Sp1-targeted inhibition of gene transcription by WP631 in transfected lymphocytes.
The binding of Sp1 transcription factor to DNA is considered a potential target for small ligands designed to interfere with gene transcription. We attempted to distinguish the direct inhibition of the Sp1-binding to DNA in vivo (cell culture) from more indirect effects due to the network of pathways that modulate cell cycle progression, which may decrease transcription without direct interference with Sp1-DNA interactions. We tested whether the Sp3 protein, whose putative binding sequence overlaps the Sp1 site, can inhibit Sp1-activated transcription and interfere with drug-DNA interactions. A well-characterized model system consisting of a wtGLUT1 (wild-type glucose transporter 1) gene promoter, or a mutated mut2GLUT1 promoter, linked to a CAT (chloramphenicol acetyltransferase) reporter gene, was used to analyze the effects of overexpressed Sp1 and Sp3 transcription factors in transiently transfected Jurkat T lymphocytes. Bisanthracycline WP631, a potent inhibitor of Sp1-activated transcription in vitro, was assayed for its ability to specifically inhibit transcription in transfected Jurkat T lymphocytes. The mut2GLUT1 promoter was used to further discriminate between the WP631 interference with Sp1-DNA complexes and Sp3-induced inhibition, since the Sp3-binding site is canceled in this promoter and replaced by a high-affinity binding site for WP631.
Mansilla
Sylvia
S
Instituto de Biología Molecular de Barcelona, CSIC, Parc Cientific de Barcelona, Josep Samitier, 1-5, E-08028 Barcelona, Spain.
Priebe
Waldemar
W
Portugal
José
J
D002455:Cell Division / Q000187:drug effects; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D003630:Daunorubicin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D019169:Jurkat Cells; D008214:Lymphocytes / Q000187:drug effects* / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection
D004268:DNA-Binding Proteins; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; C104426:WP 631; D051705:Sp3 Transcription Factor; D004247:DNA; D003630:Daunorubicin