CAMP-DEPENDENT PROTEIN-KINASE IS NECESSARY FOR INCREASED NF-E2.DNA COMPLEX-FORMATION DURING ERYTHROLEUKEMIA CELL-DIFFERENTIATION

When murine erythroleukemia (MEL) cells are induced to differentiate by hexamethylene bisacetamide (HMBA), erythroid-specific genes are transcriptionally activated; however, transcriptional activation of these genes is severely impaired in cAMP dependent protein kinase (protein kinase A)-deficient MEL cells, The transcription factor NF-EB, composed of a 45-kDa (p45) and an 18-kDa (p18) subunit, is essential for enhancer activity of the globin locus control regions (LCRs), DNA binding of NF-E2 and alpha-globin LCR enhancer activity was significantly less in HMBA-treated protein kinase A-deficient cells compared to cells containing normal protein kinase A activity; DNA binding of several other transcription factors was the same in both cell types, In parental cells, HMBA treatment and/or prolonged activation of protein kinase A increased the amount of NF-E2 . DNA complexes without change in DNA binding affinity; the expression of p45 and p18 was the same under all conditions, p45 and p18 were phosphorylated by protein kinase A in vitro, but the phosphorylation did not affect NF-E2 . DNA complexes, suggesting that protein kinase A regulates NF-E2 . DNA complex formation indirectly, e,g, by altering expression of a regulatory factor(s), Thus, protein kinase A appears to be necessary for increased NF-E2 . DNA complex formation during differentiation of MEL cells and may influence erythroid-specific gene expression through this mechanism.