Mitotic apparatus and nucleoli compartmentalization of 50,000-dalton type II regulatory subunit of cAMP-dependent protein kinase in estrogen receptor negative MDA-MB-231 human breast cancer cells.
Affinity purified RI and RII antibodies of regulatory subunits (R) of type I (RI) and type II (RII) cAMP-dependent protein kinase were utilized to determine the immunological characterization and specific compartmentalization of R in estrogen receptor negative MDA-MB-231 human breast cancer cells. The 8-azido-(32P)-cAMP binding analysis of MDA-MB-231 cell extracts exhibited 47,000- and 50,000-dalton cAMP receptor proteins. RI and RII antibodies, by immunoprecipitation, detected the 47,000- and 50,000-dalton proteins, respectively. The 47,000-dalton protein was identified as RI as it showed a similar molecular weight as of bovine RI on SDS-polyacrylamide gel electrophoresis. Although 50,000-dalton protein did not co-migrate with bovine heart 54,000-dalton RII, it was identified as RII of MDA-MB-231 cells since it was specifically precipitated with RII antibody but not with RI antibody. An indirect immunofluorescence revealed that during different phases of growth of MDA-MB-231 cells, 50,000-dalton RII was specifically compartmentalized in the mitotic spindle and nucleoli of the cells whereas RI did not exhibit a specific compartmentalization in the cells, but was distributed throughout the cell components. These results suggest specific role(s) of 50,000-dalton RII at the nuclei of MDA-MB-231 cells.
D000818:Animals; D001943:Breast Neoplasms / Q000201:enzymology / Q000648:ultrastructure*; D002417:Cattle; D002451:Cell Compartmentation; D002466:Cell Nucleolus / Q000648:ultrastructure; D000242:Cyclic AMP / Q000378:metabolism*; D005260:Female; D006801:Humans; D046911:Macromolecular Substances; D008938:Mitosis; D008970:Molecular Weight; D011494:Protein Kinases / Q000378:metabolism*; D011960:Receptors, Estrogen / Q000032:analysis*
D046911:Macromolecular Substances; D011960:Receptors, Estrogen; D000242:Cyclic AMP; D011494:Protein Kinases