Distinct cellular factors regulate the c-myb promoter through its E2F element

Most E2F-driven promoters are transiently activated around the G(1)/S trans ition. Although the promoter for the c-myb proto-oncogene harbors an E2F el ement, it is induced early in G(1) following entry into the cell cycle. Fur thermore, this promoter remains active throughout subsequent cell cycles. S ince E2F sites function as repressor elements during G(1) (due to the assoc iation of pRb with E2F factors), we investigated whether the E2F element in the c-myb promoter is regulated differently than E2F elements in promoters that are repressed during G(1). By gel shia analysis, the E2F element from the c-myb promoter was found to farm a unique complex, referred to as E2Fm yb-sp, which was not observed with E2F elements from several other promoter s. Antibodies to DP-1, E2F1 to -5, p107, or pRb failed to either supershift or block E2Fmyb-sp complex formation. Methylation interference experiments indicate that the DNA contact residues for the E2Pmyb-sp complex are disti nct from but overlapping with residues required for the binding of E2F prot eins. In addition to the identification of E2Fmyb-sp, we have found that SP -1 binds to the c-myb E2F element. Functional studies revealed that E2Fmyb- sp and/or SP-1 are required to achieve full activation of the c-myb promote r in different cell types and to maintain elevated expression of the c-myb promoter during G(1) in NIH 3T3 cells. These studies demonstrate that E2F e lements can be regulated differently through the binding of unique sets of proteins.