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Role of viral nonstructural proteins in rotavirus replication

    Francesca Arnoldi

    International Centre for Genetic Engineering & Biotechnology (ICGEB), Padriciano 99, 34012 Trieste, Italy.

    &
    Oscar R Burrone

    † Author for correspondence

    International Centre for Genetic Engineering & Biotechnology (ICGEB), Padriciano 99, 34012 Trieste, Italy.

    Published Online:https://doi.org/10.2217/17460794.4.2.185

    Studies on the molecular biology of rotavirus, the major etiologic agent of gastroenteritis in infants and young children worldwide, have so far led to a large but not exhaustive knowledge of the mechanisms by which rotavirus replicates in the host cell. While the role of rotavirus structural proteins in the replication cycle is well defined, the functions of nonstructural proteins remain poorly understood. Recent experiments of RNA interference have clearly indicated the phases of the replication cycle for which the nonstructural proteins are essentially required. In addition, biochemical studies of their interactions with other viral proteins, together with immunofluorescence experiments on cells expressing recombinant proteins in different combinations, are providing new indications of their functions. This article contains a critical collection of the most recent achievements and the current hypotheses about the roles of nonstructural proteins in virus replication.

    Papers of special note have been highlighted as: ▪ of interest ▪▪ of considerable interest

    Bibliography

    • Parashar UD, Gibson CJ, Bresse JS, Glass RI: Rotavirus and severe childhood diarrhea. Emerg. Infect. Dis.12,304–306 (2006).
    • Estes M, Kapikian A: Rotaviruses. In: Fields Virology (Volume 2, 5th Edition). Knipe DM (Ed.). Wolters Kluwer Health/Lippincott Williams & Wilkins, Philadelphia, PA, USA (2007).
    • Prasad BV, Rothnagel R, Zeng CQ et al.: Visualization of ordered genomic RNA and localization of transcriptional complexes in rotavirus. Nature382,471–473 (1996).
    • Lawton JA, Estes MK, Prasad BV: Three-dimensional visualization of mRNA release from actively transcribing rotavirus particles. Nat. Struct. Biol.4,118–121 (1997).
    • Campagna M, Eichwald C, Vascotto F, Burrone OR: RNA interference of rotavirus segment 11 mRNA reveals the essential role of NSP5 in the virus replicative cycle. J. Gen. Virol.86,1481–1487 (2005).▪▪ First demonstration using RNA interference (RNAi) of the essential role of nonstructural protein (NSP)5 in rotavirus replication.
    • Lopez T, Camacho M, Zayas M et al.: Silencing the morphogenesis of rotavirus. J. Virol.79,184–192 (2005).▪▪ First demonstration by using RNAi of the involvement of NSP4 in rotavirus morphogenesis.
    • Lopez T, Rojas M, Ayala-Breton C, Lopez S, Arias CF: Reduced expression of the rotavirus NSP5 gene has a pleiotropic effect on virus replication. J. Gen. Virol.86,1609–1617 (2005).
    • Silvestri LS, Taraporewala ZF, Patton JT: Rotavirus replication: plus-sense templates for double-stranded RNA synthesis are made in viroplasms. J. Virol.78,7763–7774 (2004).▪▪ First demonstration by RNAi that NSP2 is essentially required for rotavirus replication and that NSP1 is dispensable.
    • Silvestri LS, Tortorici MA, Vasquez-Del Carpio R, Patton JT: Rotavirus glycoprotein NSP4 is a modulator of viral transcription in the infected cell. J. Virol.79,15165–15174 (2005).▪▪ Evidence indicating the role of NSP4 in controlling the levels of viral plus-strand RNAs, maturation of viroplasms and viral progeny assembly.
    • 10  Vascotto F, Campagna M, Visintin M, Cattaneo A, Burrone OR: Effects of intrabodies specific for rotavirus NSP5 during the virus replicative cycle. J. Gen. Virol.85,3285–3290 (2004).
    • 11  Montero H, Arias CF, Lopez S: Rotavirus nonstructural protein NSP3 is not required for viral protein synthesis. J. Virol.80,9031–9038 (2006).▪▪ Evidence indicating the need of NSP3 for cellular mRNA and not for viral mRNA translation.
