1887

Abstract

Seneca Valley virus (SVV, also known as Senecavirus A), an oncolytic virus, is a nonenveloped, positive-strand RNA virus and the sole member of the genus within the family . The mechanisms of SVV entry into cells are currently almost unknown. In the present study, we found that SVV entry into HEK293T cells is acidic pH-dependent by using ammonium chloride (NHCl) and chloroquine, both of which could inhibit SVV infection. We confirmed that dynamin II is required for SVV entry by using dynasore, silencing the dynamin II protein, or expressing the dominant-negative (DN) K44A mutant of dynamin II. Then, we discovered that chlorpromazine (CPZ) treatment or knockdown of the clathrin heavy chain (CLTC) protein significantly inhibited SVV infection. In addition, overexpression of CLTC promoted SVV infection. Caveolin-1 and membrane cholesterol were also required for SVV endocytosis. Notably, utilizing genistein, EIPA or nocodazole, we observed that macropinocytosis and microtubules are not involved in SVV entry. Furthermore, overexpression of the Rab7 and Rab9 proteins but not the Rab5 or Rab11 proteins promoted SVV infection. The findings were further validated by the knockdown of four Rabs and Lamp1 proteins, indicating that after internalization, SVV is transported from late endosomes to the trans-Golgi network (TGN) or lysosomes, respectively, eventually releasing its RNA into the cytosol from the lysosomes. Our findings concretely revealed SVV endocytosis mechanisms in HEK293T cells and provided an insightful theoretical foundation for further research into SVV oncolytic mechanisms.

Funding
This study was supported by the:
  • Fundamental Research Funds for the Central Universities (Award 2662017PY108)
    • Principle Award Recipient: NotApplicable
  • National Natural Science Foundation of China (Award 31772749)
    • Principle Award Recipient: NotApplicable
  • National Natural Science Foundation of China (Award 32072841)
    • Principle Award Recipient: NotApplicable
  • National Key Research and Development Program of China (Award 2022YFD1800800)
    • Principle Award Recipient: PingQian
  • National Key Research and Development Program of China (Award 2021YFD1800300)
    • Principle Award Recipient: PingQian
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/content/journal/jgv/10.1099/jgv.0.001833
2023-03-22
2024-05-01
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References

  1. Hales LM, Knowles NJ, Reddy PS, Xu L, Hay C et al. Complete genome sequence analysis of Seneca Valley virus-001, a novel oncolytic picornavirus. J Gen Virol 2008; 89:1265–1275 [View Article] [PubMed]
    [Google Scholar]
  2. Leme RA, Alfieri AF, Alfieri AA. Update on Senecavirus infection in pigs. Viruses 2017; 9:170 [View Article] [PubMed]
    [Google Scholar]
  3. Oliveira TES, Michelazzo MMZ, Fernandes T, de Oliveira AG, Leme RA et al. Histopathological, immunohistochemical, and ultrastructural evidence of spontaneous Senecavirus A-induced lesions at the choroid plexus of newborn piglets. Sci Rep 2017; 7:16555 [View Article] [PubMed]
    [Google Scholar]
  4. Reddy PS, Burroughs KD, Hales LM, Ganesh S, Jones BH et al. Seneca Valley virus, a systemically deliverable oncolytic picornavirus, and the treatment of neuroendocrine cancers. J Natl Cancer Inst 2007; 99:1623–1633 [View Article] [PubMed]
    [Google Scholar]
  5. Qian S, Fan W, Liu T, Wu M, Zhang H et al. Seneca Valley virus suppresses host type I interferon production by targeting adaptor proteins MAVS, TRIF, and TANK for cleavage. J Virol 2017; 91:16 [View Article] [PubMed]
    [Google Scholar]
