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Journal of Virology, June 2000, p. 5133-5141, Vol. 74, No. 11
Department of Pathology and Laboratory
Medicine, Research Institute, University of British Columbia,
Vancouver, British Columbia, Canada V5Z 4H4
Received 9 September 1999/Accepted 13 March 2000
Rubella virus nonstructural proteins, translated from input genomic
RNA as a p200 polyprotein and subsequently processed into p150 and p90
by an intrinsic papain-like thiol protease, are responsible for virus
replication. To examine the effect of p200 processing on virus
replication and to study the roles of nonstructural proteins in viral
RNA synthesis, we introduced into a rubella virus infectious cDNA clone
a panel of mutations that had variable defective effects on p200
processing. The virus yield and viral RNA synthesis of these mutants
were examined. Mutations that completely abolished (C1152S and G1301S)
or largely abolished (G1301A) cleavage of p200 resulted in
noninfectious virus. Mutations that partially impaired cleavage of p200
(R1299A and G1300A) decreased virus replication. An RNase protection
assay revealed that all of the mutants synthesized negative-strand RNA
as efficiently as the wild type does but produced lower levels of
positive-strand RNA. Our results demonstrated that processing of
rubella virus nonstructural protein is crucial for virus replication
and that uncleaved p200 could function in negative-strand RNA
synthesis, whereas the cleavage products p150 and p90 are required for
efficient positive-strand RNA synthesis.
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Mutational Analysis of the Rubella Virus Nonstructural
Polyprotein and Its Cleavage Products in Virus Replication and
RNA Synthesis
*
Corresponding author. Mailing address: Department of
Pathology and Laboratory Medicine, Research Institute, University of British Columbia, Vancouver, British Columbia, Canada V5Z 4H4. Phone:
(604) 875-2473. Fax: (604) 875-2496. E-mail:
sgillam{at}interchange.ubc.ca.
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