Fig. 5. Western blot analysis of the four cPKCs isoforms. (A) Cellular extracts of GV oocytes cultured for 30 or 90 minutes were subjected to biochemical fractionation (cytoplasmic, nucleoplasmic and nuclear envelope fractions) and analyzed by immunoblotting. For each cPKC isoform, western blots were probed with an antibody against the catalytic domain that reacted equally strongly with dephosphorylated and fully phosphorylated protein. The double asterisk indicates the 80-kDa mature PKC. The single asterisk denotes the presence of the 77-kDa form, which is phosphorylated at the two C-terminal sites. The 74-kDa form, indicated by a hyphen, has either no phosphate groups or a single phosphate on the carboxyl terminus. (B) Control blots demonstrating the purity of the subcellular fractions: ß-tubulin, used as a specific marker of the cytoplasm is absent from the nuclear fraction; lamin A/C, used as a specific marker of the nuclear envelope, is present in the nuclear envelope fraction and absent from the nucleoplasm.