Inducible and conditional inhibition of human immunodeficiency virus proviral expression by vesicular stomatitis virus matrix protein

Paik, Soon-Young; Banerjea, Akhil C.; Harmison, George G.; Chen, Changjie; Schubert, Manfred

doi:10.1128/jvi.69.6.3529-3537.1995

Cited by 33 publications

(12 citation statements)

References 42 publications

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“…However, the L clone used in these experiments has been shown to complement and rescue L gene temperature-sensitive mutants of VSV at the nonpermissive temperature (31). No M gene expression was detected in either BHK or COS cells by using immunoprecipitation methods, confirming similar observations reported by other laboratories (3,4,29).…”

Section: Ifn Induction In Virus-infected Cells Requires Synthesis Orsupporting

confidence: 89%

“…Lack of promoter specificity. The M protein has been shown to be a very powerful inhibitor of transcription of target genes controlled by various viral promoters (4,29). These studies demonstrated that expression of the CAT reporter gene from a cellular promoter was as susceptible to this inhibitor of transcription as expression from viral promoters.…”

Section: Discussionmentioning

confidence: 99%

“…The goal of this study was to determine whether the wt VSV M protein inhibits transcription from a cellular (IFN) promoter in addition to transcription from the SV40, CMV, and HIV-1 viral promoters studied previously (4,29). The general experimental strategy was to cotransfect cells with pTWU54, the IFN promoter-CAT reporter gene construct described in previous reports (6,38), and one of the five VSV genes attached to the SV40 late promoter.…”

Section: Inducibility Of the Ifn Promoter In Transiently Transfected mentioning

confidence: 99%

“…Northern blot and nuclear runoff transcription analyses demonstrated a reduction in the level of the vector-encoded reporter mRNAs and transcription of the target gene, respectively. Recently, Paik et al (29) demonstrated that VSV M protein can inhibit transcription from the chromosomally integrated human immunodeficiency virus type 1 (HIV-1) provirus, suggesting that the effect of M protein is not limited to transcription from plasmids. The data also supported the role of the M protein in shutoff of host cell transcription.…”

mentioning

confidence: 99%

“…Until now, the effects of M protein on expression of target genes from only a few viral promoters, including the SV40 early promoter, the cytomegalovirus (CMV) promoter, and the HIV-1 long terminal repeat, have been examined (4,29). In this study, we determined whether the M protein had the same effect on a cellular promoter, the human IFN-␤ promoter, as on the aforementioned viral promoters.…”

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confidence: 99%

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The vesicular stomatitis virus matrix protein inhibits transcription from the human beta interferon promoter

Ferran

Lucas‐Lenard

1997

J Virol

103

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In cells infected by wild-type (wt) vesicular stomatitis virus (VSV) Indiana, host transcription is severely inhibited. DNA cotransfection studies have implicated the VSV matrix (M) protein in this process (B. L. Black and D. S. Lyles, J. Virol. 66:4058-4064, 1992). The M protein inhibited transcription not only from viral promoters in plasmids but also from the chromosomally integrated human immunodeficiency virus type 1 (HIV-1) provirus promoter (S.-Y. Paik, A. C. Banerjea, G. G. Harmison, C.-J. Chen, and M. Schubert, J. Virol. 69:3529-3537, 1995). In this study, we investigated the effect of wt VSV M protein on expression of a reporter gene under control of a cellular promoter (beta-interferon [IFN-␤] promoter), using double transient transfections in BHK and COS-1 cells. The cellular IFN-␤ promoter was as susceptible to the inhibitory effect of the M protein as the viral promoters used previously. Viral proteins N, P, and G had no significant effect on reporter gene expression. The M protein gene from VSV mutant T1026R1, which is defective in host transcription inhibition, was cloned and sequenced, and its effect on reporter gene expression was tested. The mutant M protein had a methionine-to-arginine change at position 51 in the protein sequence and did not inhibit transcription from either the IFN-␤ promoter or viral promoters. This VSV mutant is a good inducer of IFN, as opposed to the wt virus, which suppresses IFN induction. These results show that the M protein inhibits transcription from cellular as well as viral promoters and that the M protein does not regulate the IFN promoter any differently from viral promoters. While the M protein may play a role in IFN gene regulation, other viral or cellular factors that provide specificity to the induction process must also be involved.Houston for quantitating the CAT assays, R. Schloemer for providing the pTWU54 plasmid, M. Schubert for providing the VSV expression vectors, J. Rose and E. Kretzschmar for the VSV polyclonal antibody, and S. Beausoleil and L. Poliquin for the sequence of the M gene from the HR strain of VSV (Indiana).

