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Fig. 2. PLC ${zeta}$ -induced Ca²⁺ oscillations at mitosis are controlled by nuclear membrane integrity. As in Fig. 1c, MII eggs were parthenogentically activated in the presence of cytochalsin D and cultured until interphase (G2). To monitor the exact timing of NEB and NER, embryos were injected (2-6 hours before NEB) with a fluorescent probe for nuclear membrane integrity (FITC-BSA-NLS) in addition to Fura-dextran and PLC ${zeta}$ cRNA. (a) Two pronuclei can be seen with FITC-BSA-NLS. At approximately 416 minutes (415.5) after injection of PLC ${zeta}$ cRNA the first pronucleus begins to break down. By 430 minutes both pronuclei have broken down and the FITC-BSA-NLS can no longer be detected. Measuring the decrease in nuclear FITC-BSA-NLS intensity of both pronuclei (red and blue lines) it was found that NEB of the first pronucleus occurred 6.3±4.1 minutes before the initial Ca ²⁺ transient (n=12) that was indicated by the Fura-dextran fluorescence ratio (black line). (b) NER was also monitored with FITC-BSA-NLS. Measuring the accumulation of FITC-BSA-NLS into the reforming nuclei showed that the first indication of NER occurred just prior to the last Ca²⁺ transient (14.3±4.3 minutes). As in a, the red and blue lines represent fluorescence from the FITC-BSA-NLS regions and the black line is the Fura-dextran fluorescence ratio.

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