Calcium-dependent development of secondary cytostatic factor (2 degrees CSF) from Xenopus laevis oocytes and zygotes

Fresh cytosols extracted from unfertilised eggs of Xenopus laevis contain a cytostatic factor (CSF) which arrests the cell cycle at metaphase when microinjected into cleaving blastomeres. This CSF is sensitive to Ca2+ and is designated primary CSF (1 degrees CSF). During storage of Ca2(+)-containing cytosols at 2 degrees C, a stable CSF activity appears which is designated secondary CSF (2 degrees CSF). In the present study, we report that 2 degrees CSF activity can be induced in cytosols extracted from stage VI oocytes, unfertilised eggs, electrically activated eggs or blastulae, in the presence of Ca2+. Both the intensity and the rate of 2 degrees CSF development are dependent on the concentration of Ca2+ ions added to the cytosol. At Ca2+ concentrations of 5-10 mM, 2 degrees CSF activity reaches a maximum in about 7 days. Secondary CSF is relatively resistant to heat but loses all activity after 5 min at 70 degrees C. When stored at-80 degrees C, 2 degrees CSF activity remains detectable for about 6 weeks. Cytological observations show that blastomeres arrested by microinjection of 2 degrees CSF developed in cytosols of unfertilised eggs, activated eggs or blastulae contain metaphase chromosomes embedded in a bipolar spindle that has no asters developed at its poles. In contrast, blastomeres arrested by 2 degrees CSF in cytosols of stage VI oocytes contain condensed chromosomes but no spindle is formed. The mechanisms of the development of 2 degrees CSF in Ca2(+)-containing cytosols and its mode of action are discussed.