DOCSLIB.ORG

Functions of the COPII Gene Paralogs SEC23A and SEC23B Are Interchangeable in Vivo

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Functions of the COPII Gene Paralogs SEC23A and SEC23B Are Interchangeable in Vivo Functions of the COPII gene paralogs SEC23A and SEC23B are interchangeable in vivo Rami Khoriatya,b,1, Geoffrey G. Heskethc, Amélie Bernardd,2, Angela C. Weyande, Dattatreya Mellacheruvuf, Guojing Zhud, Mark J. Hoenerhoffg, Beth McGeed, Lesley Everettd,3, Elizabeth J. Adamsd,4, Bin Zhangh, Thomas L. Saundersi, Alexey I. Nesvizhskiif,j, Daniel J. Klionskyd, Jordan A. Shavitb,e, Anne-Claude Gingrasc,k, and David Ginsburga,d,e,l,m,1 aDepartment of Internal Medicine, University of Michigan, Ann Arbor, MI 48109; bProgram in Cellular and Molecular Biology, University of Michigan, Ann Arbor, MI 48109; cLunenfeld-Tanenbaum Research Institute, Sinai Health System, Toronto, M5G 1X5 ON, Canada; dLife Sciences Institute, University of Michigan, Ann Arbor, MI 48109; eDepartment of Pediatrics and Communicable Diseases, University of Michigan, Ann Arbor, MI 48109; fDepartment of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI 48109; gIn Vivo Animal Core, Unit for Laboratory Animal Medicine, University of Michigan, Ann Arbor, MI 48109; hGenomic Medicine Institute, Cleveland Clinic Lerner Research Institute, Cleveland, OH 44106; iTransgenic Animal Model Core, University of Michigan, Ann Arbor, MI 48109; jDepartment of Pathology, University of Michigan, Ann Arbor, MI 48109; kDepartment of Molecular Genetics, University of Toronto, Toronto, M5S 3H7 ON, Canada; lDepartment of Human Genetics, University of Michigan, Ann Arbor, MI 48109; and mHoward Hughes Medical Institute, University of Michigan, Ann Arbor, MI 48109 Contributed by David Ginsburg, June 29, 2018 (sent for review April 4, 2018; reviewed by Michael Boyce and Mitchell J. Weiss) Approximately one-third of the mammalian proteome is trans- The mammalian genome encodes multiple paralogs for ported from the endoplasmic reticulum-to-Golgi via COPII-coated SEC23, SEC24, SAR1, and SEC31. These paralogs have been vesicles. SEC23, a core component of coat protein-complex II (COPII), proposed to contribute to COPII coat diversity, facilitating dis- is encoded by two paralogous genes in vertebrates (Sec23a and tinct cargo selectivity and tissue-specific secretory repertoires. Sec23b). In humans, SEC23B deficiency results in congenital dysery- Alternative splicing has been described for SEC31 and SEC24C thropoietic anemia type-II (CDAII), while SEC23A deficiency results in (24, 25) and might additionally contribute to COPII diversity and a skeletal phenotype (with normal red blood cells). These distinct cargo selectivity. Alhough the SEC23 gene duplication is esti- clinical disorders, together with previous biochemical studies, sug- mated to have occurred ∼615 Mya, the two mammalian paralogs, GENETICS gest unique functions for SEC23A and SEC23B. Here we show in- SEC23A and SEC23B, encode proteins that are ∼85% identical distinguishable intracellular protein interactomes for human SEC23A and SEC23B, complementation of yeast Sec23 by both human and Significance murine SEC23A/B, and rescue of the lethality of sec23b deficiency in zebrafish by a sec23a-expressing transgene. We next demonstrate In humans, SEC23B deficiency results in congenital dysery- that a Sec23a coding sequence inserted into the murine Sec23b locus thropoietic anemia type II, a disease of abnormal red blood cell completely rescues the lethal SEC23B-deficient pancreatic pheno- development, while SEC23A deficiency results in cranio-lenticulo- type. We show that SEC23B is the predominantly expressed paralog sutural-dysplasia, a disease characterized by bone abnormalities in human bone marrow, but not in the mouse, with the reciprocal due to defective