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PMID: 30996093 Published · epublish English Journal Article Research Support, Non-U.S. Gov't

A Genome-Wide Haploid Genetic Screen Identifies Heparan Sulfate-Associated Genes and the Macropinocytosis Modulator TMED10 as Factors Supporting Vaccinia Virus Infection.

Journal of virology ·Vol. 93 ·No. 13 ·2019-00-01

Luteijn RD, van Diemen F, Blomen VA, Boer IGJ, Manikam Sadasivam S, van Kuppevelt TH, Drexler I, Brummelkamp TR, Lebbink RJ, Wiertz EJ

Abstract

Vaccinia virus is a promising viral vaccine and gene delivery candidate and has historically been used as a model to study poxvirus-host cell interactions. We employed a genome-wide insertional mutagenesis approach in human haploid cells to identify host factors crucial for vaccinia virus infection. A library of mutagenized HAP1 cells was exposed to modified vaccinia virus Ankara (MVA). Deep-sequencing analysis of virus-resistant cells identified host factors involved in heparan sulfate synthesis, Golgi organization, and vesicular protein trafficking. We validated EXT1, TM9SF2, and TMED10 (TMP21/p23/p24δ) as important host factors for vaccinia virus infection. The critical roles of EXT1 in heparan sulfate synthesis and vaccinia virus infection were confirmed. TM9SF2 was validated as a player mediating heparan sulfate expression, explaining its contribution to vaccinia virus infection. In addition, TMED10 was found to be crucial for virus-induced plasma membrane blebbing and phosphatidylserine-induced macropinocytosis, presumably by regulating the cell surface expression of the TAM receptor Axl.IMPORTANCE Poxviruses are large DNA viruses that can infect a wide range of host species. A number of these viruses are clinically important to humans, including variola virus (smallpox) and vaccinia virus. Since the eradication of smallpox, zoonotic infections with monkeypox virus and cowpox virus are emerging. Additionally, poxviruses can be engineered to specifically target cancer cells and are used as a vaccine vector against tuberculosis, influenza, and coronaviruses. Poxviruses rely on host factors for most stages of their life cycle, including attachment to the cell and entry. These host factors are crucial for virus infectivity and host cell tropism. We used a genome-wide knockout library of host cells to identify host factors necessary for vaccinia virus infection. We confirm a dominant role for heparin sulfate in mediating virus attachment. Additionally, we show that TMED10, previously not implicated in virus infections, facilitates virus uptake by modulating the cellular response to phosphatidylserine.

Keywords
TMED10 genome-wide screen heparan sulfate macropinocytosis phosphatidylserine poxvirus vaccinia virus
MeSH Terms
CRISPR-Cas Systems Cell Line, Tumor Cowpox virus/genetics DNA Viruses Gene Knockout Techniques Genetic Testing Golgi Apparatus HEK293 Cells Haploidy HeLa Cells Heparitin Sulfate/genetics,isolation & purification,metabolism Host Specificity Host-Pathogen Interactions Humans Membrane Proteins Monkeypox virus/genetics N-Acetylglucosaminyltransferases Phosphatidylserines/metabolism Pinocytosis/physiology Poxviridae/genetics Vaccinia/virology Vaccinia virus/genetics,metabolism Vesicular Transport Proteins/metabolism Virus Attachment
Chemicals
Membrane Proteins Phosphatidylserines TM9SF2 protein, human Vesicular Transport Proteins Heparitin Sulfate N-Acetylglucosaminyltransferases exostosin-1
Authors & Affiliations
10 authors, click to expand affiliations / ORCID
Luteijn Rutger D
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
van Diemen Ferdy
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
Blomen Vincent A
Netherlands Cancer Institute, Amsterdam, The Netherlands.
Boer Ingrid G J
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
Manikam Sadasivam Saravanan
Department of Membrane Biochemistry and Biophysics, Utrecht University, Utrecht, The Netherlands.
van Kuppevelt Toin H
Department of Biochemistry, Radboud University Medical Center, Nijmegen, The Netherlands.
Drexler Ingo
Institute for Virology, Universitätsklinikum Düsseldorf, Heinrich Heine University, Düsseldorf, Germany.
Brummelkamp Thijn R
Netherlands Cancer Institute, Amsterdam, The Netherlands.
Lebbink Robert Jan
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
Wiertz Emmanuel J
Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands [email protected].
