Human Enterovirus Nonstructural Protein 2C Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

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Enteroviruses contain a large number of closely related human pathogens, including poliovirus, EV71, and coxsackie viruses, and cause ~3 billion infections annually. Among the nonstructural proteins of enteroviruses or picornaviruses, protein 2CATPase is the most conserved and complex but the least understood. On the basis of sequence analyses, this protein has been predicted as a putative superfamily 3 (SF3) helicase that supposedly plays a pivotal role in enteroviral RNA replication. However, attempts to determine the helicase activity associated with 2CATPase have been unsuccessful. We found that eukaryotically expressed EV71 or CAV16 2CATPase does possess an ATP-dependent RNA helicase activity that 3′→5′ unwinds RNA helices like other SF3 helicases; surprisingly, it also functions as an RNA chaperone that remodels RNA structures in an ATP-independent manner. Moreover, we determined the domain requirements for these two RNA remodeling activities associated with 2CATPase and provide both in vitro and cellular evidence of their potential roles during viral RNA replication. Additionally, our study provides the first evidence that RNA helicase and chaperoning activities can be integrated within one protein, thereby introducing an extended view of RNA remodeling proteins.

Vyšlo v časopise: Human Enterovirus Nonstructural Protein 2C Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone. PLoS Pathog 11(7): e32767. doi:10.1371/journal.ppat.1005067
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1005067

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Zdroje

1. Bleichert F, Baserga SJ. The long unwinding road of RNA helicases. Mol Cell. 2007;27(3):339–52. doi: 10.1016/j.molcel.2007.07.014 17679086.

2. Jarmoskaite I, Russell R. RNA helicase proteins as chaperones and remodelers. Annu Rev Biochem. 2014;83:697–725. doi: 10.1146/annurev-biochem-060713-035546 24635478; PubMed Central PMCID: PMC4143424.

3. Jankowsky E. RNA helicases at work: binding and rearranging. Trends Biochem Sci. 2011;36(1):19–29. doi: 10.1016/j.tibs.2010.07.008 20813532; PubMed Central PMCID: PMC3017212.

4. Grohman JK, Gorelick RJ, Lickwar CR, Lieb JD, Bower BD, Znosko BM, et al. A guanosine-centric mechanism for RNA chaperone function. Science. 2013;340(6129):190–5. doi: 10.1126/science.1230715 23470731.

5. Rajkowitsch L, Chen D, Stampfl S, Semrad K, Waldsich C, Mayer O, et al. RNA chaperones, RNA annealers and RNA helicases. RNA Biol. 2007;4(3):118–30. doi: 10.4161/rna.4.3.5445 18347437.

6. Lorsch JR. RNA chaperones exist and DEAD box proteins get a life. Cell. 2002;109(7):797–800. doi: 10.1016/S0092-8674(02)00804-8 12110176.

7. Kadare G, Haenni AL. Virus-encoded RNA helicases. J Virol. 1997;71(4):2583–90. 9060609; PubMed Central PMCID: PMC191378.

8. Musier-Forsyth K. RNA remodeling by chaperones and helicases. RNA Biol. 2010;7(6):632–3. 21173577.

9. Frick DN. The hepatitis C virus NS3 protein: a model RNA helicase and potential drug target. Current issues in molecular biology. 2007;9(1):1–20. 17263143; PubMed Central PMCID: PMCPmc3571657.

10. Karpe YA, Aher PP, Lole KS. NTPase and 5'-RNA triphosphatase activities of Chikungunya virus nsP2 protein. PLoS One. 2011;6(7):e22336. doi: 10.1371/journal.pone.0022336 21811589; PubMed Central PMCID: PMC3139623.

11. Lee NR, Kwon HM, Park K, Oh S, Jeong YJ, Kim DE. Cooperative translocation enhances the unwinding of duplex DNA by SARS coronavirus helicase nsP13. Nucleic Acids Res. 2010;38(21):7626–36. doi: 10.1093/nar/gkq647 20671029; PubMed Central PMCID: PMC2995068.

