Fig. 6. Effects of recombinant Rpn10a and Rpn10e on cell-cycle progression in Xenopus cell-free extracts. (A) Recombinant Rpn10 proteins expressed in E.coli were added to CSF (cytostatic factor)-arrested extracts prepared from unfertilized Xenopus eggs. Cyclin B2 destruction was then measured as a function of time after triggering cell-cycle progression by the addition of 2 mM Ca²⁺. CSF extracts were periodically harvested and subjected to western blot analysis with an anti-cyclin B2 antibody. (B) Recombinant Rpn10 proteins (500 µg/ml) were added to the cell-cycle extracts prepared from activated Xenopus eggs. The extracts were periodically harvested after incubation at 23°C, and Cdc2 (histone H1) kinase activity was measured (lower panel). Chromosome condensation was monitored simultaneously by Hoechst staining (upper panel).