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Originally published In Press as doi:10.1074/jbc.M004562200 on September 27, 2000

J. Biol. Chem., Vol. 275, Issue 50, 39693-39701, December 15, 2000
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Transcriptional Activities of Reovirus RNA Polymerase in Recoated Cores
INITIATION AND ELONGATION ARE REGULATED BY SEPARATE MECHANISMS*

Diane L. FarsettaDagger §||, Kartik ChandranDagger §**, and Max L. NibertDagger §Dagger Dagger

From the Dagger  Department of Biochemistry, § Institute for Molecular Virology, and  Cell and Molecular Biology Program, University of Wisconsin-Madison, Madison, Wisconsin 53706

The particle-associated reovirus polymerase synthesizes mRNA within only certain viral particle types. Reovirus cores, subviral particles lacking outer capsid proteins µ1, sigma 3, and sigma 1, produce mRNA and abortive transcripts. Reovirus virions, which contain complete outer capsids, cannot produce mRNA and produce few abortive transcripts. Recoated cores are virion-like particles generated by the addition of recombinant outer capsid proteins to cores. We used recoated cores to analyze transcriptional regulation by reovirus outer capsid proteins. Partially recoated particles, containing less than virion amounts of µ1 and sigma 3, synthesized mRNA at levels inversely proportional to outer capsid protein levels. Fully recoated cores exhibited undetectable mRNA synthesis levels, as did virions. However, recoated cores produced high levels of abortive transcripts. Recoated core abortive transcripts remained particle-associated and appeared to inhibit further abortive transcript production. Proteolysis of recoated cores removing µ1 and sigma 3 released accumulated abortive transcripts and relieved inhibition of mRNA and abortive transcript synthesis. These results suggest transcriptional elongation, but not initiation, is blocked by virion-like amounts of µ1 and sigma 3. Particle-associated abortive transcripts may down-regulate transcriptional initiation. Minor outer capsid protein sigma 1 had no demonstrable effect on transcriptional activities. Transcriptional regulation may ensure progeny virions do not compete with transcribing particles for ribonucleoside triphosphates.

* This work was supported by National Institutes of Health Grant R29 AI39533 (to M. L. N.) and by a research grant from the Lucille P. Markey Charitable Trust to the Institute for Molecular Virology.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

This paper is dedicated to the memory of Janet L. Farsetta.

|| Also supported by National Institutes of Health Grant T32 GM08349 to the Biotechnology Training Program.

** Also supported by predoctoral fellowships from the Howard Hughes Medical Institute and from the Wisconsin Alumni Research Foundation. Current address: Dept. of Microbiology and Molecular Genetics, Harvard Medical School, Boston, MA 02115.

Dagger Dagger Received additional support as a Shaw Scientist from the Milwaukee Foundation and as a Vilas Associate from the University of Wisconsin. To whom correspondence should be addressed: Dept. of Microbiology and Molecular Genetics, Harvard Medical School, 200 Longwood Ave, Boston, MA 02115. Tel.: 617-432-4829; Fax: 617-738-7664; E-mail: mnibert@hms.harvard.edu.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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