POTENCY OF WILD-TYPE AND TEMPERATURE-SENSITIVE VESICULAR STOMATITIS-VIRUS MATRIX PROTEIN IN THE INHIBITION OF HOST-DIRECTED GENE-EXPRESSION

The matrix (M) protein of vesicular stomatitis virus (VSV) functions i n virus assembly and also appears to be involved in the inhibition of host gene expression that is a characteristic cytopathic effect of VSV infection. Previous studies have shown that expression of M protein i nhibits host-directed transcription in the absence of other viral gene products and have suggested that only small amounts of M protein are required for the inhibition. In experiments described here, the potenc y of M protein in inhibition of host-directed gene expression was dete rmined by cotransfecting different amounts of in vitro-transcribed M p rotein mRNA together with a target gene encoding chloramphenicol acety l transferase (CAT) into BHK cells or PC12 cells that had been culture d in the presence or the absence of nerve growth factor. The results o f these experiments showed that the potency of M protein was similar i n the two cell types and was not affected by the extent of differentia tion of PC12 cells. Inhibition of CAT gene expression by M protein was also independent of the nature of the promoter activating sequences o f several different RNA polymerase Ii-dependent promoters. The amount of M protein needed to give 50% inhibition of CAT expression was estim ated to be 6700-11,000 copies per cell. Earlier data that temperature- sensitive (ts) M gene mutants of VSV inhibit host transcription had be en interpreted to indicate that M protein was not involved in the inhi bition. When the amount of M protein expressed was taken into account, ts M protein was as effective as wild-type M protein in the inhibitio n of host-directed transcription at the nonpermissive temperature Thus , inhibition of host transcription by ts M mutants of VSV is due to th e potent activity of M protein, which is evident even at the low level s produced at the nonpermissive temperature (C) 1996 Academic Press, I nc.