12. Wang Q, Han Y, Qiu Y, Zhang S, Tang F, Wang Y, et al. Identification and characterization of RNA duplex unwinding and ATPase activities of an alphatetravirus superfamily 1 helicase. Virology. 2012;433(2):440–8. doi: 10.1016/j.virol.2012.08.045 22995190.

13. Roos MPR. Enteroviruses: Polioviruses, Coxsackieviruses, Echoviruses, and Newer Enteroviruses. In: Knipe DMH, Peter M., editor. Fields Virology, 5th Edition. 1. Philadelphia: Lippioncott Williams & Wilkonson; 2007. p. 840–93.

14. Browne CJ, Smith RJ, Bourouiba L. From regional pulse vaccination to global disease eradication: insights from a mathematical model of poliomyelitis. J Math Biol. 2014. doi: 10.1007/s00285-014-0810-y 25074277.

15. Banerjee R, Echeverri A, Dasgupta A. Poliovirus-encoded 2C polypeptide specifically binds to the 3'-terminal sequences of viral negative-strand RNA. J Virol. 1997;71(12):9570–8. 9371621; PubMed Central PMCID: PMC230265.

16. Rodriguez PL, Carrasco L. Poliovirus protein 2C contains two regions involved in RNA binding activity. J Biol Chem. 1995;270(17):10105–12. 7730315.

17. Paul AV, Peters J, Mugavero J, Yin J, van Boom JH, Wimmer E. Biochemical and genetic studies of the VPg uridylylation reaction catalyzed by the RNA polymerase of poliovirus. J Virol. 2003;77(2):891–904. 12502805; PubMed Central PMCID: PMCPmc140777.

18. Rieder E, Paul AV, Kim DW, van Boom JH, Wimmer E. Genetic and biochemical studies of poliovirus cis-acting replication element cre in relation to VPg uridylylation. J Virol. 2000;74(22):10371–80. 11044081; PubMed Central PMCID: PMCPmc110911.

19. Barton DJ, Flanegan JB. Synchronous replication of poliovirus RNA: initiation of negative-strand RNA synthesis requires the guanidine-inhibited activity of protein 2C. J Virol. 1997;71(11):8482–9. 9343205; PubMed Central PMCID: PMC192311.

20. Teterina NL, Gorbalenya AE, Egger D, Bienz K, Ehrenfeld E. Poliovirus 2C protein determinants of membrane binding and rearrangements in mammalian cells. J Virol. 1997;71(12):8962–72. 9371552; PubMed Central PMCID: PMC230196.

21. Aldabe R, Carrasco L. Induction of membrane proliferation by poliovirus proteins 2C and 2BC. Biochem Biophys Res Commun. 1995;206(1):64–76. doi: 10.1006/bbrc.1995.1010 7818552.

22. Gladue DP, O'Donnell V, Baker-Branstetter R, Holinka LG, Pacheco JM, Fernandez-Sainz I, et al. Foot-and-mouth disease virus nonstructural protein 2C interacts with Beclin1, modulating virus replication. J Virol. 2012;86(22):12080–90. doi: 10.1128/jvi.01610-12 22933281; PubMed Central PMCID: PMC3486479.

23. Liu Y, Wang C, Mueller S, Paul AV, Wimmer E, Jiang P. Direct interaction between two viral proteins, the nonstructural protein 2C and the capsid protein VP3, is required for enterovirus morphogenesis. PLoS Pathog. 2010;6(8):e1001066. doi: 10.1371/journal.ppat.1001066 20865167; PubMed Central PMCID: PMC2928791.

24. Wang C, Jiang P, Sand C, Paul AV, Wimmer E. Alanine scanning of poliovirus 2CATPase reveals new genetic evidence that capsid protein/2CATPase interactions are essential for morphogenesis. J Virol. 2012;86(18):9964–75. doi: 10.1128/jvi.00914-12 22761387; PubMed Central PMCID: PMC3446611.

