Functional interaction of CCAAT/enhancer-binding-protein-α basic region mutants with E2F transcription factors and DNA

The transcription factor CCAAT/enhancer-binding protein {alpha} (C/EBP{alpha}) regulates cell cycle arrest and terminal differentiation of neutrophils and adipocytes. Mutations in the basic leucine zipper domain (bZip) of C/EBP{alpha} are associated with acute myeloid leukemia. A widely used murine transforming C/EBP{alpha} basic region mutant (BRM2) entails two bZip point mutations (I294A/R297A). BRM2 has been discordantly described as defective for DNA binding or defective for interaction with E2F. We have separated the two BRM2 mutations to shed light on the intertwined reciprocity between C/EBP{alpha}-E2F-DNA interactions. Both, C/EBP{alpha} I294A and R297A retain transactivation capacity and interaction with E2F-DP. The C/EBP{alpha} R297A mutation destabilized DNA binding, whereas the C/EBP{alpha} I294A mutation enhanced binding to DNA. The C/EBP{alpha} R297A mutant, like BRM2, displayed enhanced interaction with E2F-DP but failed to repress E2F-dependent transactivation although both mutants were readily suppressed by E2F1 for transcription through C/EBP cis-regulatory sites. In contrast, the DNA binding enhanced C/EBP{alpha} I294A mutant displayed increased repression of E2F-DP mediated transactivation and resisted E2F-DP mediated repression. Thus, the efficient repression of E2F dependent S-phase genes and the activation of differentiation genes reside in the balanced DNA binding capacity of C/EBP{alpha}.