Regulation of protein phosphatase activity by regucalcin localization in rat liver nuclei.
The regulatory role of regucalcin on protein phosphatase activity in isolated rat liver nuclei was investigated. Phosphatase activity toward phosphotyrosine and phosphoserine was significantly increased by the addition of CaCl(2) (10(-5) and 10(-4) M) in the enzyme reaction mixture. Trifluoperazine (25 and 50 microM), an antagonist of calmodulin, significantly inhibited protein phosphatase activity toward phosphoserine, while it had no effect on the enzyme activity toward phosphotysine and phosphothreonine. Cyclosporin A (10(-6)-10(-4) M), an inhibitor of Ca(2+)/calmodulin-dependent protein phosphatase activity toward phosphoserine, but not phosphotyrosine and phosphoserine. Thus, Ca(2+)/calmodulin-dependent phosphatases were present in liver nuclei. Regucalcin (0.25 and 0.5 microM) had an inhibitory effect on liver nuclear phosphatase activity toward phosphotyrosine, phosphoserine, and phosphothreonine. The presence of anti-regucalcin monoclonal antibody (25 and 50 ng/ml) in the enzyme reaction mixture caused a significant elevation of nuclear phosphatase activity toward three phosphoaminoacids. An analysis with sodium sulfate-polyacrylamide gel electrophoresis suggested a possibility of localization of regucalcin in liver nuclei. Moreover, regucalcin was determined in liver nuclei using enzyme-linked immunoadsorbent assay. The present study demonstrates that the endogenous regucalcin inhibits phosphatase activity in the liver nuclei.
Omura
M
M
Laboratory of Endocrinology and Molecular Metabolism, Graduate School of Nutritional Sciences, University of Shizuoka, Shizuoka 422-8526, Japan.
Yamaguchi
M
M
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000494:pharmacology; D002265:Carboxylic Ester Hydrolases; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology; D066298:In Vitro Techniques; D047908:Intracellular Signaling Peptides and Proteins; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D051379:Mice; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D015238:Sulfotransferases
D000911:Antibodies, Monoclonal; D002135:Calcium-Binding Proteins; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; C513324:Rgn protein, mouse; D015238:Sulfotransferases; C028152:alcohol sulfotransferase; D002265:Carboxylic Ester Hydrolases; C513325:Rgn protein, rat; D010749:Phosphoprotein Phosphatases