25. Zheng Z, Li H, Zhang Z, Meng J, Mao D, Bai B, et al. Enterovirus 71 2C protein inhibits TNF-alpha-mediated activation of NF-kappaB by suppressing IkappaB kinase beta phosphorylation. J Immunol. 2011;187(5):2202–12. doi: 10.4049/jimmunol.1100285 21810613.

26. Pfister T, Wimmer E. Characterization of the nucleoside triphosphatase activity of poliovirus protein 2C reveals a mechanism by which guanidine inhibits poliovirus replication. J Biol Chem. 1999;274(11):6992–7001. 10066753.

27. Rodriguez PL, Carrasco L. Poliovirus protein 2C has ATPase and GTPase activities. J Biol Chem. 1993;268(11):8105–10. 8385138.

28. Adams P, Kandiah E, Effantin G, Steven AC, Ehrenfeld E. Poliovirus 2C protein forms homo-oligomeric structures required for ATPase activity. J Biol Chem. 2009;284(33):22012–21. doi: 10.1074/jbc.M109.031807 19520852; PubMed Central PMCID: PMC2755925.

29. Tang WF, Yang SY, Wu BW, Jheng JR, Chen YL, Shih CH, et al. Reticulon 3 binds the 2C protein of enterovirus 71 and is required for viral replication. J Biol Chem. 2007;282(8):5888–98. doi: 10.1074/jbc.M611145200 17182608.

30. Cheng Z, Yang J, Xia H, Qiu Y, Wang Z, Han Y, et al. The nonstructural protein 2C of a Picorna-like virus displays nucleic acid helix destabilizing activity that can be functionally separated from its ATPase activity. J Virol. 2013;87(9):5205–18. doi: 10.1128/jvi.00245-13 23449794; PubMed Central PMCID: PMC3624285.

31. Bystroff C, Shao Y. Fully automated ab initio protein structure prediction using I-SITES, HMMSTR and ROSETTA. Bioinformatics. 2002;18 Suppl 1:S54–61. 12169531.

32. James JA, Escalante CR, Yoon-Robarts M, Edwards TA, Linden RM, Aggarwal AK. Crystal structure of the SF3 helicase from adeno-associated virus type 2. Structure. 2003;11(8):1025–35. 12906833.

33. James JA, Aggarwal AK, Linden RM, Escalante CR. Structure of adeno-associated virus type 2 Rep40-ADP complex: insight into nucleotide recognition and catalysis by superfamily 3 helicases. Proc Natl Acad Sci U S A. 2004;101(34):12455–60. doi: 10.1073/pnas.0403454101 15310852; PubMed Central PMCID: PMC515083.

34. Pang PS, Jankowsky E, Planet PJ, Pyle AM. The hepatitis C viral NS3 protein is a processive DNA helicase with cofactor enhanced RNA unwinding. EMBO J. 2002;21(5):1168–76. doi: 10.1093/emboj/21.5.1168 11867545; PubMed Central PMCID: PMC125889.

35. Singleton MR, Dillingham MS, Wigley DB. Structure and mechanism of helicases and nucleic acid translocases. Annu Rev Biochem. 2007;76:23–50. doi: 10.1146/annurev.biochem.76.052305.115300 17506634.

36. Jarmoskaite I, Russell R. DEAD-box proteins as RNA helicases and chaperones. Wiley Interdiscip Rev RNA. 2011;2(1):135–52. doi: 10.1002/wrna.50 21297876; PubMed Central PMCID: PMC3032546.

37. Frick DN, Banik S, Rypma RS. Role of divalent metal cations in ATP hydrolysis catalyzed by the hepatitis C virus NS3 helicase: magnesium provides a bridge for ATP to fuel unwinding. J Mol Biol. 2007;365(4):1017–32. doi: 10.1016/j.jmb.2006.10.023 17084859; PubMed Central PMCID: PMC1829317.