collagen secretion (but no red blood cell defect). pattern observed in the pancreas. Taken together, these data dem- In this study, we show that SEC23A and SEC23B overlap in onstrate an equivalent function for SEC23A/B, with evolutionary function, and that the disparate phenotypes of SEC23A/ shifts in the transcription program likely accounting for the distinct SEC23B deficiency within and across species are likely due to phenotypes of SEC23A/B deficiency within and across species, a par- evolutionary shifts in gene-expression programs, rather than adigm potentially applicable to other sets of paralogous genes. distinct functions of the SEC23 paralogs. Our studies provide These findings also suggest that enhanced erythroid expression of a rationale for increased SEC23A or SEC23B expression as a the normal SEC23A gene could offer an effective therapeutic ap- therapeutic strategy for congenital dyserythropoietic anemia proach for CDAII patients. type II or cranio-lenticulo-sutural-dysplasia, respectively. endoplasmic reticulum | vesicular transport proteins | mice | Author contributions: R.K. and D.G. designed research; R.K., G.G.H., A.B., A.C.W., G.Z., transgenic | SEC23 M.J.H., B.M., L.E., E.J.A., B.Z., T.L.S., J.A.S., and A.-C.G. performed research; A.B., D.M., B.Z., T.L.S., A.I.N., D.J.K., J.A.S., and A.-C.G. contributed new reagents/analytic tools; R.K., ∼ G.G.H., D.M., G.Z., M.J.H., A.I.N., D.J.K., J.A.S., A.-C.G., and D.G. analyzed data; and n vertebrates, 8,000 secretory proteins (cargos) are cotransla- R.K., G.G.H., A.-C.G., and D.G. wrote the paper. Itionally inserted into the endoplasmic reticulum (ER) and ’ Reviewers: M.B., Duke University Medical Center; and M.J.W., St. Jude Children s subsequently transported to the Golgi apparatus via coat protein- Research Hospital. complex II (COPII)-coated vesicles before reaching their final The authors declare no conflict of interest. destination (endosomes, lysosomes, plasma membrane, or the Published under the PNAS license. extracellular space) (1–6). SAR1 and SEC23-SEC24 heterodimers Data deposition: The mass spectrometry dataset has been assigned the MassIVE ID form the inner COPII coat, with the outer coat composed of MSV000081703 and is available for FTP download at: ftp://[email protected]. SEC13-SEC31 heterotetramers. COPII coat assembly begins edu. The dataset has been deposited in the ProteomeXchange Consortium, proteomecentral. with GTP-bound SAR1 recruiting cytoplasmic SEC23-SEC24 proteomexchange.org/cgi/GetDataset) (identifier PXD008180). heterodimers to the ER membrane (7, 8), through direct in- 1To whom correspondence may be addressed. Email: [email protected] or ginsburg@ teraction between SAR1 and SEC23. SAR1-SEC23-SEC24 umich.edu. – 2Present address: Laboratory of Membrane Biogenesis, University of Bordeaux, CNRS, forms the prebudding complex, which recruits cargo proteins (9 ’ – 33882 Villenave d Ornon Cédex, France. 15)eitherviadirectinteraction(3,10,1618) or via interaction 3 – Present address: Department of Ophthalmology, University of San Francisco, San Fran- with intermediary proteins (termed receptors or adaptors) (3, 18 cisco, CA 94143. 22). Following cargo selection, SEC13-SEC31 heterotetramers are 4Present address: Human Oncology and Pathology Program, Memorial Sloan Kettering recruited, forming the outer COPII coat. SEC23 functions as a Cancer Center, New York, NY 10065. GTPase activating protein stimulating SAR1-GTP hydrolysis, a This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. process required for vesicle fission in vivo (4, 7). The SEC23 1073/pnas.1805784115/-/DCSupplemental. GTPase activity is enhanced by SEC31 (23). www.pnas.org/cgi/doi/10.1073/pnas.1805784115 PNAS Latest Articles | 1of10 Downloaded by guest on September 28, 2021 at the amino acid sequence level. The sequence identity