Ca2+ regulates hormone secretion and proopiomelanocortin gene expression in melanotrope cells via the calmodulin and the protein kinase C pathways.
The mechanism by which Ca2+ regulates proopiomelanocortin (POMC)-derived peptide secretion and POMC mRNA levels was investigated in primary cultures of porcine intermediate lobe (IL) cells maintained in serum-free medium. POMC gene expression was evaluated by the dot blot hybridization assay with a 32P-labeled DNA probe complementary to the full-length sequence of porcine POMC mRNA. Treatment of IL cells for 24 h with the calmodulin (CAM) antagonists W7 and W13 reduced POMC mRNA levels by a maximum of 50% in a dose-dependent manner (ED50 approximately 10(-8) M). Accumulation of alpha-melanocyte-stimulating hormone (alpha-MSH) in the medium was also depressed by 50% after 8 h of treatment. The role of protein kinase C (PKC) was investigated by depleting the IL cell PKC content with phorbol ester treatment. Phorbol 12-myristate 13-acetate (PMA) at 5 X 10(-8) M induced a rapid translocation of cytoplasmic PKC activity toward the membrane. After 12 h of PMA treatment, PKC activity was undetectable in either the cytoplasmic or the particulate fractions. The same dose of PMA induced a time-dependent decrease in POMC mRNA levels (50% inhibition after 24 h). The same effect was seen with the phorbol ester phorbol 12,13-dibutyrate at 5 X 10(-8) M, whereas the inactive phorbol ester 4 alpha-phorbol at 5 X 10(-8) M was without effect after 24 h of treatment. PMA treatment had a biphasic effect on alpha-MSH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)
Loeffler
J P
JP
Institut de Physiologie Générale, Université Louis Pasteur, UA 309 du CNRS, Strasbourg, France.
Kley
N
N
Louis
J C
JC
Demeneix
B A
BA
D000818:Animals; D002118:Calcium / Q000502:physiology*; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism; D005786:Gene Expression Regulation; D006728:Hormones / Q000378:metabolism*; D008543:Melanins / Q000378:metabolism*; D010703:Phorbol Esters / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D011333:Pro-Opiomelanocortin / Q000235:genetics*; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology
D002147:Calmodulin; D006728:Hormones; D008543:Melanins; D010703:Phorbol Esters; D012333:RNA, Messenger; D013449:Sulfonamides; C017967:W 7; D011333:Pro-Opiomelanocortin; C033772:N-(4-aminobutyl)-5-chloro-2-naphthalenesulfonamide; D011493:Protein Kinase C; D002118:Calcium