Calmodulin and calmodulin-binding proteins in liver cell nuclei.
Three nuclear subfractions were prepared from isolated hepatocytes nuclei. The calmodulin content in whole nuclei was 79 ng/mg of protein. The soluble fraction obtained after digestion of the nuclei with DNase I and RNase A (S1 fraction) contained 252 ng of calmodulin/mg of protein. The pellet obtained after the digestion with nucleases was treated with 1.6 M NaCl, and the soluble fraction and the residual structures obtained after the treatment were called S2 fraction and nuclear matrix, respectively. The calmodulin contents of the S2 fraction and of the nuclear matrix were 68 and 190 ng/mg of protein, respectively. If nuclei were digested only with DNase I, the calmodulin content in the soluble fraction increased to 703 ng/mg of protein, indicating that part of the nuclear calmodulin is associated with active DNA. Five nuclear calmodulin-binding proteins were identified. Two, having apparent molecular masses of 240 and 150 kDa were only found in the nuclear matrix, whereas the other three, having molecular masses of 120, 65, and 40 kDa were found in different proportions in all nuclear subfractions. A calmodulin-dependent inhibition of protein phosphorylation in the S1 fraction was discovered. Purification attempts on the calmodulin-binding proteins of the S1 subfraction by calmodulin affinity chromatography yielded four major polypeptides with apparent molecular masses of about 41, 46, and 120 (two products) kDa. These polypeptides retained the ability to inhibit protein phosphorylation but not the sensitivity to calmodulin.
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002147:Calmodulin / Q000032:analysis* / Q000502:physiology; D002148:Calmodulin-Binding Proteins / Q000032:analysis* / Q000502:physiology; D002417:Cattle; D002458:Cell Fractionation; D002467:Cell Nucleus / Q000032:analysis* / Q000648:ultrastructure; D003850:Deoxyribonuclease I / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000648:ultrastructure*; D008297:Male; D008970:Molecular Weight; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D012259:Ribonuclease, Pancreatic / Q000378:metabolism
D002147:Calmodulin; D002148:Calmodulin-Binding Proteins; D010750:Phosphoproteins; D003850:Deoxyribonuclease I; D012259:Ribonuclease, Pancreatic; D002118:Calcium