CELL-CYCLE-REGULATED ASSOCIATION OF E2F1 AND SP1 IS RELATED TO THEIR FUNCTIONAL INTERACTION

Because of the large number of growth-regulated genes containing bindi ng sites for the transcription factors Sp1 and E2F and the reported ab ility of E2F to mediate cell cycle (growth) regulation, we studied int eractions between E2F1 and Sp1, In transient transfection assays using Drosophila melanogaster SL2 cells, transfection with both Sp1 and E2F 1 expression vectors resulted in greater than 85-fold activation of tr anscription from a hamster dihydrofolate reductase reporter construct, whereas cotransfection with either the Sp1 or E2F1 expression vector resulted in 30- or <2-fold activation, respectively, Therefore, these transcription factors act synergistically in activation of dihydrofola te reductase transcription, Transient transfection studies demonstrate d that E2F1 could superactivate Sp1-dependent transcription in a promo ter containing only Sp1 sites and that Sp1 could superactivate transcr iption of promoters through E2F sites, further demonstrating that thes e factors functionally interact with one another, Coimmunoprecipitatio n studies revealed that Sp1 and E2F1 are physically associated in Dros ophila cells transfected with Sp1 and E2F1 expression vectors and in h uman cells, with maximal interaction detected in mid- to late G(1). Ad ditionally, E2F1 and Sp1 interact in vitro through specific domains of each protein, and the physical interaction and functional synergism a ppear to require the same regions, Taken together, these data demonstr ate that E2F1 and Sp1 both functionally and physically interact; there fore, through this interaction, Sp1 and E2F1 may regulate transcriptio n of genes containing binding sites for either or both factors.