IN-VITRO STIMULATION OF PROTEIN-KINASE-C BY MELATONIN

It has been shown that melatonin through binding to calmodulin acts bo th in vitro and in vivo as a potent calmodulin antagonist. It is known that calmodulin antagonists both bind to the hydrophobic domain of Ca 2+ activated calmodulin, and inhibit protein kinase C activity. In thi s work we explored the effects of melatonin on Ca2+ dependent protein kinase C activity in vitro using both a pure commercial rat brain prot ein kinase C, and a partially purified enzyme from MDCK and N1E-115 ce ll homogenates, The results showed that melatonin directly activated p rotein kinase C with a half stimulatory concentration of 1 nM. In addi tion the hormone augmented by 30% the phorbol ester stimulated protein kinase C activity and increased [H-3] PDBu binding to the kinase. In contrast, calmodulin antagonists (500 mu M) and protein kinase C inhib itors (100 mu M) abolished the enzyme activity. Melatonin analogs test ed were ineffective in increasing either protein kinase C activity or [H-3] PDBu binding. Moreover, the hormone stimulated protein kinase C autophosphorylation directly and in the presence of phorbol ester and phosphatidylserine. The results show that besides the melatonin bindin g to calmodulin, the hormone also interacts with protein kinase C only in the presence of Ca2+. They also suggest that the melatonin mechani sm of action may involve interactions with other intracellular hydroph obic and Ca2+ dependent proteins.