References (68)
68 references, click to expand
  1. Vaccinia virus envelope H3L protein binds to cell surface heparan sulfate and is important for intracellular mature virion morphogenesis and virus infection in vitro and in vivo.
    J Virol. 2000 Apr;74(7):3353-65 PMID: 10708453
  2. Chimaerins, novel non-protein kinase C phorbol ester receptors, associate with Tmp21-I (p23): evidence for a novel anchoring mechanism involving the chimaerin C1 domain.
    J Biol Chem. 2002 Feb 8;277(6):4541-50 PMID: 11689559
  3. UDP-glucuronate decarboxylase, a key enzyme in proteoglycan synthesis: cloning, characterization, and localization.
    J Biol Chem. 2002 May 10;277(19):16968-75 PMID: 11877387
  4. The block in assembly of modified vaccinia virus Ankara in HeLa cells reveals new insights into vaccinia virus morphogenesis.
    J Virol. 2002 Aug;76(16):8318-34 PMID: 12134037
  5. Identification of vaccinia virus epitope-specific HLA-A*0201-restricted T cells and comparative analysis of smallpox vaccines.
    Proc Natl Acad Sci U S A. 2003 Jan 7;100(1):217-22 PMID: 12518065
  6. Molecular cloning and identification of 3'-phosphoadenosine 5'-phosphosulfate transporter.
    J Biol Chem. 2003 Jul 11;278(28):25958-63 PMID: 12716889
  7. Synergistic control of cellular adhesion by transmembrane 9 proteins.
    Mol Biol Cell. 2003 Jul;14(7):2890-9 PMID: 12857872
  8. Human monkeypox: an emerging zoonosis.
    Lancet Infect Dis. 2004 Jan;4(1):15-25 PMID: 14720564
  9. Immunogenicity of a highly attenuated MVA smallpox vaccine and protection against monkeypox.
    Nature. 2004 Mar 11;428(6979):182-5 PMID: 15014500
  10. Construction and isolation of recombinant MVA.
    Methods Mol Biol. 2004;269:77-100 PMID: 15114009
  11. Entry of the vaccinia virus intracellular mature virion and its interactions with glycosaminoglycans.
    J Gen Virol. 2005 May;86(Pt 5):1279-90 PMID: 15831938
  12. Smallpox vaccine: the good, the bad, and the ugly.
    Clin Med Res. 2003 Apr;1(2):87-92 PMID: 15931293
  13. Tyro3 family-mediated cell entry of Ebola and Marburg viruses.
    J Virol. 2006 Oct;80(20):10109-16 PMID: 17005688
  14. Vaccinia virus 4c (A26L) protein on intracellular mature virus binds to the extracellular cellular matrix laminin.
    J Virol. 2007 Mar;81(5):2149-57 PMID: 17166913
  15. Quantitative analysis of brain and spinach leaf lipids employing silicic acid column chromatography and acetone for elution of glycolipids.
    Lipids. 1967 Jan;2(1):37-40 PMID: 17805719
  16. Cowpox virus infection: an emerging health threat.
    Curr Opin Infect Dis. 2008 Apr;21(2):153-6 PMID: 18317038
  17. Vaccinia virus uses macropinocytosis and apoptotic mimicry to enter host cells.
    Science. 2008 Apr 25;320(5875):531-5 PMID: 18436786
  18. The luminal domain of p23 (Tmp21) plays a critical role in p23 cell surface trafficking.
    Traffic. 2008 Sep;9(9):1530-50 PMID: 18627576
  19. TM9SF4 is required for Drosophila cellular immunity via cell adhesion and phagocytosis.
    J Cell Sci. 2008 Oct 15;121(Pt 20):3325-34 PMID: 18796536
  20. Vaccinia virus exhibits cell-type-dependent entry characteristics.
    Virology. 2009 Mar 15;385(2):383-91 PMID: 19162290
  21. Virus entry by macropinocytosis.
    Nat Cell Biol. 2009 May;11(5):510-20 PMID: 19404330
  22. Vaccinia virus strain differences in cell attachment and entry.
    Virology. 2009 Jun 20;389(1-2):132-40 PMID: 19428041
  23. Appraising the apoptotic mimicry model and the role of phospholipids for poxvirus entry.
    Proc Natl Acad Sci U S A. 2009 Oct 13;106(41):17517-21 PMID: 19805093
  24. p23/Tmp21 differentially targets the Rac-GAP beta2-chimaerin and protein kinase C via their C1 domains.