between creases in the remaining paralog were shown to be post- the mouse and human orthologs for SEC23A and SEC23B are translational, likely due to enhanced stability of SEC23/ ∼98% and ∼97%, respectively. SEC24 heterodimers (29). Mutations in human SEC23A result in cranio-lenticulo-sutural Previous reports suggest unique functions for SEC23A and dysplasia (CLSD), an autosomal recessive disease characterized SEC23B (29, 34–37), potentially due to distinct sets of cargo by abnormal closure of cranial fontanelles, dysmorphic facial proteins. Additionally, human SEC23A but not SEC23B was features, skeletal abnormalities, and sutural cataracts (26, 27), reported to complement yeast SEC23 (35), consistent with dis- features thought to result from defects in collagen secretion. In tinct function of the two mammalian paralogs. contrast, loss of SEC23B function results in congenital dysery- In surprising contrast to these earlier findings, we now re- thropoietic anemia type II (CDAII), an autosomal recessive port indistinguishable intracellular interactomes for human disease characterized by anemia and increased bi/multinucleated SEC23A and SEC23B and also show complementation of yeast erythroid precursors in the bone marrow (BM) (28). In mice, germline Sec23a deletion results in an open neural SEC23 by both human and murine SEC23A/B paralogs. We sec23b tube defect and compromised extraembryonic membranes, further show that the lethality resulting from disruption sec23a likely due to impaired collagen secretion, a phenotype remi- in zebrafish can be rescued by a -expressing transgene. niscent of human CLSD (29). In contrast, mice with germline Finally, we demonstrate that the Sec23a coding sequence SEC23B deficiency die perinatally, exhibiting massive pancre- inserted into the endogenous murine Sec23b locus fully rescues atic degeneration (30–32) with no anemia or other features of the mortality and severe pancreatic phenotype of SEC23B- CDAII (30, 33). In both instances, partially compensatory in- deficiency in the mouse. AB CD EF Fig. 1.
Load more

Recommended publications
  • 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]
  • A Case Report of Congenital Erythropoietic Anemia II in China with a Novel Mutation
    Annals of Hematology https://doi.org/10.1007/s00277-019-03612-2 LETTER TO THE EDITOR A case report of congenital erythropoietic anemia II in China with a novel mutation Hong Zhang1 & Wuqing Wan1 & Xiaoyan Liu1 & Chuan Wen1 & Ying Liu1 & Senlin Luo1 & Xiao Sun1 & Shizhe Liu1 Received: 19 December 2018 /Accepted: 4 January 2019 # The Author(s) 2019 Dear Editor, 53.9 μmol/L (normal, 0–21), of which 42.7 μmol/L was Congenital erythropoietic anemias (CDAs) are a indirect (normal, 0–19). G6PD deficiency was not found. group of rare inherited diseases [1]. So far, the CDAs Red blood cell folate and hemoglobin electrophoresis are mainly divided into four types (type I to type IV), gave results within normal limits. Serum vitamin B12 and the CDA type II is the most common type. It is was 736 pmol/L (normal, 133–675). Serum iron, ferritin, caused by a mutation in the SEC23B gene. To date, 67 and transferrin were all within normal limits. Erythrocyte causative mutations in the SEC23B gene have been de- osmotic fragility test was normal. Acidified glycerol he- scribed [2–5] (the complete mutational spectrum of molysis test and Coombs test were negative. Light micro- SEC23B isshowninTable1). scope observation of a bone marrow smear revealed hy- We report a patient with typical clinical manifesta- perplasia and binucleated late erythroblasts (Fig. 1a). tions and laboratory findings, a 6-year-old girl who Genetic testing of the proband, her little brother, and hadsufferedjaundiceattheageof6monthswithlow her parents performed at Shanghai Xin Peijing Medical hemoglobin levels at 80 g/L.