38. Lyons T, Murray KE, Roberts AW, Barton DJ. Poliovirus 5'-terminal cloverleaf RNA is required in cis for VPg uridylylation and the initiation of negative-strand RNA synthesis. J Virol. 2001;75(22):10696–708. doi: 10.1128/jvi.75.22.10696–10708.2001 11602711; PubMed Central PMCID: PMCPmc114651.

39. Steil BP, Barton DJ. Conversion of VPg into VPgpUpUOH before and during poliovirus negative-strand RNA synthesis. J Virol. 2009;83(24):12660–70. doi: 10.1128/jvi.01676-08 19812161; PubMed Central PMCID: PMCPmc2786823.

40. Yang J, Cheng Z, Zhang S, Xiong W, Xia H, Qiu Y, et al. A cypovirus VP5 displays the RNA chaperone-like activity that destabilizes RNA helices and accelerates strand annealing. Nucleic Acids Res. 2014;42(4):2538–54. doi: 10.1093/nar/gkt1256 24319147; PubMed Central PMCID: PMC3936753.

41. Smith RH, Kotin RM. The Rep52 gene product of adeno-associated virus is a DNA helicase with 3'-to-5' polarity. J Virol. 1998;72(6):4874–81. 9573254; PubMed Central PMCID: PMC110039.

42. DeStefano JJ, Titilope O. Poliovirus protein 3AB displays nucleic acid chaperone and helix-destabilizing activities. J Virol. 2006;80(4):1662–71. doi: 10.1128/jvi.80.4.1662–1671.2006 16439523; PubMed Central PMCID: PMC1367131.

43. Puerta-Fernandez E, Romero-Lopez C, Barroso-delJesus A, Berzal-Herranz A. Ribozymes: recent advances in the development of RNA tools. FEMS Microbiol Rev. 2003;27(1):75–97. 12697343.

44. Brown BA, Panganiban AT. Identification of a region of hantavirus nucleocapsid protein required for RNA chaperone activity. RNA Biol. 2010;7(6):830–7. 21378500; PubMed Central PMCID: PMC3073341.

45. Lam AM, Frick DN. Hepatitis C virus subgenomic replicon requires an active NS3 RNA helicase. J Virol. 2006;80(1):404–11. doi: 10.1128/jvi.80.1.404–411.2006 16352565; PubMed Central PMCID: PMC1317551.

46. Gong P, Peersen OB. Structural basis for active site closure by the poliovirus RNA-dependent RNA polymerase. Proc Natl Acad Sci U S A. 2010;107(52):22505–10. doi: 10.1073/pnas.1007626107 21148772; PubMed Central PMCID: PMCPmc3012486.

47. Mirzayan C, Wimmer E. Genetic analysis of an NTP-binding motif in poliovirus polypeptide 2C. Virology. 1992;189(2):547–55. 1322588.

48. Zuniga S, Sola I, Cruz JL, Enjuanes L. Role of RNA chaperones in virus replication. Virus Res. 2009;139(2):253–66. doi: 10.1016/j.virusres.2008.06.015 18675859.

49. Steil BP, Barton DJ. Cis-active RNA elements (CREs) and picornavirus RNA replication. Virus Res. 2009;139(2):240–52. doi: 10.1016/j.virusres.2008.07.027 18773930; PubMed Central PMCID: PMC2692539.

50. Paul AV, Wimmer E. Initiation of protein-primed picornavirus RNA synthesis. Virus Res. 2015. doi: 10.1016/j.virusres.2014.12.028 25592245.

51. Wimmer E, Hellen CU, Cao X. Genetics of poliovirus. Annual review of genetics. 1993;27:353–436. doi: 10.1146/annurev.ge.27.120193.002033 8122908.

