Transcription factors of the Sp1 family: Interaction with E2F and regulation of the murine thymidine kinase promoter

Promoters of growth and cell cycle regulated genes frequently carry binding sites for transcription factors of the E2F and Spl families. We have demonstrated recently that direct interaction between Spl and a sub-group of the E2F factors is essential for the regulation of certain promoters. We show here that the amino acids necessary for this interaction in both cases are located within the DNA binding domain. This is in line with the assumption, that the interaction between E2F and Sp-factors contributes to promoter-specificity. Cyclin A, which binds to E2F-1 in close vicinity to Spl does not interfere with this interaction. Moreover we have investigated the ability of other members of the Spl family to interact with E2F-1 and to regulate the activity of the E2F and Spl dependent murine thymidine kinase promoter. All few: factors of the Spl family are able to bind E2F-1 in co-immunoprecipitation and GST-pull down experiments. Mobility shift assays with oligonucleotides comprising the Spl, or both the Spl and the E2F binding site suggest that Spl and Sp3 supply most if not all activity binding to the GC-box of the thymidine kinase promoter in murine fibroblasts. Reporter gene assays in Drosophila melanogaster SL2 cells and murine fibroblast 3T6 cells demonstrate that the thymidine kinase promoter is activated strongly by Spl and Sp3, weakly by Sp4, and not at all by Sp2. Go-expression of E2F-1 results in synergistic activation in 3T6 but not in SL2 cells. (C) 1999 Academic Press.