Proopiomelanocortin (POMC) is a critical precursor protein in the pituitary gland that regulates adrenal steroid hormone secretion by producing the adrenocorticotropic hormone (ACTH). Corticotropin-releasing hormone (CRH) modulates ACTH release via calcium influx through the voltage-operated Ca²⁺ channels and activation of Ca²⁺/calmodulin-dependent protein kinase II (CaMKII). In this study, we aimed to investigate the role of the calcium/calmodulin-dependent protein kinase kinase/ calcium/calmodulin-dependent protein kinase IV (CaMKK/CaMKIV) signaling cascade in CRH-induced POMC expression using ACTH-producing AtT20 cells, a cell line isolated from the pituitary gland of a mouse with tumor. Protein expression levels of CaMKK and CaMKIV were determined via western blotting. POMC transcription was analyzed via real-time polymerase chain reaction and reporter gene assays, and ACTH secretion was measured via enzyme-linked immunosorbent assay. In addition, effects of constitutively active CaMKK (CaMKK-CA) and CaMKIV (CaMKIV-CA) and their dominant-negative mutants on POMC promoter activity were assessed. CRH-induced CaMKIV phosphorylation was examined via western blotting. Both CaMKK and CaMKIV were expressed in the rat pituitary tissues; three random rats were used. Moreover, 10 nM CRH significantly increased POMC transcription and ACTH secretion in AtT20 cells. Inhibition of CaMKK and protein kinase A by STO-609 and H89, respectively, suppressed CRH-induced POMC transcription. Furthermore, CaMKK-CA and CaMKIV-CA independently activated the POMC promoter. CRH rapidly induced CaMKIV phosphorylation and nuclear localization, but these effects were blocked by STO-609. Overall, these findings suggest that the CaMKK/CaMKIV signaling pathway plays a crucial role in CRH-mediated POMC transcription and ACTH secretion in AtT20 cells.
Keywords: adrenocorticotropic hormone; att20; camkk/camkiv pathway; corticotropin-releasing hormone; proopiomelanocortin.
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