Functional interaction of transcriptional coactivator ASC-2 and C/EBPalpha in granulocyte differentiation of HL-60 promyelocytic cell.

HL-60 promyelocytic cells were treated with retinoic acid (RA) to stimulate granulocyte differentiation. CCAAT/enhancer binding protein alpha (C/EBPalpha) is known to be the molecular switch during early hematopoietic developmental events that direct cells to the granulocytic pathway. Here we show that the coactivator activating signal cointegrator-2 (ASC-2) plays an important role in differentiation of HL-60 cells into granulocytes by mediating C/EBPalpha-induced gene transcription. The differentiation inducer RA increased mRNA and protein expression of ASC-2. The protein-protein interaction of C/EBPalpha and ASC-2 was detected by coimmunoprecipitation during granulocyte differentiation. Subsequently, GST-pull-down assay revealed that the N-terminal transactivation domain of C/EBPalpha could interact with ASC-2. This functional interaction of ASC-2 with C/EBPalpha drove a synergistic enhancement of C/EBPalpha-dependent transactivation and overexpression of the N-terminal C/EBPalpha protein in HL-60 cells inhibited ASC-2 responsiveness for C/EBPalpha activity in granulocyte differentiation, indicating C/EBPalpha dependency of ASC-2 activity. Taken together, these results suggest that the differentiation-dependent expressed ASC-2 protein physically and functionally interacts with C/EBPalpha and increases its transactivation activity, regulating specific gene transcription for granulocyte differentiation. Copyright 2001 Academic Press.