52. Sweeney TR, Abaeva IS, Pestova TV, Hellen CU. The mechanism of translation initiation on Type 1 picornavirus IRESs. EMBO J. 2014;33(1):76–92. doi: 10.1002/embj.201386124 24357634; PubMed Central PMCID: PMC3990684.

53. Liu Y, Wimmer E, Paul AV. Cis-acting RNA elements in human and animal plus-strand RNA viruses. Biochim Biophys Acta. 2009;1789(9–10):495–517. doi: 10.1016/j.bbagrm.2009.09.007 19781674; PubMed Central PMCID: PMC2783963.

54. Ogram SA, Flanegan JB. Non-template functions of viral RNA in picornavirus replication. Curr Opin Virol. 2011;1(5):339–46. doi: 10.1016/j.coviro.2011.09.005 22140418; PubMed Central PMCID: PMC3227123.

55. Sharma N, O'Donnell BJ, Flanegan JB. 3'-Terminal sequence in poliovirus negative-strand templates is the primary cis-acting element required for VPgpUpU-primed positive-strand initiation. J Virol. 2005;79(6):3565–77. doi: 10.1128/jvi.79.6.3565–3577.2005 15731251; PubMed Central PMCID: PMCPmc1075688.

56. Vogt DA, Andino R. An RNA element at the 5'-end of the poliovirus genome functions as a general promoter for RNA synthesis. PLoS Pathog. 2010;6(6):e1000936. doi: 10.1371/journal.ppat.1000936 20532207; PubMed Central PMCID: PMC2880563.

57. Gangaramani DR, Eden EL, Shah M, Destefano JJ. The twenty-nine amino acid C-terminal cytoplasmic domain of poliovirus 3AB is critical for nucleic acid chaperone activity. RNA Biol. 2010;7(6):820–9. 21045553; PubMed Central PMCID: PMC3072266.

58. Tang F, Xia H, Wang P, Yang J, Zhao T, Zhang Q, et al. The identification and characterization of nucleic acid chaperone activity of human enterovirus 71 nonstructural protein 3AB. Virology. 2014;464–465:353–64. doi: 10.1016/j.virol.2014.07.037 25113906.

59. Cho MW, Richards OC, Dmitrieva TM, Agol V, Ehrenfeld E. RNA duplex unwinding activity of poliovirus RNA-dependent RNA polymerase 3Dpol. J Virol. 1993;67(6):3010–8. 8388485; PubMed Central PMCID: PMCPmc237637.

60. Henn A, Bradley MJ, De La Cruz EM. ATP utilization and RNA conformational rearrangement by DEAD-box proteins. Annu Rev Biophys. 2012;41:247–67. doi: 10.1146/annurev-biophys-050511-102243 22404686.

61. Liu F, Putnam A, Jankowsky E. ATP hydrolysis is required for DEAD-box protein recycling but not for duplex unwinding. Proc Natl Acad Sci U S A. 2008;105(51):20209–14. doi: 10.1073/pnas.0811115106 19088201; PubMed Central PMCID: PMC2629341.

62. Lu G, Gong P. Crystal Structure of the full-length Japanese encephalitis virus NS5 reveals a conserved methyltransferase-polymerase interface. PLoS Pathog. 2013;9(8):e1003549. doi: 10.1371/journal.ppat.1003549 23950717; PubMed Central PMCID: PMCPmc3738499.

63. Wang Z, Qiu Y, Liu Y, Qi N, Si J, Xia X, et al. Characterization of a nodavirus replicase revealed a de novo initiation mechanism of RNA synthesis and terminal nucleotidyltransferase activity. J Biol Chem. 2013;288(43):30785–801. doi: 10.1074/jbc.M113.492728 24019510; PubMed Central PMCID: PMC3829395.

64. Wang G, Wang HJ, Zhou H, Nian QG, Song Z, Deng YQ, et al. Hydrated silica exterior produced by biomimetic silicification confers viral vaccine heat-resistance. ACS Nano. 2015;9(1):799–808. doi: 10.1021/nn5063276 25574563.