ACTIVITY OF VESICULAR STOMATITIS-VIRUS M-PROTEIN MUTANTS IN CELL ROUNDING IS CORRELATED WITH THE ABILITY TO INHIBIT HOST GENE-EXPRESSION AND IS NOT CORRELATED WITH VIRUS ASSEMBLY FUNCTION

In addition to its role in virus assembly, the matrix (M) protein of v esicular stomatitis virus (VSV) is involved in virus-induced cell roun ding and inhibition of host-directed gene expression. Previous experim ents have shown that two M protein mutants genetically dissociate the ability of M protein to inhibit host-directed gene expression from its function in virus assembly: M protein from tsO82 virus is fully funct ional in virus assembly but defective in the inhibition of host-direct ed gene expression, while the MN1 deletion mutant, which lacks amino a cids 4-21, inhibits host-directed gene expression but cannot function in virus assembly. Experiments presented here compared cell rounding i nduced by these two mutant M proteins to that of wt M protein. BHK cel ls were transfected with M protein mRNA transcribed in vitro, and the extent of cell rounding was evaluated at 24 hr posttransfection. The M N1 protein was nearly as effective as wt M protein in the induction of cell rounding, while tsO82 M protein expressed from transfected RNA w as not able to induce cell rounding above that observed in negative co ntrols without M protein, although it did cause BHK cells to have a le ss elongated shape. These results indicate that the ability of MN1 and tsO82 M proteins to induce cell rounding is not correlated with their virus assembly function. Instead the cell rounding activity of these mutants is correlated with their ability to inhibit host-directed gene expression. Previous data suggesting that these two cytopathic activi ties could be dissociated can be readily accounted for by quantitative differences in M protein expression required. Infection of either BHK cells or L cells with tsO82 virus induced cell rounding, although cel l rounding was delayed relative to that following infection with wt VS V, suggesting that tsO82 M protein retains some cytopathic activity. T he distribution of actin, vimentin, and tubulin in transfected cells w as determined by fluorescence microscopy. In cells transfected with ts O82 M mRNA, these cytoskeletal elements were indistinguishable from th ose of negative control transfected cells. In cells rounded as a resul t of transfection with wt M or MN1 mRNA, actin-containing filaments we re reorganized into a thick perinuclear ring but were not depolymerize d. In contrast, tubulin and vimentin appeared to be diffusely distribu ted throughout the cytoplasm of rounded cells. These results support t he idea that cell rounding induced by M protein results from the depol ymerization of microtubules and/or intermediate filaments. (C) 1997 Ac ademic Press.