CREB (cAMP response element binding protein) and C/EBP alpha (CCAAT/enhancer binding protein) are required for the superstimulation of phosphoenolpyruvate carboxykinase gene transcription by adenoviral E1a and cAMP

In the present study, we observed superstimulated levels of cAMP-stimulated transcription from the phosphoenolpyruvate carboxykinase (PEPCK) gene prom oter in cells infected with wild-type adenovirus expressing 12 S and 13 S E 1a proteins, or in cells expressing 13 S E1a alone. cAMP-stimulated transcr iption was inhibited in cells expressing only 12 S E1a, but slightly elevat ed in cells expressing E1a proteins with mutations in conserved regions 1 o r 2, leading us to conclude that the superstimulation was mediated by conse rved region 3 of 13 S E1a. E1a failed to enhance cAMP-stimulated transcript ion from promoters containing mutations that abolish binding by cAMP respon se element binding protein (CREB) or CCAAT/enhancer binding proteins (C/EBP s). This result was supported by experiments in which expression of dominan t-negative CREB and/or C/EBP proteins repressed E1a- and cAMP-stimulated tr anscription from the PEPCK gene promoter. In reconstitution experiments usi ng a Gal4-responsive promoter, E1a enhanced cAMP-stimulated transcription w hen chimaeric Gal4-CREB and Gal4-C/EBP alpha were co-expressed. Phosphoryla tion of CREB on serine-133 was stimulated in cells treated with dibutyryl c AMP, whereas phosphorylation of C/EBP alpha was increased by E1a expression . Our data support a model in which cAMP agonists increase CREB activity an d stimulate PEPCK gene transcription, a process that is enhanced by E1a thr ough the phosphorylation of C/EBP alpha.