MULTIPLE FUNCTIONAL DOMAINS OF AML1 - PU.1 AND C EBP-ALPHA SYNERGIZE WITH DIFFERENT REGIONS OF AML1/

Control elements of many genes are regulated by multiple activators wo rking in concert to confer the maximal level of expression, but the me chanism of such synergy is not completely understood. The promoter of the human macrophage colony-stimulating factor (M-CSF) receptor presen ts an excellent model with which we can study synergistic, tissue-spec ific activation for two reasons. First, myeloid-specific expression of the M-CSF receptor is regulated transcriptionally by three factors wh ich are crucial for normal hematopoiesis: PU.1, AML1, and C/EBP alpha. Second, these proteins interact in such a way as to demonstrate at le ast two examples of synergistic activation. We have shown that AML1 an d C/EBP alpha activate the M-CSF receptor promoter in a synergistic ma nner. As we report here, AML1 also synergizes, and interacts physicall y, with PU.1. Detailed analysis of the physical and functional interac tion of AML1 with PU.1 and C/EEP alpha has revealed that the proteins contact one another through their DNA-binding domains and that AML1 ex hibits cooperative DNA binding with C/EBP alpha but not with PU.1. Thi s difference in DNA-binding abilities may explain, in part, the differ ences observed in synergistic activation. Furthermore, the activation domains of all three factors are required for synergistic activation, and the region of AML1 required for synergy with PU.1 is distinct from that required for synergy with C/EBP alpha. These observations presen t the possibility that synergistic activation is mediated by secondary proteins contacted through the activation domains of AML1, C/EBP alph a, and PU.1.