    [Show full text]
  • Sec24a Antibody A
    C 0 2 - t Sec24A Antibody a e r o t S Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) 8 7 Web: [email protected] 6 www.cellsignal.com 9 # 3 Trask Lane Danvers Massachusetts 01923 USA For Research Use Only. Not For Use In Diagnostic Procedures. Applications: Reactivity: Sensitivity: MW (kDa): Source: UniProt ID: Entrez-Gene Id: WB, IP H Mk Endogenous 120 Rabbit O95486 10802 Product Usage Information 8. Merte, J. et al. (2010) Nat Cell Biol 12, 41-6; sup pp 1-8. 9. Wansleeben, C. et al. (2010) Development 137, 1067-73. Application Dilution Western Blotting 1:1000 Immunoprecipitation 1:50 Storage Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody. Specificity / Sensitivity Sec24A Antibody detects endogenous levels of total Sec24A protein. This antibody does not cross-react with other members of Sec24 family. Species Reactivity: Human, Monkey Source / Purification Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Phe52 of human Sec24A protein. Antibodies are purified by protein A and peptide affinity chromatography. Background Coat Protein Complex II (COPII) is composed of five cytosolic proteins: Sec23/24 complex, Sec13/31 complex, and Sar1. COPII coat is located at the ER/Golgi interface and is involved in transport of newly synthesized proteins from the ER to the Golgi apparatus (1). COPII formation is initiated through the binding of the activated G protein, Sar1, to the Sec23/24 complex, thereby forming a prebudding complex that directly binds target molecules (1-3).
    [Show full text]
  • Murine SEC24D Can Substitute Functionally for SEC24C in Vivo
    bioRxiv preprint doi: https://doi.org/10.1101/284398; this version posted March 22, 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. Functional Overlap Between Mouse SEC24C and SEC24D Murine SEC24D Can Substitute Functionally for SEC24C in vivo Elizabeth J. Adams1,2, Rami Khoriaty2,3, Anna Kiseleva1, Audrey C. A. Cleuren1,7, Kärt Tomberg1,4, Martijn A. van der Ent3, Peter Gergics4, K. Sue O’Shea5, Thomas L. Saunders3, David Ginsburg1-4,6,7 1From the Life Sciences Institute, 2Program in Cellular and Molecular Biology, 3Department of Internal Medicine, 4Departement of Human Genetics, 5Department of Cell and Developmental Biology, 6Department of Pediatrics and the 7Howard Hughes Medical Institute, University of Michigan, Ann Arbor, MI 48109 ABSTRACT The COPII component SEC24 mediates the recruitment of transmembrane cargoes or cargo adaptors into newly forming COPII vesicles on the ER membrane. Mammalian genomes encode four Sec24 paralogs (Sec24a-d), with two subfamilies based on sequence homology (SEC24A/B and C/D), though little is known about their comparative functions and cargo-specificities. Complete deficiency for Sec24d results in very early embryonic lethality in mice (before the 8 cell stage), with later embryonic lethality (E 7.5) observed in Sec24c null mice. To test the potential overlap in function between SEC24C/D, we employed dual recombinase mediated cassette exchange to generate a Sec24cc-d allele, in which the C-terminal 90% of SEC24C has been replaced by SEC24D coding sequence. In contrast to the embryonic lethality at E7.5 of SEC24C-deficiency, Sec24cc-d/c-d pups survive to term, though dying shortly after birth.
    [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]
  • Genetic Drivers of Pancreatic Islet Function
    | INVESTIGATION Genetic Drivers of Pancreatic Islet Function Mark P. Keller,*,1 Daniel M. Gatti,†,1 Kathryn L. Schueler,* Mary E. Rabaglia,* Donnie S. Stapleton,* Petr Simecek,† Matthew Vincent,† Sadie Allen,‡ Aimee Teo Broman,§ Rhonda Bacher,§ Christina Kendziorski,§ Karl W. Broman,§ Brian S. Yandell,** Gary A. Churchill,†,2 and Alan D. Attie*,2 *Department of Biochemistry, §Department of Biostatistics and Medical Informatics, and **Department of Horticulture, University of Wisconsin–Madison, Wisconsin 53706-1544, †The Jackson Laboratory, Bar Harbor, Maine 06409, and ‡Maine School of Science and Mathematics, Limestone, Maine 06409, ORCID IDs: 0000-0002-7405-5552 (M.P.K.); 0000-0002-4914-6671 (K.W.B.); 0000-0001-9190-9284 (G.A.C.); 0000-0002-0568-2261 (A.D.A.) ABSTRACT The majority of gene loci that have been associated with type 2 diabetes play a role in pancreatic islet function. To evaluate the role of islet gene expression in the etiology of diabetes, we sensitized a genetically diverse mouse population with a Western diet high in fat (45% kcal) and sucrose (34%) and carried out genome-wide association mapping of diabetes-related phenotypes. We quantified mRNA abundance in the islets and identified 18,820 expression QTL. We applied mediation analysis to identify candidate causal driver genes at loci that affect the abundance of numerous transcripts. These include two genes previously associated with monogenic diabetes (PDX1 and HNF4A), as well as three genes with nominal association with diabetes-related traits in humans (FAM83E, IL6ST, and SAT2). We grouped transcripts into gene modules and mapped regulatory loci for modules enriched with transcripts specific for a-cells, and another specific for d-cells.
