JVI Accepts, published online ahead of print on 4 November 2009

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J. Virol. doi:10.1128/JVI.01571-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Localization of mammalian orthoreovirus proteins to cytoplasmic factory-like structures via nonoverlapping regions of μNS

Cathy L. Miller^*, Michelle M. Arnold, Teresa J. Broering, Craig E. Hastings, and Max L. Nibert^*

Department of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Avenue, and Program in Virology, Division of Medical Sciences, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115; and Department of Veterinary Microbiology and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa 50011

^* To whom correspondence should be addressed. Email: clm{at}iastate.edu. mnibert{at}hms.harvard.edu.

Virally induced structures called viral factories form throughout the cytoplasm of cells infected with mammalian orthoreoviruses (MRV). When expressed alone in cells, MRV nonstructural protein μNS forms factory-like structures very similar in appearance to viral factories, suggesting it is involved in forming the structural matrix of these structures. μNS also associates with MRV core particles; core proteins μ2, 1, 2, 3, and 2; and RNA-binding nonstructural protein NS. These multiple associations result in recruitment or retention of these viral proteins or particles at factory-like structures. In this study, we identified the regions of μNS necessary and sufficient for these associations and additionally examined the localization of viral RNA synthesis in infected cells. We found that short regions within the amino-terminal 220 residues of μNS are necessary for association with core particles and necessary and sufficient for association with proteins μ2, 1, 2, 2, and NS. We also found that only protein 3 associates with the carboxyl-terminal one-third of μNS and that viral RNA is synthesized within viral factories. These results suggest that μNS may act as a cytoplasmic scaffolding protein involved in localizing and coordinating viral replication or assembly intermediates for efficient production of progeny core particles during MRV infection.