Abstract
The poliovirus-encoded, membrane-associated polypeptide 2C is believed to be required for initiation and elongation of RNA synthesis. We have expressed and purified recombinant, histidine-tagged 2C and examined its ability to bind to the first 100 nucleotides of the poliovirus 5' untranslated region of the positive strand and its complementary 3'-terminal negative-strand RNA sequences. Results presented here demonstrate that the 2C polypeptide specifically binds to the 3'-terminal sequences of poliovirus negative-strand RNA. Since this region is believed to form a stable cloverleaf structure, a number of mutations were constructed to examine which nucleotides and/or structures within the cloverleaf are essential for 2C binding. Binding of 2C to the 3'-terminal cloverleaf of the negative-strand RNA is greatly affected when the conserved sequence, UGUUUU, in stem a of the cloverleaf is altered. Mutational studies suggest that interaction of 2C with the 3'-terminal cloverleaf of negative-strand RNA is facilitated when the sequence UGUUUU is present in the context of a double-stranded structure. The implication of 2C binding to negative-strand RNA in viral replication is discussed.
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- Aldabe R., Carrasco L. Induction of membrane proliferation by poliovirus proteins 2C and 2BC. Biochem Biophys Res Commun. 1995 Jan 5;206(1):64–76. doi: 10.1006/bbrc.1995.1010. [DOI] [PubMed] [Google Scholar]
- Andino R., Rieckhof G. E., Achacoso P. L., Baltimore D. Poliovirus RNA synthesis utilizes an RNP complex formed around the 5'-end of viral RNA. EMBO J. 1993 Sep;12(9):3587–3598. doi: 10.1002/j.1460-2075.1993.tb06032.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Andino R., Rieckhof G. E., Baltimore D. A functional ribonucleoprotein complex forms around the 5' end of poliovirus RNA. Cell. 1990 Oct 19;63(2):369–380. doi: 10.1016/0092-8674(90)90170-j. [DOI] [PubMed] [Google Scholar]
- Argos P., Kamer G., Nicklin M. J., Wimmer E. Similarity in gene organization and homology between proteins of animal picornaviruses and a plant comovirus suggest common ancestry of these virus families. Nucleic Acids Res. 1984 Sep 25;12(18):7251–7267. doi: 10.1093/nar/12.18.7251. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bernstein H. D., Sarnow P., Baltimore D. Genetic complementation among poliovirus mutants derived from an infectious cDNA clone. J Virol. 1986 Dec;60(3):1040–1049. doi: 10.1128/jvi.60.3.1040-1049.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bienz K., Egger D., Pasamontes L. Association of polioviral proteins of the P2 genomic region with the viral replication complex and virus-induced membrane synthesis as visualized by electron microscopic immunocytochemistry and autoradiography. Virology. 1987 Sep;160(1):220–226. doi: 10.1016/0042-6822(87)90063-8. [DOI] [PubMed] [Google Scholar]
- Bienz K., Egger D., Pfister T., Troxler M. Structural and functional characterization of the poliovirus replication complex. J Virol. 1992 May;66(5):2740–2747. doi: 10.1128/jvi.66.5.2740-2747.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bienz K., Egger D., Troxler M., Pasamontes L. Structural organization of poliovirus RNA replication is mediated by viral proteins of the P2 genomic region. J Virol. 1990 Mar;64(3):1156–1163. doi: 10.1128/jvi.64.3.1156-1163.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cho M. W., Teterina N., Egger D., Bienz K., Ehrenfeld E. Membrane rearrangement and vesicle induction by recombinant poliovirus 2C and 2BC in human cells. Virology. 1994 Jul;202(1):129–145. doi: 10.1006/viro.1994.1329. [DOI] [PubMed] [Google Scholar]
- Echeverri A. C., Dasgupta A. Amino terminal regions of poliovirus 2C protein mediate membrane binding. Virology. 1995 Apr 20;208(2):540–553. doi: 10.1006/viro.1995.1185. [DOI] [PubMed] [Google Scholar]
- Harris K. S., Xiang W., Alexander L., Lane W. S., Paul A. V., Wimmer E. Interaction of poliovirus polypeptide 3CDpro with the 5' and 3' termini of the poliovirus genome. Identification of viral and cellular cofactors needed for efficient binding. J Biol Chem. 1994 Oct 28;269(43):27004–27014. [PubMed] [Google Scholar]
- Johnson K. L., Sarnow P. Three poliovirus 2B mutants exhibit noncomplementable defects in viral RNA amplification and display dosage-dependent dominance over wild-type poliovirus. J Virol. 1991 Aug;65(8):4341–4349. doi: 10.1128/jvi.65.8.4341-4349.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kitamura N., Semler B. L., Rothberg P. G., Larsen G. R., Adler C. J., Dorner A. J., Emini E. A., Hanecak R., Lee J. J., van der Werf S. Primary structure, gene organization and polypeptide expression of poliovirus RNA. Nature. 1981 Jun 18;291(5816):547–553. doi: 10.1038/291547a0. [DOI] [PubMed] [Google Scholar]
