REGULATION OF THE RAT PROOPIOMELANOCORTIN GENE-EXPRESSION IN ATT-20 CELLS .1. EFFECTS OF THE COMMON SECRETAGOGUES

Although the effects of the various secretagogues on corticotropin (AC TH) secretion have been well studied, their effects on the POMC gene e xpression have not been thoroughly characterized. In this study, we es tablished a new model system using the AtT20 mouse corticotroph tumor cell line transfected stably with a plasmid containing 0.7 kb of the r at POMC 5' promoter-luciferase fusion gene. The responsiveness to exog enous CRH improved markedly when the cells were cultured with low seru m medium (1% FBS) compared with serum rich medium (10%). Using this cu lture condition, we examined the effects of not only CRH but also othe r secretagogues such as catecholamines, vasopressin, and angiotensin I I, upon the transcriptional activity of the POMC gene. CRH stimulated POMC promoter activity (3.5-fold increase) as well as cAMP generation and ACTH secretion in a dose- and time-dependent manner, with the maxi mal effect being observed 3-5 h after the start of incubation. Catecho lamines, especially epinephrine (10 nM and above), also stimulated all parameters, although less potently than CRH, and the effect was mimic ked by the beta-, but not alpha-adrenergic, agonist, suggesting the in volvement of the beta-adrenergic receptor. The combined effects of epi nephrine and CRH were greater in all parameters than those of CRH alon e, and the effects of both hormones were completely blocked by H89, an inhibitor of protein kinase A. Vasopressin and angiotensin II showed minimal effects on POMC expression. Our results suggest that 1) catech olamines, as well as CRH, positively regulate the POMC gene at physiol ogical concentrations; 2) the cAMP-PKA. system is the common intracell ular signaling pathway for CRH and catecholamines; and 3) vasopressin and angiotensin II also have weak but significant stimulatory effects on POMC promoter activity.