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Four hours after infection of BHK cells by vesicular stomatitis virus (VSV), the rate of total protein synthesis was about 65% that of uninfected cells and synthesis of the 12 to 15 predominant cellular polypeptides was reduced to a level about 25% that of control cells. As determined by in vitro translation of isolated RNA and both one-and two-dimensional gel analyses of the products, all predominant cellular mRNA's remained intact and translatable after infection. The total amount of translatable mRNA per cell increased about threefold after infection; this additional mRNA directed synthesis of the five VSV structural proteins. To determine the subcellular localization of cellular and viral mRNA before and after infection, RNA from various sizes of polysomes and nonpolysomal ribonucleoproteins (RNPs) was isolated from infected and noninfected cells and translated in vitro. Over 80% of most predominant species of cellular mRNA was bound to polysomes in control cells, and over 60% was bound in infected cells. Only 2 of the 12 predominant species of translatable cellular mRNA's were localized to the RNP fraction, both in infected and in uninfected cells. The average size of polysomes translating individual cellular mRNA's was reduced about two-to threefold after infection. For example, in uninfected cells, actin (molecular weight 42,000) mRNA was found predominantly on polysomes with 12 ribosomes; after infection it was found on polysomes with five ribosomes, the same size of polysomes that were translating VSV N (molecular weight 52,000) and M (molecular weight 35,000) mRNA. We conclude that the inhibition of cellular protein synthesis after VSV infection is due, in large measure, to competition for ribosomes by a large excess of viral mRNA. The efficiency of initiation of translation on cellular and viral mRNA's is about the same in infected cells;

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Infection of animal cells by vesicular stomatitis virus (VSV) results in inhibition of translation of cellular mRNA. We showed previously that, in BHK cells infected by the Glasgow isolate of VSV Indiana, this is due to competition during the initiation step of protein synthesis of viral and cellular mRNA for a constant, limiting number of ribosomes. We show here that infection of the same cells with the San Juan isolate of VSV resulted in a more rapid shutoff of host protein synthesis and that this was paralleled by a more rapid accumulation of viral mRNA. Extending our conclusion that shutoff is due to mRNA competition, we show further that the average size of polysomes translating viral and cellular nmRNA was threefold smaller in cells infected by VSV San Juan than by VSV Glasgow, which, in turn, was about one-half that of uninfected cells. In all cases, cellular and viral mRNA's which encoded the same-sized polypeptides were found on the same-sized polysomes, a result indicating that the efficiency of translation of both types of mRNA's is about the same in the infected cell. Also, there was no preferential sequestration of viral or cellular mRNA's in ribonucleoprotein particles. Additional correlations between the levels of viral mRNA's and the inhibition of protein synthesis came from studies of three other wild-type VSV strains and also from studies with Vero and L cells. In particular, the rate of shutoff of Lcell protein synthesis after infection by any VSV isolate was slower than that in BHK cells, and this was correlated with a slower rate of accumulation of viral mRNA. VSV temperature-sensitive mutants which synthesized, at the nonpermissive temperature, no VSV mRNA failed to inhibit synthesis of cellular proteins. Stanners and co-workers (C. P. Stanners, A. M. Francoeur, and T. Lam, Cell 11:273-281, 1977) claimed that VSV mutant Rl inhibited synthesis of L cell protein synthesis less rapidly than did its parent wild-type strain HR. They concluded that this effect was due to a mutation in an unspecified VSV protein, "P." We found, in both L and BHK cells, that Rl infection resulted in a slightly slower inhibition of cellular mRNA translation than did HR infection and that this was correlated with a slightly reduced accumulation of VSV mRNA. The level of VSV mRNA, rather than any specific VSV protein, appeared to be the key factor in determining the rate of shutoff of host protein synthesis. Infection of mammalian cells by vesicular stomatitis virus (VSV) results in inhibition of synthesis of cellular protein and RNA and eventual cell death. Neither replication of the genomic RNA nor production of infectious virus is necessary for the inhibition of cellular protein synthesis; however, some transcription of the negative-stranded viral genome into mRNA is essential (3, 4, 6-8). Much, if not all, of the inhibition of cellular protein synthesis is manifested at the translational level: after infection cellular mRNA remains intact and fully functional in in vitro translation systems, yet is translated in th...

Vesicular stomatitis virus (VSV) particles formed at early times after infection contain only one-third the amount of viral glycoprotein (G protein), relative to the major internal structural proteins M and N, as is found in particles released later. These "early" particles also have a lower density in equilibrium sucrose gradients than do those formed later; however, the sedimentation velocity and specific infectivity of these two classes of particles are the same. VSV-infected cells also release virus-like particles which sediment considerably faster than authentic virions and contain a higher-than-normal proportion of the VSV G protein relative to internal VSV proteins. These particles have a reduced specific infectivity but a normal density in sucrose gradients. All classes of VSV virions contain a constant proportion of M and N polypeptides. The ratio of G protein to M or N protein, in contrast, can vary over a sixfold range; this implies that an interaction between a precise number of surface G proteins with either of the underlying M and N proteins is not a prerequisite for budding of infectious viral particles from the cell surface.A vesicular stomatitis virus (VSV) particle, like those of most lipid-containing animal viruses, is formed by budding from the plasma membrane of an infected cell (reviewed in references 3, 11, 12, 18, and 21). This complex process is not well understood, but must reflect the structural organization of the viral proteins in the infected cell and in the virion. The transmembrane (5,6,13,16,20) viral glycoprotein (G protein) is imbedded in the plasma membrane; it becomes, by far, the major protein exposed on the surfaces of infected cells and on the surface of the virion (2,3,7,21). The M protein is apparently localized in the inner surface of the virus membrane and may serve as a "bridge" between the C; protein and the viral nucleocapsid, which consists of one molecule of viral RNA and the other three virus-encoded proteins, N, NS, and L (11,12,17,21,22).The VSV budding process is not totally spe-