, UMC Utrecht Brain Center, University Medical Centre Utrecht, Utrecht University, Utrecht 3584 CG, Netherlands 3 Department of Human Genetics, Leiden University Medical Center, Leiden, 2333 ZC, The Netherlands 4 Department of Psychiatry, Amsterdam UMC, Vrije Universiteit Amsterdam, Amsterdam Neuroscience, Amsterdam, 1081 HV, The Netherlands 5 Biobank-based Integrated Omics Study Consortium. For a complete list of authors, see the acknowledgements. 6 Department of Internal Medicine, Erasmus Medical Centre, Rotterdam, The Netherlands. 7 Centre for Molecular and Biomolecular Informatics, Radboud Institute for Molecular Life Sciences, Radboud University
    [Show full text]
  • Ykt6 Membrane-To-Cytosol Cycling Regulates Exosomal Wnt Secretion
    bioRxiv preprint doi: https://doi.org/10.1101/485565; this version posted December 3, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Ykt6 membrane-to-cytosol cycling regulates exosomal Wnt secretion Karen Linnemannstöns1,2, Pradhipa Karuna1,2, Leonie Witte1,2, Jeanette Kittel1,2, Adi Danieli1,2, Denise Müller1,2, Lena Nitsch1,2, Mona Honemann-Capito1,2, Ferdinand Grawe3,4, Andreas Wodarz3,4 and Julia Christina Gross1,2* Affiliations: 1Hematology and Oncology, University Medical Center Goettingen, Goettingen, Germany. 2Developmental Biochemistry, University Medical Center Goettingen, Goettingen, Germany. 3Molecular Cell Biology, Institute I for Anatomy, University of Cologne Medical School, Cologne, Germany 4Cluster of Excellence-Cellular Stress Response in Aging-Associated Diseases (CECAD), Cologne, Germany *Correspondence: Dr. Julia Christina Gross, Hematology and Oncology/Developmental Biochemistry, University Medical Center Goettingen, Justus-von-Liebig Weg 11, 37077 Goettingen Germany Abstract Protein trafficking in the secretory pathway, for example the secretion of Wnt proteins, requires tight regulation. These ligands activate Wnt signaling pathways and are crucially involved in development and disease. Wnt is transported to the plasma membrane by its cargo receptor Evi, where Wnt/Evi complexes are endocytosed and sorted onto exosomes for long-range secretion. However, the trafficking steps within the endosomal compartment are not fully understood. The promiscuous SNARE Ykt6 folds into an auto-inhibiting conformation in the cytosol, but a portion associates with membranes by its farnesylated and palmitoylated C-terminus. Here, we demonstrate that membrane detachment of Ykt6 is essential for exosomal Wnt secretion.
    [Show full text]
  • Supporting Information Structure and Protein Interaction-Based Gene
    Supporting Information Structure and Protein Interaction-based Gene Ontology Annotations Reveal Likely Functions of Uncharacterized Proteins on Human Chromosome 17 Chengxin Zhang, Xiaoqiong Wei, Gilbert S. Omenn, and Yang Zhang Table of Contents Text S1. SPARQL query for curating the list of uPE1 proteins from neXtProt database version 2018-01-17 ................................................................................................................................... S2 Text S1. SPARQL query for curating the initial list of unannotated proteins from neXtProt database ......................................................................................................................................... S3 Text S3. Benchmark set of 100 well annotated PE1 proteins randomly selected from chromosome 17 ............................................................................................................................. S4 Table S1. Fmax of different programs for GO prediction using UniProt and neXtProt annotation as gold standard for the benchmark set of 100 chromosome 17 PE1 proteins ............................ S5 Table S2. Fmax of “structure” component and the final consensus prediction in COFACTOR for a benchmark set of 59 proteins with experimental structure covering at least 30% of target sequence ....................................................................................................................................... S6 Table S3. COFACTOR predicted functions for 66 uPE1 protein on chromosome 17 ...............
    [Show full text]
  • An Integrated Approach to Comprehensively Map the Molecular Context of Proteins
    bioRxiv preprint doi: https://doi.org/10.1101/264788; this version posted February 13, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Liu et al. An integrated approach to comprehensively map the molecular context of proteins Xiaonan Liu1,2, Kari Salokas1,2, Fitsum Tamene1,2,3 and Markku Varjosalo1,2,3* 1Institute of Biotechnology, University of Helsinki, Helsinki 00014, Finland 2Helsinki Institute of Life Science, University of Helsinki, Helsinki 00014, Finland 3Proteomics Unit, University of Helsinki, Helsinki 00014, Finland *Corresponding author Abstract: Protein-protein interactions underlie almost all cellular functions. The comprehensive mapping of these complex cellular networks of stable and transient associations has been made available by affi nity purifi cation mass spectrometry (AP-MS) and more recently by proximity based labelling methods such as BioID. Due the advancements in both methods and MS instrumentation, an in-depth analysis of the whole human proteome is at grasps. In order to facilitate this, we designed and optimized an integrated approach utilizing MAC-tag combining both AP-MS and BioID in a single construct. We systematically applied this approach to 18 subcellular localization markers and generated a molecular context database, which can be used to defi ne molecular locations for any protein of interest. In addition, we show that by combining the AP-MS and BioID results we can also obtain interaction distances within a complex. Taken together, our combined strategy off ers comprehensive approach for mapping physical and functional protein interactions. Introduction: Majority of proteins do not function in isolation geting the endogenous bait protein, allowing and their interactions with other proteins defi ne purifi cation of the bait protein together with the their cellular functions.
    [Show full text]
  • The Alopecia Areata Phenotype Is Induced by the Water Avoidance Stress Test in Cchcr1-Deficient Mice
    bioRxiv preprint doi: https://doi.org/10.1101/2020.12.16.423031; this version posted December 16, 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. The alopecia areata phenotype is induced by the water avoidance stress test in cchcr1- deficient mice. Qiao-Feng Zhao1, Nagisa Yoshihara1, Atsushi Takagi1, Etsuko Komiyama1, Akira Oka3, Shigaku Ikeda*1, 2 1Department of Dermatology and Allergology, and 2Atopy (Allergy) Research Center, Juntendo University Graduate School of Medicine, Tokyo, Japan 3The Institute of Medical Sciences, Tokai University, Kanagawa, Japan *Corresponding author: Shigaku Ikeda, MD, PhD, Department of Dermatology and Allergology, Juntendo University Graduate School of Medicine, 2-1-1 Hongo, Bunkyo- ku, Tokyo 113-8421, Japan Tel: 81-3-5802-1089, Fax: 81-3-3813-2205, E-mail: [email protected] Running title: AA phenotype induced in cchcr1-deficient mice COI: The authors have no conflicts of interest to declare. p. 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.12.16.423031; this version posted December 16, 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. ABSTRACT Background: We recently discovered a nonsynonymous variant in the coiled-coil alpha- helical rod protein 1 (CCHCR1) gene within the alopecia areata (AA) risk haplotype; mice engineered to carry the risk allele displayed a hair loss phenotype.
    [Show full text]