Requirement of an E1A-sensitive coactivator for long-range transactivationby the beta-globin locus control region

Four erythroid-specific DNase I-hypersensitive sites at the 5'-end of the b eta-globin locus confer high-level transcription to the beta-globin genes, To identify coactivators that mediate long-range transactivation by this lo cus control region (LCR), we assessed the influence of E1A, an inhibitor of the CBP/p300 histone acetylase, on LCR function. E1A strongly inhibited tr ansactivation of A gamma- and beta-globin promoters by the HS2, HS2-HS3, an d HS1-HS4 subregions of the LCR in human K562 and mouse erythroleukemia cel ls. Short- and long-range transactivation mediated by the LCR were equally sensitive to E1A. The E1A sensitivity was apparent in transient and stable transfection assays, and E1A inhibited expression of the endogenous gamma-g lobin genes, Only sites for NF-E2 within HS2 were required for E1A sensitiv ity in K562 cells, and EIA abolished transactivation mediated by the activa tion domain of NF-E2. E1A mutants defective in CBP/p300 binding only weakly inhibited HS2-mediated transactivation, whereas a mutant defective in reti noblastoma protein binding strongly inhibited transactivation. Expression o f CBP/p300 potentiated HS2-mediated transactivation. Moreover, expression o f GAL4-CBP strongly increased transactivation of a reporter containing HS2 with a GAL4 site substituted for the NF-E2 sites. Thus, we propose that a C BP/p300-containing coactivator complex is the E1A-sensitive factor importan t for LCR function.