Cholecystokinin stimulates the down-regulation of CTP:phosphocholine cytidylyltransferase in pancreatic acinar cells.
Stimulation of rat pancreatic acinar cells with cholecystokinin (CCK) is known to result in a significant inhibition of CTP:phosphocholine cytidylyltransferase (CT), a rate-limiting enzyme in phosphatidylcholine biosynthesis. Immunoprecipitation of CT from 32P-labeled acinar cells revealed that CCK treatment also caused a marked reduction in CT phosphate levels. The effects of CCK were maximal over 60 min and dependent on concentration, exhibiting an EC50 of 800 pM. Other calcium mobilizing secretagogues such as carbamylcholine (100 microM) and bombesin (10 nM) also reduced CT phosphate levels to 20 and 39% of control, respectively. Treatment of cells with thapsigargin and/or 12-O-tetradecanoyl-phorbol-13-acetate established that a combination of increased intracellular Ca2+ and protein kinase C activation was necessary to decrease phosphorylated CT content. Conversely, secretin (10 nM) or 8-(4-chlorophenylthio)-cAMP (100 microM) added alone had no effects. Use of the compound JMV-180 indicated CCK was acting through the low affinity state of the CCKA receptor to reduce CT phosphate levels. Further, the decrease in phosphorylated CT caused by CCK was blocked by the phosphatase inhibitors okadaic acid (3 microM) and calyculin A (100 nM). Finally, immunoblotting from whole cell lysates revealed CT was partially degraded in response to CCK, providing a novel mechanism by which the inhibition of CT enzyme activity occurs in response to the hormone. Moreover, this degradation was also blocked by a phosphatase inhibitor. These data suggest that the dephosphorylation of either CT itself or some other regulatory molecule(s) which mediates the CCK-induced protease activation may play a central role in reducing CT enzyme levels in acinar cells.
Groblewski
G E
GE
Department of Physiology, University of Michigan, School of Medicine, Ann Arbor 48109.
Wang
Y
Y
Ernst
S A
SA
Kent
C
C
Williams
J A
JA
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000819:agonists / Q000378:metabolism; D002478:Cells, Cultured; D002766:Cholecystokinin / Q000494:pharmacology*; D019916:Choline-Phosphate Cytidylyltransferase; D015536:Down-Regulation / Q000187:drug effects; D005455:Fluorescent Antibody Technique; D008297:Male; D009713:Nucleotidyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010179:Pancreas / Q000187:drug effects* / Q000201:enzymology; D010710:Phosphates / Q000378:metabolism; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D017207:Rats, Sprague-Dawley
D010710:Phosphates; D002766:Cholecystokinin; D009713:Nucleotidyltransferases; D019916:Choline-Phosphate Cytidylyltransferase; D010749:Phosphoprotein Phosphatases; D002118:Calcium