    • 12  Padilla-Noriega L, Paniagua O, GuzmaN, Leon S: Rotavirus protein NSP3 shuts off host cell protein synthesis. Virology298,1–7 (2002).
    • 13  Piron M, Vende P, Cohen J, Poncet D: Rotavirus RNA-binding protein NSP3 interacts with eIF4GI and evicts the poly(A) binding protein from eIF4F. EMBO J.17,5811–5821 (1998).
    • 14  Vende P, Piron M, Castagne N, Poncet D: Efficient translation of rotavirus mRNA requires simultaneous interaction of NSP3 with the eukaryotic translation initiation factor eIF4G and the mRNA 3´ end. J. Virol.74,7064–7071 (2000).
    • 15  Patton JT, Taraporewala Z, Chen D et al.: Effect of intragenic rearrangement and changes in the 3´ consensus sequence on NSP1 expression and rotavirus replication. J. Virol.75,2076–2086 (2001).
    • 16  Barro M, Patton JT: Rotavirus NSP1 inhibits expression of type I interferon by antagonizing the function of interferon regulatory factors IRF3, IRF5, and IRF7. J. Virol.81,4473–4481 (2007).
    • 17  Gonzalez SA, Mattion NM, Bellinzoni R, Burrone OR: Structure of rearranged genome segment 11 in two different rotavirus strains generated by a similar mechanism. J. Gen. Virol.70(Pt 6),1329–1336 (1989).
    • 18  Kojima K, Taniguchi K, Urasawa T, Urasawa S: Sequence analysis of normal and rearranged NSP5 genes from human rotavirus strains isolated in nature: implications for the occurrence of the rearrangement at the step of plus strand synthesis. Virology224,446–452 (1996).
    • 19  Gorziglia M, Nishikawa K, Fukuhara N: Evidence of duplication and deletion in super short segment 11 of rabbit rotavirus Alabama strain. Virology170,587–590 (1989).
    • 20  Lopez S, Arias CF: Early steps in rotavirus cell entry. Curr. Top. Microbiol. Immunol.309,39–66 (2006).
    • 21  Petrie BL, Greenberg HB, Graham DY, Estes MK: Ultrastructural localization of rotavirus antigens using colloidal gold. Virus Res.1,133–152 (1984).
    • 22  Altenburg BC, Graham DY, Estes MK: Ultrastructural study of rotavirus replication in cultured cells. J. Gen. Virol.46,75–85 (1980).
    • 23  Esparza J, Gorziglia M, Gil F, Romer H: Multiplication of human rotavirus in cultured cells: an electron microscopic study. J. Gen. Virol.47,461–472 (1980).
    • 24  Au KS, Chan WK, Burns JW, Estes MK: Receptor activity of rotavirus nonstructural glycoprotein NS28. J. Virol.63,4553–4562 (1989).
    • 25  Musalem C, Espejo RT: Release of progeny virus from cells infected with simian rotavirus SA11. J. Gen. Virol.66(Pt 12),2715–2724 (1985).
    • 26  Jourdan N, Maurice M, Delautier D et al.: Rotavirus is released from the apical surface of cultured human intestinal cells through nonconventional vesicular transport that bypasses the Golgi apparatus. J. Virol.71,8268–8278 (1997).
    • 27  Taraporewala Z, Chen D, Patton JT: Multimers formed by the rotavirus nonstructural protein NSP2 bind to RNA and have nucleoside triphosphatase activity. J. Virol.73,9934–9943 (1999).
    • 28  Schuck P, Taraporewala Z, McPhie P, Patton JT: Rotavirus nonstructural protein NSP2 self-assembles into octamers that undergo ligand-induced conformational changes. J. Biol. Chem.276,9679–9687 (2001).
    • 29  Kumar M, Jayaram H, Vasquez-Del Carpio R et al.: Crystallographic and biochemical analysis of rotavirus NSP2 with nucleotides reveals a nucleoside diphosphate kinase-like activity. J. Virol.81,12272–12284 (2007).
    • 30  Vasquez-Del Carpio R, Gonzalez-Nilo FD, Riadi G, Taraporewala ZF, Patton JT: Histidine triad-like motif of the rotavirus NSP2 octamer mediates both RTPase and NTPase activities. J. Mol. Biol.362,539–554 (2006).
    • 31  Vende P, Taraporewala ZF, Patton JT: RNA-binding activity of the rotavirus phosphoprotein NSP5 includes affinity for double-stranded RNA. J. Virol.76,5291–5299 (2002).
