CREB is one component of the binding complex of the Ces-2/E2A-HLF binding element and is an integral part of the interleukin-3 survival signal

The Ces-2/E2A-HLF binding element (CBE) is recognized by Caenorhabditis ele gans death specification gene product Ces-2 and human acute lymphocytic leu kemia oncoprotein E2A-HLF, In an attempt to identify a cellular CBE-binding protein(s) that may be involved in apoptosis regulation in mammals, multip le nuclear binding complexes of CBE were identified in various mammalian ce ll lines and tissues by electrophoretic mobility shift assay. Cyclic AMP (c AMP)-responsive element (CRE)-binding protein (CREB) was present in one maj or CBE complex of Ba/F3 and TF-1 cells, and both in vitro-translated and Es cherichia coli-synthesized CREB bound to CBE, Activation of CREB by cAMP-el evating chemicals or the catalytic subunit of protein kinase A (PKAc) resul ted in induction of the CBE-driven reporter gene. Stimulation of Ba/F3 cell s with interleukin-3 (IL-3) promptly induced phosphorylation of CREB at ser ine(133) partially via a PKA-dependent pathway. Consistently, Ba/F3 cell su rvival in the absence of IL-3 was prolonged by activation of PKA. Conversel y, treatment of cells with a PKA inhibitor or expression of the dominant ne gative forms of the regulatory subunit type I of PKA and CREB overrode the survival activity of IL-3, Last, the bcl-2 gene was demonstrated to be one candidate cellular target of the CREB-containing CBE complex, as mutations in the CRE and CBE sites significantly reduced the IL-3 inducibility of the bcl-2 promoter. Together, our results suggest that CREB is one cellular co unterpart of Ces-2/E2A-HLF and is part of IL-3 dependent apoptosis regulati on in hematopoietic cells.