REGULATION OF RAT-LIVER MICROSOMAL CHOLESTEROL ESTER HYDROLASE BY REVERSIBLE PHOSPHORYLATION

The regulation of neutral cholesterol ester hydrolase activity by chan ges in its phosphorylation state was studied in rat liver microsomes. Treatment with cAMP-dependent protein kinase resulted in increased enz yme activity, which was further enhanced by the addition of cAMP and M gATP. Consistent activations were also achieved with MgCl2 and MgATP, the magnesium effect being abolished by ethylenediaminetetraacetic aci d and adenosine triphosphate. Cholesterol ester hydrolase was activate d twofold by free calcium and Ca2+/calmodulin; this latter effect was blocked by the chelator ethylene-glycol-bis(beta-aminoethyl ether)N,N, N',N'-tetraacetic acid and the calmodulin antagonist trifluoperazine. The phosphatase inhibitors pyrophosphate and glycerophosphate led to m arked and dose-dependent increases in esterase activity, whereas okada ic acid elicited no effect. Further more, pyrophosphate and okadaic ac id did not change the increases in enzyme activity promoted by Ca2+, C a2+/calmodulin, Mg2+ and MgATP. Cholesterol ester hydrolase was inacti vated in a concentration-dependent manner by nonspecific alkaline phos phatases. In cAMP-dependent protein kinase/cAMP- or Ca2+/calmodulin-ac tivated microsomes, a time-dependent loss of activation in cholesteryl oleate hydrolysis was caused by alkaline phosphatase. These findings suggest that microsomal cholesterol ester hydrolase is activated throu gh cAMP and Ca2+/calmodulin phosphorylation, whereas enzyme deactivati on is dependent on phosphatase action.