    Mol Biol Cell. 2010 Apr 15;21(8):1398-408 PMID: 20164256
  25. Vaccinia virus strains use distinct forms of macropinocytosis for host-cell entry.
    Proc Natl Acad Sci U S A. 2010 May 18;107(20):9346-51 PMID: 20439710
  26. Simultaneous high-resolution analysis of vaccinia virus and host cell transcriptomes by deep RNA sequencing.
    Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11513-8 PMID: 20534518
  27. Vaccinia virus A25 and A26 proteins are fusion suppressors for mature virions and determine strain-specific virus entry pathways into HeLa, CHO-K1, and L cells.
    J Virol. 2010 Sep;84(17):8422-32 PMID: 20538855
  28. Genetic associations of variants in genes encoding HIV-dependency factors required for HIV-1 infection.
    J Infect Dis. 2010 Dec 15;202(12):1836-45 PMID: 21083371
  29. The soluble serum protein Gas6 bridges virion envelope phosphatidylserine to the TAM receptor tyrosine kinase Axl to mediate viral entry.
    Cell Host Microbe. 2011 Apr 21;9(4):286-98 PMID: 21501828
  30. The remaining smallpox stocks: the healthiest outcome.
    Lancet. 2012 Jan 7;379(9810):10-2 PMID: 21601271
  31. Tmp21, a novel MHC-I interacting protein, preferentially binds to Β2-microglobulin-free MHC-I heavy chains.
    BMB Rep. 2011 Jun;44(6):369-74 PMID: 21699748
  32. Expression profiling of the intermediate and late stages of poxvirus replication.
    J Virol. 2011 Oct;85(19):9899-908 PMID: 21795349
  33. Ebola virus entry requires the cholesterol transporter Niemann-Pick C1.
    Nature. 2011 Aug 24;477(7364):340-3 PMID: 21866103
  34. Poxvirus host cell entry.
    Curr Opin Virol. 2012 Feb;2(1):20-7 PMID: 22440962
  35. Poxvirus cell entry: how many proteins does it take?
    Viruses. 2012 May;4(5):688-707 PMID: 22754644
  36. Direct identification of ligand-receptor interactions on living cells and tissues.
    Nat Biotechnol. 2012 Oct;30(10):997-1001 PMID: 22983091
  37. Orthopoxvirus species and strain differences in cell entry.
    Virology. 2012 Nov 25;433(2):506-12 PMID: 22999097
  38. Heparan sulfate biosynthesis: regulation and variability.
    J Histochem Cytochem. 2012 Dec;60(12):898-907 PMID: 23042481
  39. The enterohemorrhagic Escherichia coli effector protein NleF binds mammalian Tmp21.
    Vet Microbiol. 2013 May 31;164(1-2):164-70 PMID: 23434013
  40. Deciphering the glycosylome of dystroglycanopathies using haploid screens for lassa virus entry.
    Science. 2013 Apr 26;340(6131):479-83 PMID: 23519211
  41. Recombinant MVA vaccines: dispelling the myths.
    Vaccine. 2013 Sep 6;31(39):4247-51 PMID: 23523407
  42. Poxvirus DNA replication.
    Cold Spring Harb Perspect Biol. 2013 Sep 01;5(9):null PMID: 23838441
  43. Viruses PLAY DEAD to TAMe interferon responses.
    Cell Host Microbe. 2013 Aug 14;14(2):117-8 PMID: 23954149
  44. Enveloped viruses disable innate immune responses in dendritic cells by direct activation of TAM receptors.
    Cell Host Microbe. 2013 Aug 14;14(2):136-47 PMID: 23954153
  45. Human immunodeficiency virus and heparan sulfate: from attachment to entry inhibition.
    Front Immunol. 2013 Nov 20;4:385 PMID: 24312095
  46. A high-coverage shRNA screen identifies TMEM129 as an E3 ligase involved in ER-associated protein degradation.
    Nat Commun. 2014 May 08;5:3832 PMID: 24807418
  47. Virus entry. Lassa virus entry requires a trigger-induced receptor switch.
    Science. 2014 Jun 27;344(6191):1506-10 PMID: 24970085
  48. Modified vaccinia virus ankara (MVA) as production platform for vaccines against influenza and other viral respiratory diseases.