    [Show full text]
  • Association of Gene Ontology Categories with Decay Rate for Hepg2 Experiments These Tables Show Details for All Gene Ontology Categories
    Supplementary Table 1: Association of Gene Ontology Categories with Decay Rate for HepG2 Experiments These tables show details for all Gene Ontology categories. Inferences for manual classification scheme shown at the bottom. Those categories used in Figure 1A are highlighted in bold. Standard Deviations are shown in parentheses. P-values less than 1E-20 are indicated with a "0". Rate r (hour^-1) Half-life < 2hr. Decay % GO Number Category Name Probe Sets Group Non-Group Distribution p-value In-Group Non-Group Representation p-value GO:0006350 transcription 1523 0.221 (0.009) 0.127 (0.002) FASTER 0 13.1 (0.4) 4.5 (0.1) OVER 0 GO:0006351 transcription, DNA-dependent 1498 0.220 (0.009) 0.127 (0.002) FASTER 0 13.0 (0.4) 4.5 (0.1) OVER 0 GO:0006355 regulation of transcription, DNA-dependent 1163 0.230 (0.011) 0.128 (0.002) FASTER 5.00E-21 14.2 (0.5) 4.6 (0.1) OVER 0 GO:0006366 transcription from Pol II promoter 845 0.225 (0.012) 0.130 (0.002) FASTER 1.88E-14 13.0 (0.5) 4.8 (0.1) OVER 0 GO:0006139 nucleobase, nucleoside, nucleotide and nucleic acid metabolism3004 0.173 (0.006) 0.127 (0.002) FASTER 1.28E-12 8.4 (0.2) 4.5 (0.1) OVER 0 GO:0006357 regulation of transcription from Pol II promoter 487 0.231 (0.016) 0.132 (0.002) FASTER 6.05E-10 13.5 (0.6) 4.9 (0.1) OVER 0 GO:0008283 cell proliferation 625 0.189 (0.014) 0.132 (0.002) FASTER 1.95E-05 10.1 (0.6) 5.0 (0.1) OVER 1.50E-20 GO:0006513 monoubiquitination 36 0.305 (0.049) 0.134 (0.002) FASTER 2.69E-04 25.4 (4.4) 5.1 (0.1) OVER 2.04E-06 GO:0007050 cell cycle arrest 57 0.311 (0.054) 0.133 (0.002)
    [Show full text]
  • Congenital Dyserythropoietic Anemia Type II
    Punzo et al. Orphanet Journal of Rare Diseases 2011, 6:89 http://www.ojrd.com/content/6/1/89 RESEARCH Open Access Congenital Dyserythropoietic Anemia Type II: molecular analysis and expression of the SEC23B Gene Francesca Punzo1,2, Aida M Bertoli-Avella1, Saverio Scianguetta2, Fulvio Della Ragione3, Maddalena Casale2, Luisa Ronzoni4, Maria D Cappellini4, Gianluca Forni5, Ben A Oostra1 and Silverio Perrotta2* Abstract Background: Congenital dyserythropoietic anemia type II (CDAII), the most common form of CDA, is an autosomal recessive condition. CDAII diagnosis is based on invasive, expensive, and time consuming tests that are available only in specialized laboratories. The recent identification of SEC23B mutations as the cause of CDAII opens new possibilities for the molecular diagnosis of the disease. The aim of this study was to characterize molecular genomic SEC23B defects in 16 unrelated patients affected by CDAII and correlate the identified genetic alterations with SEC23B transcript and protein levels in erythroid precursors. Methods: SEC23B was sequenced in 16 patients, their relatives and 100 control participants. SEC23B transcript level were studied by quantitative PCR (qPCR) in peripheral erythroid precursors and lymphocytes from the patients and healthy control participants. Sec23B protein content was analyzed by immunoblotting in samples of erythroblast cells from CDAII patients and healthy controls. Results: All of the investigated cases carried SEC23B mutations on both alleles, with the exception of two patients in which a single heterozygous mutation was found. We identified 15 different SEC23B mutations, of which four represent novel mutations: p.Gln214Stop, p.Thr485Ala, p.Val637Gly, and p.Ser727Phe. The CDAII patients exhibited a 40-60% decrease of SEC23B mRNA levels in erythroid precursors when compared with the corresponding cell type from healthy participants.