  6. Wen W, Li X, Yin M, Wang H, Qin L et al. Selective autophagy receptor SQSTM1/ p62 inhibits Seneca Valley virus replication by targeting viral VP1 and VP3. Autophagy 2021; 17:3763–3775 [View Article] [PubMed]
    [Google Scholar]
  7. Zhou X, Liang W-F, Si G-B, Li J-H, Chen Z-F et al. Buffalo-origin Seneca Valley virus in China: first report, isolation, genome characterization, and evolution analysis. Front Vet Sci 2021; 8:730701 [View Article] [PubMed]
    [Google Scholar]
  8. Xue Q, Liu H, Zhu Z, Yang F, Xue Q et al. Seneca Valley virus 3C protease negatively regulates the type I interferon pathway by acting as a viral deubiquitinase. Antiviral Research 2018; 160:183–189 [View Article] [PubMed]
    [Google Scholar]
  9. Liu T, Li X, Wu M, Qin L, Chen H et al. Seneca Valley virus 2C and 3Cpro induce apoptosis via mitochondrion-mediated intrinsic pathway. Front Microbiol 2019; 10:1202 [View Article] [PubMed]
    [Google Scholar]
  10. Fernandes MHV, Maggioli MF, Otta J, Joshi LR, Lawson S et al. Senecavirus A 3C protease mediates host cell apoptosis late in infection. Front Immunol 2019; 10:363 [View Article] [PubMed]
    [Google Scholar]
  11. Wen W, Li X, Wang H, Zhao Q, Yin M et al. Seneca Valley virus 3C protease induces pyroptosis by directly cleaving porcine Gasdermin D. J Immunol 2021; 207:189–199 [View Article] [PubMed]
    [Google Scholar]
  12. Jackson T, Sheppard D, Denyer M, Blakemore W, King AM. The epithelial integrin alphavbeta6 is a receptor for foot-and-mouth disease virus. J Virol 2000; 74:4949–4956 [View Article] [PubMed]
    [Google Scholar]
  13. Hussain KM, Leong KLJ, Ng MM-L, Chu JJH. The essential role of clathrin-mediated endocytosis in the infectious entry of human enterovirus 71. J Biol Chem 2011; 286:309–321 [View Article] [PubMed]
    [Google Scholar]
  14. Kim C, Bergelson JM. Echovirus 7 entry into polarized intestinal epithelial cells requires clathrin and Rab7. mBio 2012; 3:e00304-11 [View Article] [PubMed]
    [Google Scholar]
  15. O’Donnell V, Larocco M, Baxt B. Heparan sulfate-binding foot-and-mouth disease virus enters cells via caveola-mediated endocytosis. J Virol 2008; 82:9075–9085 [View Article] [PubMed]
    [Google Scholar]
  16. Han S-C, Guo H-C, Sun S-Q, Jin Y, Wei Y-Q et al. Productive entry of foot-and-mouth disease virus via macropinocytosis independent of phosphatidylinositol 3-kinase. Sci Rep 2016; 6:19294 [View Article] [PubMed]
    [Google Scholar]
  17. Cao L, Zhang R, Liu T, Sun Z, Hu M et al. Seneca Valley virus attachment and uncoating mediated by its receptor anthrax toxin receptor 1. Proc Natl Acad Sci 2018; 115:13087–13092 [View Article] [PubMed]
    [Google Scholar]
  18. Jia M, Sun M, Tang Y-D, Zhang Y-Y, Wang H et al. Senecavirus A entry into host cells is dependent on the cholesterol-mediated endocytic pathway. Front Vet Sci 2022; 9:840655 [View Article] [PubMed]
    [Google Scholar]
  19. Hou L, Tong X, Pan Y, Shi R, Liu C et al. Seneca Valley virus enters PK-15 cells via Caveolae-mediated endocytosis and macropinocytosis dependent on low-pH, Dynamin, Rab5, and Rab7. J Virol 2022; 96:e0144622 [View Article] [PubMed]
    [Google Scholar]
  20. Giranda VL, Heinz BA, Oliveira MA, Minor I, Kim KH et al. Acid-induced structural changes in human rhinovirus 14: possible role in uncoating. Proc Natl Acad Sci 1992; 89:10213–10217 [View Article] [PubMed]
    [Google Scholar]
  21. Wang X, Peng W, Ren J, Hu Z, Xu J et al. A sensor-adaptor mechanism for enterovirus uncoating from structures of EV71. Nat Struct Mol Biol 2012; 19:424–429 [View Article] [PubMed]
    [Google Scholar]
  22. Su C, Zheng C. When Rab GTPases meet innate immune signaling pathways. Cytokine Growth Factor Rev 2021; 59:95–100 [View Article] [PubMed]
