Molecular mechanisms of visna virus Tat: identification of the targets for transcriptional activation and evidence for a post-transcriptional effect.
Visna virus is a pathogenic lentivirus of sheep that is distantly related to the primate lentiviruses, including the human immunodeficiency virus type 1 (HIV-1). Replication of HIV-1 in cell culture requires the expression of a virus-encoded protein, Tat, which is a potent trans-activator of viral gene expression. Visna virus encodes an analogous Tat protein that greatly increases gene expression directed by the visna viral LTR. This report uses a stable vero cell line that constitutively expresses visna virus Tat to investigate the molecular mechanism of action of Tat on viral gene expression. Transient expression assays, using the visna virus LTR to drive transcription of the bacterial gene for chloramphenicol acetyltransferase (CAT), demonstrate that Tat trans-activates gene expression by increasing steady-state mRNA levels. The increase in steady-state mRNA levels is sufficient to account for the increase in protein observed and is due, in part, to an increase in the rate of transcription initiation. Tat mediates the accumulation of mRNA through AP-4 and AP-1 binding sites located in the U3 region of the LTR. Deletion of the upstream AP-1 and AP-4 binding sites results in a residual low level of trans-activation by Tat. Further experiments, using LTRs with R-U5 sequences deleted to +10, demonstrate AP-1 and AP-4 mediated responses to TAT at the RNA level, but no increase was observed in CAT protein.
Gdovin
S L
SL
Department of Immunology and Infectious Diseases, Johns Hopkins University School of Hygiene and Public Health, Baltimore, Maryland 21205.
Clements
J E
JE
D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000502:physiology*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D012091:Repetitive Sequences, Nucleic Acid; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014790:Visna-maedi virus / Q000235:genetics*
D004268:DNA-Binding Proteins; D015696:Gene Products, tat; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D014157:Transcription Factors; C055473:enhancer-binding protein AP-4