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Wild-type (wt) vesicular stomatitis virus (VSV) strains stimulate plasmacytoid dendritic cells (pDC) through

Matrix (M) protein mutants of vesicular stomatitis virus (VSV), such as rM51R-M virus, are less virulent than wild-type (wt) VSV strains due to their inability to suppress innate immunity. Studies presented here show that when inoculated intranasally into mice, rM51R-M virus was cleared from nasal mucosa by day 2 postinfection and was attenuated for spread to the central nervous system, in contrast to wt VSV, thus accounting for its reduced virulence. However, it stimulated an antibody response similar to that in mice infected with the wt virus, indicating that it has the ability to induce adaptive immunity in vivo without causing disease. These results support the use of M protein mutants of VSV as vaccine vectors.Recombinant vesicular stomatitis virus (VSV)-based vectors expressing foreign proteins are currently being explored as vaccines due to their ability to induce strong immune responses and protect against challenges with a variety of pathogens, including human immunodeficiency virus and influenza virus (7,12,(19)(20)(21). Although laboratory-adapted strains of VSV are generally nonpathogenic in humans and nonhuman primates, the potential for VSV vectors to cause disease in humans has been addressed by the development of vectors that are attenuated for virus growth by deleting or mutating the viral glycoprotein (G protein) or by rearranging the natural gene order of VSV (4,5,8,18). In many cases, this has the undesired effect of reducing the level of antigen expression and reducing the subsequent immune response. An alternative strategy is to attenuate viral pathogenicity by reducing the ability of the virus to suppress host innate immune responses without compromising the yield of infectious progeny. This strategy has the advantages of high levels of antigen expression and also the enhancement of adaptive immune responses through the activation of innate immune responses.Wild-type (wt) strains of VSV effectively suppress the host innate immune response through the inhibition of host gene expression by the viral matrix (M) protein (2, 23). M protein is a multifunctional protein that is involved in the shutoff of host transcription, nuclear cytoplasmic transport, and translation during virus infection (13). Studies previously carried out in our laboratory demonstrate that a recombinant M protein mutant of VSV, rM51R-M virus, containing an arginine for methionine substitution at position 51 of the M protein sequence, is defective at inhibiting host gene expression (2). Therefore, in contrast to its isogenic recombinant wt counterpart (rwt virus), rM51R-M virus stimulates the expression of genes involved in host innate immune responses. Furthermore, rM51R-M virus does not cause disease in mice (1), nor do other strains of VSV containing the M51R M protein mutation (25). These results suggest that the suppression of host innate immune responses by M protein is a major determinant of virulence for VSV and that rM51R-M virus would be a safer and more effective vaccine vector than wt VSV strains. Although we an...

The matrix (M) protein of vesicular stomatitis virus (VSV) is a potent inhibitor in vivo of transcription by all three host RNA polymerases (RNAP). In the case of host RNA polymerase II (RNAPII), the inhibition is due to lack of activity of the TATA-binding protein (TBP), which is a subunit of the basal transcription factor TFIID. Despite the potency of M protein-induced inhibition in vivo, experiments presented here show that M protein cannot directly inactivate TFIID in vitro. Addition of M protein to nuclear extracts from uninfected cells did not inhibit transcription activity, indicating that the inhibition is indirect and is mediated through host factors. The host factors that are known to regulate TBP activity include phosphorylation by host kinases and association with different TBP-associated factor (TAF) subunits. However, TBP in VSV-infected cells was found to be assembled normally with its TAF subunits, as shown by ion exchange high-pressure liquid chromatography and sedimentation velocity analysis. A normal pattern of phosphorylation of TBP in VSVinfected cells was also observed by pH gradient gel electrophoresis. Collectively, these data indicate that M protein inactivates TBP activity in RNAPII-dependent transcription by a novel mechanism, since the known mechanisms for regulating TBP activity cannot account for the inhibition.