    [Google Scholar]
  23. Ren J, Wang X, Hu Z, Gao Q, Sun Y et al. Picornavirus uncoating intermediate captured in atomic detail. Nat Commun 2013; 4:1929 [View Article] [PubMed]
    [Google Scholar]
  24. Vanlandingham PA, Ceresa BP. Rab7 regulates late endocytic trafficking downstream of multivesicular body biogenesis and cargo sequestration. J Biol Chem 2009; 284:12110–12124 [View Article] [PubMed]
    [Google Scholar]
  25. Arighi CN, Hartnell LM, Aguilar RC, Haft CR, Bonifacino JS. Role of the mammalian retromer in sorting of the cation-independent mannose 6-phosphate receptor. J Cell Biol 2004; 165:123–133 [View Article] [PubMed]
    [Google Scholar]
  26. Barbero P, Bittova L, Pfeffer SR. Visualization of Rab9-mediated vesicle transport from endosomes to the trans-Golgi in living cells. J Cell Biol 2002; 156:511–518 [View Article] [PubMed]
    [Google Scholar]
  27. Wilcke M, Johannes L, Galli T, Mayau V, Goud B et al. Rab11 regulates the compartmentalization of early endosomes required for efficient transport from early endosomes to the trans-golgi network. J Cell Biol 2000; 151:1207–1220 [View Article] [PubMed]
    [Google Scholar]
  28. Clague MJ. Molecular aspects of the endocytic pathway. Biochem J 1998; 336:271–282 [View Article] [PubMed]
    [Google Scholar]
  29. Ng EL, Gan BQ, Ng F, Tang BL. Rab GTPases regulating receptor trafficking at the late endosome-lysosome membranes. Cell Biochem Funct 2012; 30:515–523 [View Article] [PubMed]
    [Google Scholar]
  30. Qian S, Fan W, Qian P, Chen H, Li X. Isolation and full-genome sequencing of Seneca Valley virus in piglets from China, 2016. Virol J 2016; 13:173 [View Article] [PubMed]
    [Google Scholar]
  31. Zhu Y-Z, Xu Q-Q, Wu D-G, Ren H, Zhao P et al. Japanese encephalitis virus enters rat neuroblastoma cells via a pH-dependent, dynamin and caveola-mediated endocytosis pathway. J Virol 2012; 86:13407–13422 [View Article] [PubMed]
    [Google Scholar]
  32. Silva MC, Guerrero-Plata A, Gilfoy FD, Garofalo RP, Mason PW. Differential activation of human monocyte-derived and plasmacytoid dendritic cells by West Nile virus generated in different host cells. J Virol 2007; 81:13640–13648 [View Article] [PubMed]
    [Google Scholar]
  33. Wen W, Zheng Z, Wang H, Zhao Q, Yin M et al. Seneca Valley virus induces DHX30 cleavage to antagonize its antiviral effects. J Virol 2022; 96:e0112122 [View Article] [PubMed]
    [Google Scholar]
  34. Liu W, Li X, Zhang H, Hao G, Shang X et al. Evaluation of immunoreactivity and protection efficacy of Seneca Valley virus inactivated vaccine in finishing pigs based on screening of inactivated agents and adjuvants. Vaccines 2022; 10:631 [View Article] [PubMed]
    [Google Scholar]
  35. Schoggins JW, Wilson SJ, Panis M, Murphy MY, Jones CT et al. A diverse range of gene products are effectors of the type I interferon antiviral response. Nature 2011; 472:481–485 [View Article] [PubMed]
    [Google Scholar]
  36. Levy HC, Bostina M, Filman DJ, Hogle JM. Catching a virus in the act of RNA release: a novel poliovirus uncoating intermediate characterized by cryo-electron microscopy. J Virol 2010; 84:4426–4441 [View Article] [PubMed]
    [Google Scholar]
  37. Marsh M, Helenius A. Virus entry: open sesame. Cell 2006; 124:729–740 [View Article] [PubMed]
    [Google Scholar]
  38. Sloan RD, Kuhl BD, Mesplède T, Münch J, Donahue DA et al. Productive entry of HIV-1 during cell-to-cell transmission via dynamin-dependent endocytosis. J Virol 2013; 87:8110–8123 [View Article] [PubMed]
    [Google Scholar]
  39. Holla P, Ahmad I, Ahmed Z, Jameel S. Hepatitis E virus enters liver cells through a dynamin-2, clathrin and membrane cholesterol-dependent pathway. Traffic 2015; 16:398–416 [View Article] [PubMed]
