Cyclic AMP-mediated inhibition of transcription of the malic enzyme gene in chick embryo hepatocytes in culture - Characterization of a cis-acting element far upstream of the promoter

Glucagon, acting via cAMP, inhibits transcription of the malic enzyme gene in chick embryo hepatocytes. In transiently transfected hepatocytes, fragments from the 5'-flanking DNA of the malic enzyme gene confer cAMP responsiveness to linked reporter genes, The major inhibitory cAMP response element at -3180/-3114 base pairs (bp) is similar to the consensus binding site for AP1. DNA fragments from -3134/-3115, -1713/-944, and -413/-147 bp also contain inhibitory cAMP response elements. The negative action of cAMP is mimicked by overexpression of the catalytic subunit of protein kinase A, inhibited by overexpression of a specific inhibitor of protein kinase A, and inhibited by overexpression of the T3 receptor; these results indicate involvement of the classical eukaryotic pathway for cAMP action and suggest interaction between the T3 and cAMP pathways. Sequence-specific complexes form between nuclear proteins and a DNA fragment containing -3192/-3158 bp of 5'-flanking DNA. In nuclear extracts prepared from cells treated with chlorophenylthio-cyclic AMP and T3, the complexes have different masses than those formed with extracts from cells treated with T3 alone. Antibodies to c-Fos or ATF-2 inhibit formation of the complex formed by proteins from cells treated with chlorophenylthio-cyclic AMP and T3 but not by those from cells treated with T3 alone. These results suggest an important role for c-Fos and ATF-2 in glucagon-nediated inhibition of transcription of the malic enzyme gene.