    [Show full text]
  • SEC24A Identified As an Essential Mediator of Thapsigargin-Induced
    Chidawanyika et al. Cell Death Discovery (2018) 4:115 https://doi.org/10.1038/s41420-018-0135-5 Cell Death Discovery ARTICLE Open Access SEC24A identified as an essential mediator of thapsigargin-induced cell death in a genome-wide CRISPR/Cas9 screen Tamutenda Chidawanyika1, Elizabeth Sergison 2, Michael Cole2,3, Kenneth Mark1 and Surachai Supattapone1,4 Abstract Endoplasmic reticulum (ER) stress from accumulated misfolded proteins in the ER can activate the unfolded protein response (UPR). The UPR acts either to restore proteostasis or to activate cell death pathways if the stress cannot be resolved. The key downstream effectors in these pathways have been studied extensively. However, in comparison, stressor-specific key mediators are not as well characterized. In this study, we sought to identify and compare the genes that are necessary for cell death induced by three classic pharmacological ER stressors with different mechanisms of action: thapsigargin, tunicamycin, and brefeldin A. We conducted genome-wide CRISPR/Cas9-based loss-of-function screens against these agents in HAP1 cells, which are a near-haploid cell line. Our screens confirmed that MFSD2A and ARF4, which were identified in previous screens, are necessary for tunicamycin- and brefeldin A- induced cytotoxicity, respectively. We identified a novel gene, SEC24A, as an essential gene for thapsigargin-induced cytotoxicity in HAP1 cells. Further experiments showed that the ability of SEC24A to facilitate ER stress-induced cell death is specific to thapsigargin and that SEC24A acts upstream of the UPR. These findings show that the genes required for ER stress-induced cell death are specific to the agent used to induce ER stress and that the resident ER cargo receptor protein SEC24A is an essential mediator of thapsigargin-induced UPR and cell death.
    [Show full text]
  • Consequences of Mutations in the Genes of the ER Export Machinery COPII in Vertebrates
    Biomedical Sciences Publications Biomedical Sciences 1-22-2020 Consequences of mutations in the genes of the ER export machinery COPII in vertebrates Chung-Ling Lu Iowa State University, [email protected] Jinoh Kim Iowa State University, [email protected] Follow this and additional works at: https://lib.dr.iastate.edu/bms_pubs Part of the Cellular and Molecular Physiology Commons, Molecular Biology Commons, and the Molecular Genetics Commons The complete bibliographic information for this item can be found at https://lib.dr.iastate.edu/ bms_pubs/81. For information on how to cite this item, please visit http://lib.dr.iastate.edu/ howtocite.html. This Article is brought to you for free and open access by the Biomedical Sciences at Iowa State University Digital Repository. It has been accepted for inclusion in Biomedical Sciences Publications by an authorized administrator of Iowa State University Digital Repository. For more information, please contact [email protected]. Consequences of mutations in the genes of the ER export machinery COPII in vertebrates Abstract Coat protein complex II (COPII) plays an essential role in the export of cargo molecules such as secretory proteins, membrane proteins, and lipids from the endoplasmic reticulum (ER). In yeast, the COPII machinery is critical for cell viability as most COPII knockout mutants fail to survive. In mice and fish, homozygous knockout mutants of most COPII genes are embryonic lethal, reflecting the essentiality of the COPII machinery in the early stages of vertebrate development. In humans, COPII mutations, which are often hypomorphic, cause diseases having distinct clinical features. This is interesting as the fundamental cellular defect of these diseases, that is, failure of ER export, is similar.
