Sp1-p53 heterocomplex mediates activation of HTLV-I long terminal repeat by 12-O-tetradecanoylphorbol-13-acetate that is antagonized by protein kinase C.
We have previously demonstrated that 12-O-tetradecanoylphorbol-13-acetate (TPA) activates human T-cell leukemia virus type-I long terminal repeat (LTR) in Jurkat cells by a protein kinase C (PKC)-independent mechanism involving a posttranslational activation of Sp1 binding to an Sp1 site located within the Ets responsive region-1 (ERR-1). By employing the PKC inhibitor, bisindolylmaleimide I and cotransfecting the reporter LTR construct with a vector expressing PKC-alpha, we demonstrated, in the present study, that this effect of TPA was not only independent of, but actually antagonized by, PKC. Electrophoretic mobility shift assays together with antibody-mediated supershift and immuno-coprecipitation analyses, revealed that the posttranslational activation of Sp1 was exerted by inducing the formation of Sp1-p53 heterocomplex capable of binding to the Sp1 site in ERR-1. Furthermore, we demonstrated that Jurkat cells contain both wild-type (w.t.) and mutant forms of p53 and we detected both of them in this complex at variable combinations; some molecules of the complex contained either the w.t. or the mutant p53 separately, whereas others contained the two of them together. Finally, we showed that the Sp1-p53 complexes could bind also to an Sp1 site present in the promoter of another gene such as the cyclin-dependent kinase inhibitor p21(WAF-1), but not to consensus recognition sequences of the w.t. p53. Therefore, we speculate that there might be several other PKC-independent biological effects of TPA which result from interaction of such Sp1-p53 complexes with Sp1 recognition sites residing in the promoters of a wide variety of cellular and viral genes.
Torgeman
A
A
Department of Microbiology and Immunology, Ben Gurion University of the Negev, Beer Sheva 84105, Israel.
Mor-Vaknin
N
N
Zelin
E
E
Ben-Aroya
Z
Z
Löchelt
M
M
Flügel
R M
RM
Aboud
M
M
D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D016384:Consensus Sequence / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics; D016572:Cyclosporine / Q000494:pharmacology; D004247:DNA / Q000235:genetics / Q000378:metabolism; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D015368:Human T-lymphotropic virus 1 / Q000235:genetics*; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D019169:Jurkat Cells; D046911:Macromolecular Substances; D008301:Maleimides / Q000494:pharmacology; D009154:Mutation / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D020218:Response Elements / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D020079:Terminal Repeat Sequences / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D015533:Transcriptional Activation / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism*
C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D007211:Indoles; D046911:Macromolecular Substances; D008301:Maleimides; D016329:Sp1 Transcription Factor; D016159:Tumor Suppressor Protein p53; D016572:Cyclosporine; D004247:DNA; D011493:Protein Kinase C; C088060:bisindolylmaleimide; D013755:Tetradecanoylphorbol Acetate