Fig. 6. XCds1 is indirectly inactivated by cyclin B-Cdc2 kinase. (A) Extracts prepared from immature oocytes were immunoprecipitated with control rabbit IgG (open circles) or anti-XCds1 antibody (closed circles). The immunoprecipitates were incubated with starfish cyclin B-Cdc2 kinase. Phosphorylation was measured as a function of incubation time. (B) After 30 minutes incubation with cyclin B-Cdc2 kinase in A, phosphorylated proteins were resolved by SDS-PAGE and visualized by autoradiography (top). The arrow indicates the position corresponding to XCds1. Western blot analysis confirmed that equivalent amounts of XCds1 were recovered in the immunoprecipitates with anti-XCds1 antibody (bottom). The immunoprecipitates were subjected to XCds1 kinase assay, using GST-XCdc25C(254-316)-WT as a substrate (middle). Phosphorylation at Ser-287 on XCdc25C was visualized by western blot with anti-phospho human Cdc25C (Ser-216) antibody. (C) Immature oocytes were injected with 10 ng of mRNA for LacZ or Xenopus cyclin B1. Cyclin B1 mRNA-injected oocytes and progesterone-treated oocytes were collected when they underwent GVBD. XCds1 was immunoprecipitated with anti-XCds1 antibody and assayed for its kinase activity (top). Western blot analysis confirmed that equivalent amounts of XCds1 were recovered in the immunoprecipitates with anti-XCds1 antibody (middle). The same extracts were subjected to total histone H1 kinase assay (bottom).