    • 32  Poncet D, Lindenbaum P, L’Haridon R, Cohen J: In vivo and in vitro phosphorylation of rotavirus NSP5 correlates with its localization in viroplasms. J. Virol.71,34–41 (1997).
    • 33  Torres-Vega MA, Gonzalez RA, Duarte M et al.: The C-terminal domain of rotavirus NSP5 is essential for its multimerization, hyperphosphorylation and interaction with NSP6. J. Gen. Virol.81,821–830 (2000).
    • 34  Afrikanova I, Miozzo MC, Giambiagi S, Burrone O: Phosphorylation generates different forms of rotavirus NSP5. J. Gen. Virol.77(Pt 9),2059–2065 (1996).
    • 35  Blackhall J, Fuentes A, Hansen K, Magnusson G: Serine protein kinase activity associated with rotavirus phosphoprotein NSP5. J. Virol.71,138–144 (1997).
    • 36  Welch SK, Crawford SE, Estes MK: Rotavirus SA11 genome segment 11 protein is a nonstructural phosphoprotein. J. Virol.63,3974–3982 (1989).
    • 37  Gonzalez SA, Burrone OR: Rotavirus NS26 is modified by addition of single O-linked residues of N-acetylglucosamine. Virology182,8–16 (1991).
    • 38  Fabbretti E, Afrikanova I, Vascotto F, Burrone OR: Two non-structural rotavirus proteins, NSP2 and NSP5, form viroplasm-like structures in vivo. J. Gen. Virol.80(Pt 2),333–339 (1999).▪▪ First demonstration of the ability of NSP2 and NSP5 to form viroplasm-like structures when co-expressed in uninfected cells.
    • 39  Gallegos CO, Patton JT: Characterization of rotavirus replication intermediates: a model for the assembly of single-shelled particles. Virology172,616–627 (1989).
    • 40  Patton JT, Jones MT, Kalbach AN, He YW, Xiaobo J: Rotavirus RNA polymerase requires the core shell protein to synthesize the double-stranded RNA genome. J. Virol.71,9618–9626 (1997).
    • 41  Tortorici MA, Broering TJ, Nibert ML, Patton JT: Template recognition and formation of initiation complexes by the replicase of a segmented double-stranded RNA virus. J. Biol. Chem.278,32673–32682 (2003).
    • 42  Patton JT, Vasquez-Del Carpio R, Tortorici MA, Taraporewala ZF: Coupling of rotavirus genome replication and capsid assembly. Adv. Virus Res.69,167–201 (2007).
    • 43  Taraporewala ZF, Schuck P, Ramig RF, Silvestri L, Patton JT: Analysis of a temperature-sensitive mutant rotavirus indicates that NSP2 octamers are the functional form of the protein. J. Virol.76,7082–7093 (2002).
    • 44  Jayaram H, Taraporewala Z, Patton JT, Prasad BV: Rotavirus protein involved in genome replication and packaging exhibits a HIT-like fold. Nature417,311–315 (2002).
    • 45  Carpio RV, Gonzalez-Nilo FD, Jayaram H et al.: Role of the histidine triad-like motif in nucleotide hydrolysis by the rotavirus RNA-packaging protein NSP2. J. Biol. Chem.279,10624–10633 (2004).
    • 46  Taraporewala ZF, Jiang X, Vasquez-Del Carpio R et al.: Structure-function analysis of rotavirus NSP2 octamer by using a novel complementation system. J. Virol.80,7984–7994 (2006).▪ First demonstration of the need of the NTPase activity of NSP2 for rotavirus genome synthesis, but not for viroplasm formation.
    • 47  Taraporewala ZF, Patton JT: Identification and characterization of the helix-destabilizing activity of rotavirus nonstructural protein NSP2. J. Virol.75,4519–4527 (2001).
    • 48  Afrikanova I, Fabbretti E, Miozzo MC, Burrone OR: Rotavirus NSP5 phosphorylation is up-regulated by interaction with NSP2. J. Gen. Virol.79(Pt 11),2679–2686 (1998).
    • 49  Eichwald C, Rodriguez JF, Burrone OR: Characterisation of rotavirus NSP2/NSP5 interaction and dynamics of viroplasms formation. J. Gen. Virol.85,625–634 (2004).
    • 50  Jiang X, Jayaram H, Kumar M et al.: Cryoelectron microscopy structures of rotavirus NSP2–NSP5 and NSP2–RNA complexes: implications for genome replication. J. Virol.80,10829–10835 (2006).
    • 51  Kattoura MD, Chen X, Patton JT: The rotavirus RNA-binding protein NS35 (NSP2) forms 10S multimers and interacts with the viral RNA polymerase. Virology202,803–813 (1994).
