DETAILED ANALYSIS OF THE BASIC DOMAIN OF THE E2F1 TRANSCRIPTION FACTOR INDICATES THAT IT IS UNIQUE AMONG BHLH PROTEINS

The E2F1 transcription factor binds to sites within the promoters of a number of cell cycle regulated genes through a basic-helix-loop-helix motif (bHLH). It is shown here that the basic region of E2F1 is disti nct from that of all other bHLH proteins. The center of the basic regi on contains a helix breaking proline-glycine pair, (P122, G123), imply ing a turn within this region. This is in contrast to the known bHLH c ontaining proteins where the basic region is alpha-helical. Substituti on of P122 and G123 with alanines results in a significant reduction i n DNA binding levels, with the predicted formation of an alpha-helix. Also in contrast to other bHLH proteins, mutations generated in conser ved basic residues of E2F1 do not effect DNA binding. In addition, a s ingle leucine (191) between helix no. 2 and the leucine zipper is requ ired for DNA binding while the leucine zipper itself is not necessary. Finally, E2F1 interacts with ail of the G-residues in the sequence GG CGG-GAAA while the A-residues are not required for DNA binding. The un iqueness of the E2F1 DNA binding domain is likely to play a role in it s binding a DNA site that is distinct from that of all other bHLH prot eins (CACGTG).