Fig. 4. PLC-induced Ca²⁺ oscillations are regulated by nuclear import. MII-arrested mouse eggs were microinjected with Fura-dextran and PLC cRNA. Intracellular Ca²⁺ was monitored by changes in fluorescence excitation ratio. Time zero indicates approximately the point at which PLC was microinjected. The trace in a shows a series of Ca²⁺ oscillations in a control egg injected with PLC cRNA. In b, parallel treated eggs were also injected with 10 mg/ml (pipette concentration) WGA, which significantly (P<0.0001) prolonged the oscillations (control: mean 444.2±142.0 minutes, n=14; WGA: mean 826.5±104.3 minutes, n=13). MII eggs were also injected with Fura-dextran and Myc-tagged versions of either control PLC or PLC^K377E to monitor the nuclear localisation. Trace ci shows a series of Ca²⁺ oscillations in a control egg injected with Myc-PLC cRNA. In di, parallel eggs were injected with Myc-PLC^K377E. Oscillations were significantly (P<0.0001) prolonged with the NLS mutant (control: mean 335.9±93.2 minutes, n=20; K377E: mean 702.2±162.8 minutes, n=22). Embryos were fixed and stained (see Fig. 3). Confocal and brightfield images were simultaneously obtained (cii and dii) showing the absence of Myc-PLC^K377E in the nucleoplasm.