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Journal of Virology, April 2003, p. 4566-4576, Vol. 77, No. 8
0022-538X/03/$08.00+0 DOI: 10.1128/JVI.77.8.4566-4576.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
Reovirus
NS Protein Localizes to Inclusions through an Association Requiring the µNS Amino Terminus
Cathy L. Miller,1 Teresa J. Broering,1 John S. L. Parker,1 Michelle M. Arnold,1,2 and Max L. Nibert1,2*
Department of Microbiology and Molecular Genetics,1
Virology Training Program, Harvard Medical School, Boston, Massachusetts 021152
Received 11 October 2002/
Accepted 23 January 2003
Cells infected with mammalian reoviruses contain phase-dense inclusions, called viral factories, in which viral replication and assembly are thought to occur. The major reovirus nonstructural protein µNS forms morphologically similar phase-dense inclusions when expressed in the absence of other viral proteins, suggesting it is a primary determinant of factory formation. In this study we examined the localization of the other major reovirus nonstructural protein,
NS. Although
NS colocalized with µNS in viral factories during infection, it was distributed diffusely throughout the cell when expressed in the absence of µNS. When coexpressed with µNS,
NS was redistributed and colocalized with µNS inclusions, indicating that the two proteins associate in the absence of other viral proteins and suggesting that this association may mediate the localization of
NS to viral factories in infected cells. We have previously shown that µNS residues 1 to 40 or 41 are both necessary and sufficient for µNS association with the viral microtubule-associated protein µ2. In the present study we found that this same region of µNS is required for its association with
NS. We further dissected this region, identifying residues 1 to 13 of µNS as necessary for association with
NS, but not with µ2. Deletion of
NS residues 1 to 11, which we have previously shown to be required for RNA binding by that protein, resulted in diminished association of
NS with µNS. Furthermore, when treated with RNase, a large portion of
NS was released from µNS coimmunoprecipitates, suggesting that RNA contributes to their association. The results of this study provide further evidence that µNS plays a key role in forming the reovirus factories and recruiting other components to them.
* Corresponding author. Mailing address: Dept. of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Ave., Boston, MA 02115. Phone: (617) 645-3680. Fax: (617) 738-7664. E-mail:
mnibert{at}hms.harvard.edu.
Journal of Virology, April 2003, p. 4566-4576, Vol. 77, No. 8
0022-538X/03/$08.00+0 DOI: 10.1128/JVI.77.8.4566-4576.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.
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