    • 52  Arnoldi F, Campagna M, Eichwald C, Desselberger U, Burrone OR: Interaction of rotavirus polymerase VP1 with nonstructural protein NSP5 is stronger than that with NSP2. J. Virol.81,2128–2137 (2007).▪ First description of the interaction of NSP5 with the rotavirus polymerase in the absence of other rotavirus proteins or viral RNA.
    • 53  Bar-Magen T, Spencer E, Patton JT: An ATPase activity associated with the rotavirus phosphoprotein NSP5. Virology369,389–399 (2007).
    • 54  Vende P, Tortorici MA, Taraporewala ZF, Patton JT: Rotavirus NSP2 interferes with the core lattice protein VP2 in initiation of minus- strand synthesis. Virology313,261–273 (2003).
    • 55  Berois M, Sapin C, Erk I, Poncet D, Cohen J: Rotavirus nonstructural protein NSP5 interacts with major core protein VP2. J. Virol.77,1757–1763 (2003).
    • 56  Mohan KV, Muller J, Atreya CD: The N- and C-terminal regions of rotavirus NSP5 are the critical determinants for the formation of viroplasm-like structures independent of NSP2. J. Virol.77,12184–12192 (2003).
    • 57  Sen A, Agresti D, Mackow ER: Hyperphosphorylation of the rotavirus NSP5 protein is independent of serine 67 or NSP2, and the intrinsic insolubility of NSP5 is regulated by cellular phosphatases. J. Virol.80,1807–1816 (2006).
    • 58  Campagna M, Burrone OR: Fusion of tags induces spurious phosphorylation of rotavirus NSP5. J. Virol.80,8283–8284; author reply 8284–8285 (2006).
    • 59  Eichwald C, Jacob G, Muszynski B, Allende JE, Burrone OR: Uncoupling substrate and activation functions of rotavirus NSP5: phosphorylation of Ser-67 by casein kinase 1 is essential for hyperphosphorylation. Proc. Natl Acad. Sci. USA101,16304–16309 (2004).
    • 60  Campagna M, Budini M, Arnoldi F et al.: Impaired hyperphosphorylation of rotavirus NSP5 in cells depleted of casein kinase 1α is associated with the formation of viroplasms with altered morphology and a moderate decrease in virus replication. J. Gen. Virol.88,2800–2810 (2007).
    • 61  Blackhall J, Munoz M, Fuentes A, Magnusson G: Analysis of rotavirus nonstructural protein NSP5 phosphorylation. J. Virol.72,6398–6405 (1998).
    • 62  Eichwald C, Vascotto F, Fabbretti E, Burrone OR: Rotavirus NSP5: mapping phosphorylation sites and kinase activation and viroplasm localization domains. J. Virol.76,3461–3470 (2002).
    • 63  Schmid S, Mayer D, Schneider U, Schwemmle M: Functional characterization of the major and minor phosphorylation sites of the P protein of Borna disease virus. J. Virol.81,5497–5507 (2007).
    • 64  Bergmann CC, Maass D, Poruchynsky MS, Atkinson PH, Bellamy AR: Topology of the non-structural rotavirus receptor glycoprotein NS28 in the rough endoplasmic reticulum. EMBO J.8,1695–1703 (1989).
    • 65  Both GW, Siegman LJ, Bellamy AR, Atkinson PH: Coding assignment and nucleotide sequence of simian rotavirus SA11 gene segment 10: location of glycosylation sites suggests that the signal peptide is not cleaved. J. Virol.48,335–339 (1983).
    • 66  Xu A, Bellamy AR, Taylor JA: Immobilization of the early secretory pathway by a virus glycoprotein that binds to microtubules. EMBO J.19,6465–6474 (2000).
    • 67  Berkova Z, Crawford SE, Trugnan G et al.: Rotavirus NSP4 induces a novel vesicular compartment regulated by calcium and associated with viroplasms. J. Virol.80,6061–6071 (2006).
    • 68  Mir KD, Parr RD, Schroeder F, Ball JM: Rotavirus NSP4 interacts with both the amino- and carboxyl-termini of caveolin-1. Virus Res.126,106–115 (2007).
    • 69  Parr RD, Storey SM, Mitchell DM et al.: The rotavirus enterotoxin NSP4 directly interacts with the caveolar structural protein caveolin-1. J. Virol.80,2842–2854 (2006).
    • 70  Storey SM, Gibbons TF, Williams CV et al.: Full-length, glycosylated NSP4 is localized to plasma membrane caveolae by a novel raft isolation technique. J. Virol.81,5472–5483 (2007).
    • 71  Sapin C, Colard O, Delmas O et al.: Rafts promote assembly and atypical targeting of a nonenveloped virus, rotavirus, in Caco-2 cells. J. Virol.76,4591–4602 (2002).
