Drosophila skpA, a component of SCF ubiquitin ligases, regulates centrosome duplication during the cell cycle. T.D. Murphy 1, C.S. Garafola 1, C.D. Kennedy 2, G.H. Karpen 2. 1) Department of Embryology, Carnegie Inst Washington, Baltimore, MD; 2) MBVL, Salk Institute for Biological Studies, La Jolla, CA.
Centrosome duplication must be precisely coupled to the DNA replication cycle to ensure that each cell has two and only two centrosomes at the onset of mitosis. Drosophila skpA, a putative component of SCF ubiquitin ligases, regulates the link between the centrosome and nuclear duplication cycles. Lethal skpA null mutants generated by imprecise excision of a P element exhibit dramatic centrosome overduplication in neuroblasts while maintaining a normal DNA content. Surprisingly, many mutant cells are able to organize bipolar spindles and execute a normal anaphase in the presence of extra functional centrosomes. skpA mutant cells also show increased localization of g-tubulin to centrosomes during interphase and an altered nuclear morphology. Studies in other systems have shown that cyclin E/cdk2 activity is necessary to initiate centrosome duplication and cyclin E is targeted for degradation by an SCF complex, suggesting that centrosome overduplication in skpA mutant cells may arise from aberrant accumulation of cyclin E. Consistent with this hypothesis, 35% of skpA mutant cells have high cyclin E levels, compared to 2% of cells in wildtype controls. However, overexpression of cyclin E with multiple heat shocks only partially phenocopies the skpA mutant phenotype and does not result in centrosome overduplication, suggesting that skpA mutant cells have a second defect. We propose that centrosome duplication is controlled by an unknown centrosome licensing factor (CLF) produced and loaded onto centrosomes early in the cell cycle when cyclin E/cdk2 activity is low, and that excess CLF is degraded by a SKPa-based SCF complex. skpA mutant cells would accumulate CLF allowing centrosome duplication to reinitiate in the same cell cycle, and this defect would be enhanced by elevated levels of cyclin E.