Cell Cycle-Regulated Association of E2F1 and Sp1 Is Related to Their Functional Interaction

Abstract

Because of the large number of growth-regulated genes containing binding sites for the transcription factors Sp1 and E2F and the reported ability of E2F to mediate cell cycle (growth) regulation, we studied interactions between E2F1 and Sp1. In transient transfection assays using Drosophila melanogaster SL2 cells, transfection with both Sp1 and E2F1 expression vectors resulted in greater than 85-fold activation of transcription 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 dihydrofolate reductase transcription. Transient transfection studies demonstrated that E2F1 could superactivate Sp1-dependent transcription in a promoter containing only Sp1 sites and that Sp1 could superactivate transcription of promoters through E2F sites, further demonstrating that these factors functionally interact with one another. Coimmunoprecipitation studies revealed that Sp1 and E2F1 are physically associated in Drosophila cells transfected with Sp1 and E2F1 expression vectors and in human cells, with maximal interaction detected in mid-to late G 1 . Additionally, E2F1 and Sp1 interact in vitro through specific domains of each protein, and the physical interaction and functional synergism appear to require the same regions. Taken together, these data demonstrate that E2F1 and Sp1 both functionally and physically interact; therefore, through this interaction, Sp1 and E2F1 may regulate transcription of genes containing binding sites for either or both factors.

Retinoblastoma protein associates with SP1 and activates the hamster dihydrofolate reductase promoter

“…Two reports describe a novel interaction between Sp1 and members of the E2F1 family, including the superactivation of Sp1-mediated transcription by E2F (Karlseder et al, 1996;Lin et al, 1996). Therefore, Sp1 is bound by both E2F and Rb proteins, each of which can increase Sp1-mediated transcription.…”

“…On the one hand, Sp1 interacts with other proteins such as p107 (Datta et al, 1995), p53 (Borellini and Glazer, 1993) and E2F (Karlseder et al, 1996;Lin et al, 1996); the interaction with p107 represses Sp1-dependent transcription whereas the interaction with E2F leads to synergistic activation of dhfr transcription. On the other hand, the tumor suppressor Rb is involved in the control of transcription of several genes.…”

Regulation of E2F-1 gene expression in avian cells

“…A similar observation has been made with the murine promoter (Hsiao et al, 1994). Azizkhan's laboratory has shown that E2F-1 can transactivate the hamster dihydrofolate reductase (DHFR) promoter via its SP-1 site, when E2F-1 can directly interact with SP-1 factor (Lin et al, 1996). As the quail E2F-1 promoter possesses SP-1 sites, some of which are occupied in vivo, one might argue that the E2F-1 transactivation on the E2F site deleted promoter is due to the E2F-1/SP-1 complex bound on SP-1 sites.…”

Transcriptional regulation of the hepatocyte growth factor (HGF) gene by the Sp family of transcription factors

“…However, as mentioned above, we can not exclude the possibility that Sp1 and Sp3 bound to the major Sp1 site cooperate with other -HSS 5 Figure 7 Identi®cation of DNaseI hypersensitive sites in the 5'-anking region of the mouse HGF promoter. Freshly isolated nuclei from NIH3T3 ®broblasts were treated with the indicated amounts of DNaseI, then their genomic DNA was extracted, transcription factors bound to other sites in the 5'-anking region of the mouse HGF promoter, since it is known that Sp1 cooperates with several transcription factors such as p53 (Gualberto and Baldwin, 1996), GATA-1 (Merika and Orkin, 1995), YY1 (Seto et al, 1993), E1a (Wagner and Green, 1994) and E2F (Lin et al, 1996). Control of HGF gene expression is complex.…”