FIGURE 4:

K71M Gwl protein induces oocyte maturation. (A) Oocyte morphology. Oocytes were treated with progesterone or injected with buffer or WT or K71M Gwl proteins purified from non-OA treated (interphase) Sf9 cells. After incubation overnight, GVBD was assessed by white-spot formation. An oocyte that did not undergo GVBD with K71M Gwl was designated “G2” (e.g., upper oocyte, right panel). (B) Analysis of K71M Gwl expressing oocytes. The oocytes in A were lysed and Western blotted for Gwl, cyclin B1, and pY15 Cdc2, as indicated. At this exposure level, the shifted form of endogenous Gwl in progesterone-treated (GVBD) oocytes is less apparent. (C) Maturation induced by K71M Gwl requires protein synthesis. Active Gwl was purified from OA-treated Sf9 cells as described previously and microinjected into oocytes, followed by incubation in the absence and presence of cycloheximide (CHX, 10 μg/ml). After 6 h, GVBD was assessed by white-spot formation.