JVI Figure table search 04
Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Supplemental material
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Sosnovtsev, S. V.
Right arrow Articles by Green, K. Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Sosnovtsev, S. V.
Right arrow Articles by Green, K. Y.

 Previous Article  |  Next Article 

Journal of Virology, August 2006, p. 7816-7831, Vol. 80, No. 16
0022-538X/06/$08.00+0     doi:10.1128/JVI.00532-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Cleavage Map and Proteolytic Processing of the Murine Norovirus Nonstructural Polyprotein in Infected Cells{dagger}

Stanislav V. Sosnovtsev,1* Gaël Belliot,1,{ddagger} Kyeong-OK Chang,1,§ Victor G. Prikhodko,1 Larissa B. Thackray,2 Christiane E. Wobus,2 Stephanie M. Karst,2 Herbert W. Virgin,2 and Kim Y. Green1

Laboratory of Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland,1 Department of Pathology and Immunology, Washington University School of Medicine, St. Louis, Missouri2

Received 14 March 2006/ Accepted 23 May 2006

Murine norovirus (MNV) is presently the only member of the genus Norovirus in the Caliciviridae that can be propagated in cell culture. The goal of this study was to elucidate the proteolytic processing strategy of MNV during an authentic replication cycle in cells. A proteolytic cleavage map of the ORF1 polyprotein was generated, and the virus-encoded 3C-like (3CL) proteinase (Pro) mediated cleavage at five dipeptide cleavage sites, 341E/G342, Q705/N706, 870E/G871, 994E/A995, and 1177Q/G1178, that defined the borders of six proteins with the gene order p38.3 (Nterm)-p39.6 (NTPase)-p18.6-p14.3 (VPg)-p19.2 (Pro)-p57.5 (Pol). Bacterially expressed MNV 3CL Pro was sufficient to mediate trans cleavage of the ORF1 polyprotein containing the mutagenized Pro sequence into products identical to those observed during cotranslational processing of the authentic ORF1 polyprotein in vitro and to those observed in MNV-infected cells. Immunoprecipitation and Western blot analysis of proteins produced in virus-infected cells demonstrated efficient cleavage of the proteinase-polymerase precursor. Evidence for additional processing of the Nterm protein in MNV-infected cells by caspase 3 was obtained, and Nterm sequences 118DRPD121 and 128DAMD131 were mapped as caspase 3 cleavage sites by site-directed mutagenesis. The availability of the MNV nonstructural polyprotein cleavage map in concert with a permissive cell culture system should facilitate studies of norovirus replication.

* Corresponding author. Mailing address: 50 South Drive MSC8007, Building 50, Room 6316, Bethesda, MD 20892-8007. Phone: (301) 594-1666. Fax: (301) 480-5031. E-mail: ss216m{at}nih.gov.

{dagger} Supplemental material for this article may be found at http://jvi.asm.org/.

{ddagger} Present address: Laboratoire de Virologie, Centre National de Référence des Virus Entériques, UFR Médecine-CHU de Dijon, 7 boulevard Jeanne d'Arc, 21079 Dijon CEDEX, France.

§ Present address: Department of Diagnostic Medicine/Pathobiology, Kansas State University, Manhattan, KS 66506.

Journal of Virology, August 2006, p. 7816-7831, Vol. 80, No. 16
0022-538X/06/$08.00+0     doi:10.1128/JVI.00532-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:



Home Help [Feedback] [For Subscribers] [Archive] [Search] [Contents]
J. Bacteriol. Mol. Cell. Biol. Microbiol. Mol. Biol. Rev.
Clin. Vaccine Immunol. ALL ASM JOURNALS

Copyright © 2006 by the American Society for Microbiology. All rights reserved.