- Li J. P., Baltimore D. An intragenic revertant of a poliovirus 2C mutant has an uncoating defect. J Virol. 1990 Mar;64(3):1102–1107. doi: 10.1128/jvi.64.3.1102-1107.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Li J. P., Baltimore D. Isolation of poliovirus 2C mutants defective in viral RNA synthesis. J Virol. 1988 Nov;62(11):4016–4021. doi: 10.1128/jvi.62.11.4016-4021.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mirzayan C., Wimmer E. Biochemical studies on poliovirus polypeptide 2C: evidence for ATPase activity. Virology. 1994 Feb 15;199(1):176–187. doi: 10.1006/viro.1994.1110. [DOI] [PubMed] [Google Scholar]
- Mirzayan C., Wimmer E. Genetic analysis of an NTP-binding motif in poliovirus polypeptide 2C. Virology. 1992 Aug;189(2):547–555. doi: 10.1016/0042-6822(92)90578-d. [DOI] [PubMed] [Google Scholar]
- Pelletier J., Sonenberg N. Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature. 1988 Jul 28;334(6180):320–325. doi: 10.1038/334320a0. [DOI] [PubMed] [Google Scholar]
- Racaniello V. R., Baltimore D. Cloned poliovirus complementary DNA is infectious in mammalian cells. Science. 1981 Nov 20;214(4523):916–919. doi: 10.1126/science.6272391. [DOI] [PubMed] [Google Scholar]
- Rivera V. M., Welsh J. D., Maizel J. V., Jr Comparative sequence analysis of the 5' noncoding region of the enteroviruses and rhinoviruses. Virology. 1988 Jul;165(1):42–50. doi: 10.1016/0042-6822(88)90656-3. [DOI] [PubMed] [Google Scholar]
- Rodríguez P. L., Carrasco L. Poliovirus protein 2C contains two regions involved in RNA binding activity. J Biol Chem. 1995 Apr 28;270(17):10105–10112. doi: 10.1074/jbc.270.17.10105. [DOI] [PubMed] [Google Scholar]
- Rodríguez P. L., Carrasco L. Poliovirus protein 2C has ATPase and GTPase activities. J Biol Chem. 1993 Apr 15;268(11):8105–8110. [PubMed] [Google Scholar]
- Roehl H. H., Parsley T. B., Ho T. V., Semler B. L. Processing of a cellular polypeptide by 3CD proteinase is required for poliovirus ribonucleoprotein complex formation. J Virol. 1997 Jan;71(1):578–585. doi: 10.1128/jvi.71.1.578-585.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Roehl H. H., Semler B. L. Poliovirus infection enhances the formation of two ribonucleoprotein complexes at the 3' end of viral negative-strand RNA. J Virol. 1995 May;69(5):2954–2961. doi: 10.1128/jvi.69.5.2954-2961.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rohll J. B., Percy N., Ley R., Evans D. J., Almond J. W., Barclay W. S. The 5'-untranslated regions of picornavirus RNAs contain independent functional domains essential for RNA replication and translation. J Virol. 1994 Jul;68(7):4384–4391. doi: 10.1128/jvi.68.7.4384-4391.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Schlegel A., Giddings T. H., Jr, Ladinsky M. S., Kirkegaard K. Cellular origin and ultrastructure of membranes induced during poliovirus infection. J Virol. 1996 Oct;70(10):6576–6588. doi: 10.1128/jvi.70.10.6576-6588.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Shen Y., Igo M., Yalamanchili P., Berk A. J., Dasgupta A. DNA binding domain and subunit interactions of transcription factor IIIC revealed by dissection with poliovirus 3C protease. Mol Cell Biol. 1996 Aug;16(8):4163–4171. doi: 10.1128/mcb.16.8.4163. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Skinner M. A., Racaniello V. R., Dunn G., Cooper J., Minor P. D., Almond J. W. New model for the secondary structure of the 5' non-coding RNA of poliovirus is supported by biochemical and genetic data that also show that RNA secondary structure is important in neurovirulence. J Mol Biol. 1989 May 20;207(2):379–392. doi: 10.1016/0022-2836(89)90261-1. [DOI] [PubMed] [Google Scholar]
- Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
- Teterina N. L., Kean K. M., Gorbalenya A. E., Agol V. I., Girard M. Analysis of the functional significance of amino acid residues in the putative NTP-binding pattern of the poliovirus 2C protein. J Gen Virol. 1992 Aug;73(Pt 8):1977–1986. doi: 10.1099/0022-1317-73-8-1977. [DOI] [PubMed] [Google Scholar]
- Wimmer E., Hellen C. U., Cao X. Genetics of poliovirus. Annu Rev Genet. 1993;27:353–436. doi: 10.1146/annurev.ge.27.120193.002033. [DOI] [PubMed] [Google Scholar]
- Ypma-Wong M. F., Dewalt P. G., Johnson V. H., Lamb J. G., Semler B. L. Protein 3CD is the major poliovirus proteinase responsible for cleavage of the P1 capsid precursor. Virology. 1988 Sep;166(1):265–270. doi: 10.1016/0042-6822(88)90172-9. [DOI] [PubMed] [Google Scholar]