A ROLE FOR A BENT DNA-STRUCTURE IN E2F-MEDIATED TRANSCRIPTION ACTIVATION

We examined the role of promoter architecture, as well as that of the DNA-bending capacity of the E2F transcription factor family, in the ac tivation of transcription, DNA phasing analysis revealed that a consen sus E2F site in the E2F1 promoter possesses an inherent bend with a ne t magnitude of 40 +/- 2 degrees and with an orientation toward the maj or groove relative to the center of the E2F site, The inherent DNA ben d is reversed upon binding of E2F, generating a net bend with a magnit ude of 25 +/- 3 degrees oriented toward the minor groove relative to t he center of the E2F site. We also found that three members of the E2F family, in conjunction with the DP1 protein, bend the DNA toward the minor groove, suggesting that DNA bending is a characteristic of the e ntire E2F family, The Rb-E2F complex, on the other hand, does not reve rse the intrinsic DNA bend, Analysis of a series of E2F1 deletion muta nts defined E2F1 sequences which are not required for DNA binding but are necessary for the DNA-bending capacity of E2F. An internal region of E2F1, previously termed the marked box, which is highly homologous among E2F family members, was particularly important in DNA bending, W e also found that a bent DNA structure can be a contributory component in the activation of the E2F1 promoter but is not critical in the rep ression of that promoter in quiescent cells, This finding: suggests th at E2F exhibits characteristics typical of modular transcription facto rs, with independent DNA-binding and transcriptional activation functi ons, but also has features of architectural factors that alter DNA str ucture.