    [Google Scholar]
  40. Krieger SE, Kim C, Zhang L, Marjomaki V, Bergelson JM. Echovirus 1 entry into polarized Caco-2 cells depends on dynamin, cholesterol, and cellular factors associated with macropinocytosis. J Virol 2013; 87:8884–8895 [View Article] [PubMed]
    [Google Scholar]
  41. Li M, Zhang D, Li C, Zheng Z, Fu M et al. Characterization of Zika virus endocytic pathways in human glioblastoma cells. Front Microbiol 2020; 11:242 [View Article] [PubMed]
    [Google Scholar]
  42. Macia E, Ehrlich M, Massol R, Boucrot E, Brunner C et al. Dynasore, a cell-permeable inhibitor of dynamin. Dev Cell 2006; 10:839–850 [View Article] [PubMed]
    [Google Scholar]
  43. Oh P, McIntosh DP, Schnitzer JE. Dynamin at the neck of caveolae mediates their budding to form transport vesicles by GTP-driven fission from the plasma membrane of endothelium. J Cell Biol 1998; 141:101–114 [View Article] [PubMed]
    [Google Scholar]
  44. Damke H, Baba T, Warnock DE, Schmid SL. Induction of mutant dynamin specifically blocks endocytic coated vesicle formation. J Cell Biol 1994; 127:915–934 [View Article] [PubMed]
    [Google Scholar]
  45. Ferguson SM, De Camilli P. Dynamin, a membrane-remodelling GTPase. Nat Rev Mol Cell Biol 2012; 13:75–88 [View Article] [PubMed]
    [Google Scholar]
  46. Mercer J, Schelhaas M, Helenius A. Virus entry by endocytosis. Annu Rev Biochem 2010; 79:803–833 [View Article] [PubMed]
    [Google Scholar]
  47. Mercer J, Helenius A. Virus entry by macropinocytosis. Nat Cell Biol 2009; 11:510–520 [View Article] [PubMed]
    [Google Scholar]
  48. Mercer J, Helenius A. Vaccinia virus uses macropinocytosis and apoptotic mimicry to enter host cells. Science 2008; 320:531–535 [View Article] [PubMed]
    [Google Scholar]
  49. Yin X, Ambardekar C, Lu Y, Feng Z. Distinct entry mechanisms for nonenveloped and quasi-enveloped hepatitis E viruses. J Virol 2016; 90:4232–4242 [View Article] [PubMed]
    [Google Scholar]
  50. Taylor MP, Koyuncu OO, Enquist LW. Subversion of the actin cytoskeleton during viral infection. Nat Rev Microbiol 2011; 9:427–439 [View Article] [PubMed]
    [Google Scholar]
  51. Walsh D, Naghavi MH. Exploitation of cytoskeletal networks during early viral infection. Trends Microbiol 2019; 27:39–50 [View Article] [PubMed]
    [Google Scholar]
  52. Matlin KS, Reggio H, Helenius A, Simons K. Pathway of vesicular stomatitis virus entry leading to infection. J Mol Biol 1982; 156:609–631 [View Article] [PubMed]
    [Google Scholar]
  53. Lozach P-Y, Mancini R, Bitto D, Meier R, Oestereich L et al. Entry of bunyaviruses into mammalian cells. Cell Host Microbe 2010; 7:488–499 [View Article] [PubMed]
    [Google Scholar]
  54. Jackson T, Mould AP, Sheppard D, King AMQ. Integrin alphavbeta1 is a receptor for foot-and-mouth disease virus. J Virol 2002; 76:935–941 [View Article] [PubMed]
    [Google Scholar]
  55. Rahn E, Petermann P, Hsu MJ, Rixon FJ, Knebel-Mörsdorf D. Entry pathways of herpes simplex virus type 1 into human keratinocytes are dynamin- and cholesterol-dependent. PLoS One 2011; 6:e25464 [View Article] [PubMed]
    [Google Scholar]
  56. Coyne CB, Shen L, Turner JR, Bergelson JM. Coxsackievirus entry across epithelial tight junctions requires occludin and the small GTPases Rab34 and Rab5. Cell Host Microbe 2007; 2:181–192 [View Article] [PubMed]
    [Google Scholar]
  57. Huttunen M, Waris M, Kajander R, Hyypiä T, Marjomäki V. Coxsackievirus A9 infects cells via nonacidic multivesicular bodies. J Virol 2014; 88:5138–5151 [View Article] [PubMed]
    [Google Scholar]
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