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Journal of Virology, November 1998, p. 8597-8604, Vol. 72, No. 11
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Reovirus Growth in Cell Culture Does Not Require the Full Complement of Viral Proteins: Identification of a sigma 1s-Null Mutant

Steven E. Rodgers,1,2 Jodi L. Connolly,1,2 James D. Chappell,2,3 and Terence S. Dermody1,2,3,*

Departments of Microbiology and Immunology 1 and Pediatrics3 and Elizabeth B. Lamb Center for Pediatric Research,2 Vanderbilt University School of Medicine, Nashville, Tennessee 37232

Received 9 March 1998/Accepted 6 July 1998

The reovirus sigma 1s protein is a 14-kDa nonstructural protein encoded by the S1 gene segment. The S1 gene has been linked to many properties of reovirus, including virulence and induction of apoptosis. Although the function of sigma 1s is not known, the sigma 1s open reading frame is conserved in all S1 gene sequences determined to date. In this study, we identified and characterized a variant of type 3 reovirus, T3C84-MA, which does not express sigma 1s. To facilitate these experiments, we generated two monoclonal antibodies (MAbs) that bind different epitopes of the sigma 1s protein. Using these MAbs in immunoblot and immunofluorescence assays, we found that L929 (L) cells infected with T3C84-MA do not contain sigma 1s. To determine whether sigma 1s is required for reovirus infection of cultured cells, we compared the growth of T3C84-MA and its parental strain, T3C84, in L cells and Madin-Darby canine kidney (MDCK) cells. After 48 h of growth, yields of T3C84-MA were equivalent to yields of T3C84 in L cells and were fivefold lower than yields of T3C84 in MDCK cells. After 7 days of growth following adsorption at a low multiplicity of infection, yields of T3C84-MA and T3C84 did not differ significantly in either L cells or MDCK cells. To determine whether sigma 1s is required for apoptosis induced by reovirus infection, T3C84-MA and T3C84 were tested for their capacity to induce apoptosis, using an acridine orange staining assay. In these experiments, the percentages of apoptotic cells following infection with T3C84-MA and T3C84 were equivalent. These findings indicate that nonstructural protein sigma 1s is not required for reovirus growth in cell culture and does not influence the capacity of reovirus to induce apoptosis. Therefore, reovirus replication does not require the full complement of virally encoded proteins.

* Corresponding author. Mailing address: Lamb Center for Pediatric Research, D7235 MCN, Vanderbilt University School of Medicine, Nashville, TN 37232. Phone: (615) 343-9943. Fax: (615) 343-9723. E-mail: terry.dermody{at}mcmail.vanderbilt.edu.

Journal of Virology, November 1998, p. 8597-8604, Vol. 72, No. 11
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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