Selection of RNA replicons capable of persistent noncytopathic replication in mammalian cells.

Abstract:

:The natural life cycle of alphaviruses, a group of plus-strand RNA viruses, involves transmission to vertebrate hosts via mosquitoes. Chronic infections are established in mosquitoes (and usually in mosquito cell cultures), but infection of susceptible vertebrate cells typically results in rapid shutoff of host mRNA translation and cell death. Using engineered Sindbis virus RNA replicons expressing puromycin acetyltransferase as a dominant selectable marker, we identified mutations allowing persistent, noncytopathic replication in BHK-21 cells. Two of these adaptive mutations involved single-amino-acid substitutions in the C-terminal portion of nsP2, the viral helicase-protease. At one of these loci, nsP2 position 726, numerous substitution mutations were created and characterized in the context of RNA replicons and infectious virus. Our results suggest a direct correlation between the level of viral RNA replication and cytopathogenicity. This work also provides a series of alphavirus replicons for noncytopathic gene expression studies (E. V. Agapov, I. Frolov, B. D. Lindenbach, B. M. Prágai, S. Schlesinger, and C. M. Rice, Proc. Natl. Acad. Sci. USA 95:12989-12994, 1998) and a general strategy for selecting RNA viral mutants adapted to different cellular environments.

journal_name

J Virol

journal_title

Journal of virology

authors

Frolov I,Agapov E,Hoffman TA Jr,Prágai BM,Lippa M,Schlesinger S,Rice CM

doi

10.1128/JVI.73.5.3854-3865.1999

subject

Has Abstract

pub_date

1999-05-01 00:00:00

pages

3854-65

issue

5

eissn

0022-538X

issn

1098-5514

journal_volume

73

pub_type

杂志文章