    [Show full text]
  • Mutations in the Coat Complex II Component SEC23B
    Yang et al. Cell Death and Disease (2020) 11:157 https://doi.org/10.1038/s41419-020-2358-7 Cell Death & Disease ARTICLE Open Access Mutations in the coat complex II component SEC23B promote colorectal cancer metastasis Chunyuan Yang1,NanChen2,XiangLi1, Dan Lu3,ZhiyuanHou3,YuhuaLi3,YanJin3,JinGu2 and Yuxin Yin1,3 Abstract Metastasis is the leading cause of death for colorectal cancer (CRC). However, the protein transport process involved in CRC metastasis remains unclear. In this report, we use whole-exome sequencing and bioinformatics analysis to identify somatic mutations in CRC samples and found mutations of the protein transport gene Sec23 homolog B (SEC23B)in patients with metachronous liver metastasis. We show that deletion of SEC23B suppresses the membrane localization of adhesion proteins and augments cell mobility. SEC23B mutations either cause a premature stop (C649T) or impair its protein transport activity (C1467G and T488C + G791A + G2153A). Furthermore, SEC23B mutations inhibit the transport of epithelial cell adhesion molecule (EPCAM) and CD9 molecule, thereby attenuating cell adhesion and promoting invasiveness both in vitro and in vivo. Taken together, these data demonstrate the important impact of SEC23B mutations on metastasis, and we propose that SEC23B is a potential suppressor of CRC metastasis. Introduction pathway consists of a set of organelles and cargo-bearing Colorectal cancer (CRC) is one of the most prevalent vesicles10,11. The first step of this pathway is the transport 1234567890():,; 1234567890():,; 1234567890():,; 1234567890():,; cancers worldwide. According to the World Health of protein from endoplasmic reticulum (ER) to Golgi Organization (WHO), nearly 1.1 million individuals are apparatus, mediated by the coat protein II (COPII) com- diagnosed with CRC each year1.
    [Show full text]
  • Stx5-Mediated ER-Golgi Transport in Mammals and Yeast
    cells Review Stx5-Mediated ER-Golgi Transport in Mammals and Yeast Peter TA Linders 1 , Chiel van der Horst 1, Martin ter Beest 1 and Geert van den Bogaart 1,2,* 1 Tumor Immunology Lab, Radboud University Medical Center, Radboud Institute for Molecular Life Sciences, Geert Grooteplein 28, 6525 GA Nijmegen, The Netherlands 2 Department of Molecular Immunology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands * Correspondence: [email protected]; Tel.: +31-50-36-35230 Received: 8 July 2019; Accepted: 25 July 2019; Published: 26 July 2019 Abstract: The soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) syntaxin 5 (Stx5) in mammals and its ortholog Sed5p in Saccharomyces cerevisiae mediate anterograde and retrograde endoplasmic reticulum (ER)-Golgi trafficking. Stx5 and Sed5p are structurally highly conserved and are both regulated by interactions with other ER-Golgi SNARE proteins, the Sec1/Munc18-like protein Scfd1/Sly1p and the membrane tethering complexes COG, p115, and GM130. Despite these similarities, yeast Sed5p and mammalian Stx5 are differently recruited to COPII-coated vesicles, and Stx5 interacts with the microtubular cytoskeleton, whereas Sed5p does not. In this review, we argue that these different Stx5 interactions contribute to structural differences in ER-Golgi transport between mammalian and yeast cells. Insight into the function of Stx5 is important given its essential role in the secretory pathway of eukaryotic cells and its involvement in infections and neurodegenerative diseases. Keywords: syntaxin 5; Golgi apparatus; endoplasmic reticulum; membrane trafficking; secretory pathway 1. Introduction The secretory pathway is essential for secretion of cytokines, hormones, growth factors, and extracellular matrix proteins, as well as for the delivery of receptors and transporters to the cell membrane and lytic proteins to endo-lysosomal compartments.
    [Show full text]