    Viruses. 2014 Jul 17;6(7):2735-61 PMID: 25036462
  49. A single immunization with modified vaccinia virus Ankara-based influenza virus H7 vaccine affords protection in the influenza A(H7N9) pneumonia ferret model.
    J Infect Dis. 2015 Mar 1;211(5):791-800 PMID: 25246535
  50. Safety and immunogenicity of a modified-vaccinia-virus-Ankara-based influenza A H5N1 vaccine: a randomised, double-blind phase 1/2a clinical trial.
    Lancet Infect Dis. 2014 Dec;14(12):1196-207 PMID: 25455987
  51. Human infection with a zoonotic orthopoxvirus in the country of Georgia.
    N Engl J Med. 2015 Mar 26;372(13):1223-30 PMID: 25806914
  52. Viral apoptotic mimicry.
    Nat Rev Microbiol. 2015 Aug;13(8):461-9 PMID: 26052667
  53. Gene essentiality and synthetic lethality in haploid human cells.
    Science. 2015 Nov 27;350(6264):1092-6 PMID: 26472760
  54. A Haploid Genetic Screen Identifies Heparan Sulfate Proteoglycans Supporting Rift Valley Fever Virus Infection.
    J Virol. 2015 Nov 18;90(3):1414-23 PMID: 26581979
  55. An orthopoxvirus-based vaccine reduces virus excretion after MERS-CoV infection in dromedary camels.
    Science. 2016 Jan 1;351(6268):77-81 PMID: 26678878
  56. An essential receptor for adeno-associated virus infection.
    Nature. 2016 Feb 4;530(7588):108-12 PMID: 26814968
  57. Bridging the Gap between Glycosylation and Vesicle Traffic.
    Front Cell Dev Biol. 2016 Mar 08;4:15 PMID: 27014691
  58. Genome-Wide Screening Uncovers the Significance of N-Sulfation of Heparan Sulfate as a Host Cell Factor for Chikungunya Virus Infection.
    J Virol. 2017 Jun 9;91(13): PMID: 28404855
  59. Cytomegalovirus gp40/m152 Uses TMED10 as ER Anchor to Retain MHC Class I.
    Cell Rep. 2018 Jun 5;23(10):3068-3077 PMID: 29874591
  60. Tmp21 and p24A, two type I proteins enriched in pancreatic microsomal membranes, are members of a protein family involved in vesicular trafficking.
    J Biol Chem. 1996 Jul 19;271(29):17183-9 PMID: 8663407
  61. A major transmembrane protein of Golgi-derived COPI-coated vesicles involved in coatomer binding.
    J Cell Biol. 1996 Dec;135(5):1239-48 PMID: 8947548
  62. Highly attenuated modified vaccinia virus Ankara (MVA) as an effective recombinant vector: a murine tumor model.
    Vaccine. 1997 Mar;15(4):387-94 PMID: 9141209
  63. p23, a major COPI-vesicle membrane protein, constitutively cycles through the early secretory pathway.
    Proc Natl Acad Sci U S A. 1997 Oct 14;94(21):11393-8 PMID: 9326620
  64. Immunosuppressive effects of apoptotic cells.
    Nature. 1997 Nov 27;390(6658):350-1 PMID: 9389474
  65. A27L protein mediates vaccinia virus interaction with cell surface heparan sulfate.
    J Virol. 1998 Feb;72(2):1577-85 PMID: 9445060
  66. Macrophages that have ingested apoptotic cells in vitro inhibit proinflammatory cytokine production through autocrine/paracrine mechanisms involving TGF-beta, PGE2, and PAF.
    J Clin Invest. 1998 Feb 15;101(4):890-8 PMID: 9466984
  67. gp25L/emp24/p24 protein family members of the cis-Golgi network bind both COP I and II coatomer.
    J Cell Biol. 1998 Feb 23;140(4):751-65 PMID: 9472029
  68. Molecular cloning and characterization of the human and mouse UDP-glucose dehydrogenase genes.
    J Biol Chem. 1998 Sep 25;273(39):25117-24 PMID: 9737970
Article Info
Journal
Journal of virology
Abbr.
J Virol
ISSN
1098-5514
DOI
10.1128/JVI.02160-18
Email
[email protected]
Published
2019-00-01
Epub
2019-00-14
Language
English
Region
United States
NLM ID
0113724
PMCID
PMC6580964
Subset
IM
External Links
PubMed source DOI: 10.1128/JVI.02160-18 PMC Full Text View journal details
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