Fig. 5. Upregulation of Wee1 activity by tyrosine autophosphorylation is required for normal gastrulation. (A) Both cells of a two-cell embryo were injected with RNAs encoding KD-Wee1, YYY/FFF-Wee1 or WT-Wee1. Anti-Flag immune complexes isolated from lysates prepared at stages 5, 8, 9 and 10 were examined by immunoblot analysis using anti-phosphotyrosine and anti-FLAG antibodies. (B,C) Embryos injected with MO-Wee1 alone, MO-Wee1+RNA (WT), MO-Wee1+RNA (KD) or MO-Wee1+RNA (YYY/FFF) were scored for gastrulation defects at stage 11. Number of embryos examined: MO-Wee1 (n=17); MO-Wee1+WT Wee1 (n=34); MO-Wee1+KD-Wee1 (n=42); MO-Wee1+YYY/FFF-Wee1 (n=35). Rescue RNAs were injected at 2 ng/embryo. Lysates from the injected embryos were examined by immunoblot analysis using anti-Wee1 antibody.