    • 72  Huang H, Schroeder F, Zeng C et al.: Membrane interactions of a novel viral enterotoxin: rotavirus nonstructural glycoprotein NSP4. Biochemistry40,4169–4180 (2001).
    • 73  Huang H, Schroeder F, Estes MK, McPherson T, Ball JM: Interaction(s) of rotavirus non-structural protein 4 (NSP4) C-terminal peptides with model membranes. Biochem. J.380,723–733 (2004).
    • 74  Meyer JC, Bergmann CC, Bellamy AR: Interaction of rotavirus cores with the nonstructural glycoprotein NS28. Virology171,98–107 (1989).
    • 75  Bugarcic A, Taylor JA: Rotavirus nonstructural glycoprotein NSP4 is secreted from the apical surfaces of polarized epithelial cells. J. Virol.80,12343–12349 (2006).
    • 76  Zhang M, Zeng CQ, Morris AP, Estes MK: A functional NSP4 enterotoxin peptide secreted from rotavirus-infected cells. J. Virol.74,11663–11670 (2000).
    • 77  Dong Y, Zeng CQ, Ball JM, Estes MK, Morris AP: The rotavirus enterotoxin NSP4 mobilizes intracellular calcium in human intestinal cells by stimulating phospholipase C-mediated inositol 1,4,5-trisphosphate production. Proc. Natl Acad. Sci. USA94,3960–3965 (1997).
    • 78  Ball JM, Tian P, Zeng CQ, Morris AP, Estes MK: Age-dependent diarrhea induced by a rotaviral nonstructural glycoprotein. Science272,101–104 (1996).
    • 79  Zambrano JL, Diaz Y, Pena F et al.: Silencing of rotavirus NSP4 or VP7 expression reduces alterations in Ca2+ homeostasis induced by infection of cultured cells. J. Virol.82,5815–5824 (2008).
    • 80  Sen A, Sen N, Mackow ER: The formation of viroplasm-like structures by the rotavirus NSP5 protein is calcium regulated and directed by a C-terminal helical domain. J. Virol.81,11758–11767 (2007).
    • 81  Cuadras MA, Bordier BB, Zambrano JL, Ludert JE, Greenberg HB: Dissecting rotavirus particle–raft interaction with small interfering RNAs: insights into rotavirus transit through the secretory pathway. J. Virol.80,3935–3946 (2006).
    • 82  Mattion NM, Cohen J, Aponte C, Estes MK: Characterization of an oligomerization domain and RNA-binding properties on rotavirus nonstructural protein NS34. Virology190,68–83 (1992).
    • 83  Piron M, Delaunay T, Grosclaude J, Poncet D: Identification of the RNA-binding, dimerization, and eIF4GI-binding domains of rotavirus nonstructural protein NSP3. J. Virol.73,5411–5421 (1999).
    • 84  Montero H, Rojas M, Arias CF, Lopez S: Rotavirus infection induces the phosphorylation of eIF2α but prevents the formation of stress granules. J. Virol.82,1496–1504 (2008).
    • 85  Harb M, Becker MM, Vitour D et al.: Nuclear localization of cytoplasmic poly(A)-binding protein (PABP-C1) upon rotavirus infection involves interaction of NSP3 with eIF4G and RoXaN. J. Virol.82(22),11283–11293 (2008).
    • 86  Poncet D, Laurent S, Cohen J: Four nucleotides are the minimal requirement for RNA recognition by rotavirus non-structural protein NSP3. EMBO J.13,4165–4173 (1994).
    • 87  Poncet D, Aponte C, Cohen J: Rotavirus protein NSP3 (NS34) is bound to the 3´ end consensus sequence of viral mRNAs in infected cells. J. Virol.67,3159–3165 (1993).
    • 88  Deo RC, Groft CM, Rajashankar KR, Burley SK: Recognition of the rotavirus mRNA 3´ consensus by an asymmetric NSP3 homodimer. Cell108,71–81 (2002).
    • 89  Komoto S, Sasaki J, Taniguchi K: Reverse genetics system for introduction of site-specific mutations into the double-stranded RNA genome of infectious rotavirus. Proc. Natl Acad. Sci. USA103,4646–4651 (2006).
    • 90  Kobayashi T, Antar AA, Boehme KW et al.: A plasmid-based reverse genetics system for animal double-stranded RNA viruses. Cell Host Microbe1,147–157 (2007).
    • 91  Boyce M, Roy P: Recovery of infectious bluetongue virus from RNA. J. Virol.81,2179–2186 (2007).
    • 92  Boyce M, Celma CC, Roy P: Development of reverse genetics systems for bluetongue virus: recovery of infectious virus from synthetic RNA transcripts. J. Virol.82,8339–8348 (2008).