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Section: Ifn Induction In Virus-infected Cells Requires Synthesis Orsupporting

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Section: Inducibility Of the Ifn Promoter In Transiently Transfected mentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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The vesicular stomatitis virus matrix protein inhibits transcription from the human beta interferon promoter

Ferran

Lucas‐Lenard

1997

J Virol

103

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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

Lyles

McKenzie

1997

Virology

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In addition to its role in virus assembly, the matrix (M) protein of vesicular stomatitis virus (VSV) is involved in virus-induced cell rounding and inhibition of host-directed gene expression. Previous experiments 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 functional in virus assembly but defective in the inhibition of host-directed gene expression, while the MN1 deletion mutant, which lacks amino acids 4-21, inhibits host-directed gene expression but cannot function in virus assembly. Experiments presented here compared cell rounding induced by these two mutant M proteins to that of wt M protein. BHK cells were transfected with M protein mRNA transcribed in vitro, and the extent of cell rounding was evaluated at 24 hr posttransfection. The MN1 protein was nearly as effective as wt M protein in the induction of cell rounding, while tsO82 M protein expressed from transfected RNA was not able to induce cell rounding above that observed in negative controls without M protein, although it did cause BHK cells to have a less 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 activities 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 cell rounding was delayed relative to that following infection with wt VSV, suggesting that tsO82 M protein retains some cytopathic activity. The distribution of actin, vimentin, and tubulin in transfected cells was determined by fluorescence microscopy. In cells transfected with tsO82 M mRNA, these cytoskeletal elements were indistinguishable from those of negative control transfected cells. In cells rounded as a result of transfection with wt M or MN1 mRNA, actin-containing filaments were reorganized into a thick perinuclear ring but were not depolymerized. In contrast, tubulin and vimentin appeared to be diffusely distributed throughout the cytoplasm of rounded cells. These results support the idea that cell rounding induced by M protein results from the depolymerization of microtubules and/or intermediate filaments.

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Identification of a Consensus Mutation in M Protein of Vesicular Stomatitis Virus from Persistently Infected Cells That Affects Inhibition of Host-Directed Gene Expression

Ahmed

Lyles

1997

Virology

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In addition to its function in virus assembly, the viral matrix (M) protein of vesicular stomatitis virus (VSV) inhibits host-directed gene expression. The goal of this study was to determine whether sequence changes in M protein contribute to a reduced shut off of host gene expression in cells persistently infected with VSV. Viruses isolated from L cells persistently infected with VSV inhibited host RNA synthesis more slowly than wild-type (wt) VSV. M genes of the persistent viral population were cloned and sequenced. One mutation, an N to D change at position 163 of the protein sequence (N163D), was common to all the molecular clones. The N163D M protein was synthesized from transfected mRNA at a rate that was 30% of that of wt M protein, but was turned over at a rate that was similar to that of wt M protein. Transfection of mRNA encoding N163D M protein inhibited expression of a cotransfected target gene encoding chloramphenicol acetyl transferase (CAT), but the inhibition was 6 to 10 times less effective than transfection of equivalent amounts of wt M mRNA. This difference could not be accounted for by differences in translation of CAT mRNA. Thus, when the differences in M protein expression were taken into account, N163D M protein was 2 to 3 times less effective than wt M protein in the inhibition of host-directed gene expression, similar to the differences in host transcription observed in virus-infected cells. Point mutations in addition to the N163D mutation were found in about half of the M gene molecular clones. The M gene of an independently isolated molecular clone, N163D.2, contained two additional point mutations in its carboxy terminal region. N163D.2 M protein was highly defective in inhibition of host gene expression and was turned over more rapidly than wt M protein. These results support the idea that M gene mutations contribute to a reduced cytopathic effect in cells persistently infected with VSV.

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Inducible and conditional inhibition of human immunodeficiency virus proviral expression by vesicular stomatitis virus matrix protein

Cited by 33 publications

References 42 publications

The vesicular stomatitis virus matrix protein inhibits transcription from the human beta interferon promoter

The vesicular stomatitis virus matrix protein inhibits transcription from the human beta interferon promoter

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

Identification of a Consensus Mutation in M Protein of Vesicular Stomatitis Virus from Persistently Infected Cells That Affects Inhibition of Host-Directed Gene Expression

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