GA/GC-rich sequence confers Tat responsiveness to human neurotropic virus promoter, JCVL, in cells derived from central nervous system.
The human immunodeficiency virus type I (HIV-1) nuclear protein, Tat, is a potent transactivating factor that stimulates the rate of transcription of responsive promoters. Evidently, to exert its activity, Tat requires to be localized in close proximity of the transcription initiation site. Previous studies in our laboratory have demonstrated that Tat has the capacity to increase transcriptional activity of the late gene of a human neurotropic virus JC (JCV) in glial cells. In the present study, using deletion mutation analysis, we have identified a region upstream of the JCV late RNA start sites, termed upstream target (upTAR), that positively responds to Tat activation in glial cells. Using synthetic oligonucleotides spanning the upTAR sequence linked to a heterologous promoter, we have identified a GA/GC-rich region (GGAGGCGGAGGC) that confers TAT responsiveness preferentially on glial cell lines. Using gel mobility-shift and UV cross-linking assays, we have demonstrated that four major complexes (a-d) from glial and HeLa (non-glial) cells interact with the upTAR sequence. Whereas the molecular weights of the participant proteins in these complexes are similar in both glial and non-glial extracts, glial cells are enriched in proteins that form a major c complex. Interestingly, the participant proteins in complex c are developmentally regulated during brain development. The possible role of these proteins in increasing local concentrations of Tat in the vicinity of the JCV late RNA start sites is discussed.
Chowdhury
M
M
Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Kundu
M
M
Khalili
K
K
D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D001921:Brain / Q000382:microbiology*; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D015696:Gene Products, tat / Q000235:genetics*; D015497:HIV-1 / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D066298:In Vitro Techniques; D007577:JC Virus / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009457:Neuroglia / Q000382:microbiology*; D009687:Nuclear Proteins / Q000235:genetics; D011401:Promoter Regions, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D017384:Sequence Deletion; D015533:Transcriptional Activation; D054322:tat Gene Products, Human Immunodeficiency Virus
D004268:DNA-Binding Proteins; D015696:Gene Products, tat; D009687:Nuclear Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus