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SCFCyclin F controls centrosome homeostasis and mitotic fidelity through CP110 degradation

Abstract

Generally, F-box proteins are the substrate recognition subunits of SCF (Skp1–Cul1–F-box protein) ubiquitin ligase complexes, which mediate the timely proteolysis of important eukaryotic regulatory proteins1,2. Mammalian genomes encode roughly 70 F-box proteins, but only a handful have established functions3,4. The F-box protein family obtained its name from Cyclin F (also called Fbxo1), in which the F-box motif (the 40-amino-acid domain required for binding to Skp1) was first described5. Cyclin F, which is encoded by an essential gene, also contains a cyclin box domain, but in contrast to most cyclins, it does not bind or activate any cyclin-dependent kinases (CDKs)5,6,7. However, like other cyclins, Cyclin F oscillates during the cell cycle, with protein levels peaking in G2. Despite its essential nature and status as the founding member of the F-box protein family, Cyclin F remains an orphan protein, whose functions are unknown. Starting from an unbiased screen, we identified CP110, a protein that is essential for centrosome duplication, as an interactor and substrate of Cyclin F. Using a mode of substrate binding distinct from other F-box protein–substrate pairs, CP110 and Cyclin F physically associate on the centrioles during the G2 phase of the cell cycle, and CP110 is ubiquitylated by the SCFCyclin F ubiquitin ligase complex, leading to its degradation. siRNA-mediated depletion of Cyclin F in G2 induces centrosomal and mitotic abnormalities, such as multipolar spindles and asymmetric, bipolar spindles with lagging chromosomes. These phenotypes were reverted by co-silencing CP110 and were recapitulated by expressing a stable mutant of CP110 that cannot bind Cyclin F. Finally, expression of a stable CP110 mutant in cultured cells also promotes the formation of micronuclei, a hallmark of chromosome instability. We propose that SCFCyclin F-mediated degradation of CP110 is required for the fidelity of mitosis and genome integrity.

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Figure 1: Cyclin F and CP110 interact and colocalize to the centrosomes.
Figure 2: CP110 is targeted for ubiquitylation and degradation by SCF Cyclin F during the G2 phase of the cell cycle.
Figure 3: Cyclin F silencing induces centrosome and mitotic aberrations.
Figure 4: The failure to degrade CP110 causes centrosome and mitotic defects.

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Acknowledgements

We thank S. Elledge for Cyclin FFlox/– and Cyclin F−/− MEFs, J.R. Skaar for reading the manuscript and F.M. Forrester for technical help. M.P. is grateful to T.M. Thor for continuous support. This work was funded by fellowships from the American Italian Cancer Foundation to V.D’A. and V.D., a grant from the March of Dimes (1-FY08-372) to B.D. and grants from the National Institutes of Health (R01-GM057587, R37-CA076584 and R21-AG032560) to M.P. A.S., L.F. and M.P.W. are supported by the Stowers Institute for Medical Research. V.D’A. is a Leukemia & Lymphoma Society Fellow. M.P. is an Investigator with the Howard Hughes Medical Institute.

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V.D’A. and V.D. performed and planned all experiments and helped to write the manuscript. M.P. coordinated the study, oversaw the results, and wrote the manuscript. S.V. and B.D. provided reagents, advice and assistance with the analysis of γ-tubulin and Centrin 2 foci. A.S., L.F. and M.P.W. performed the mass spectrometry analysis of the Cyclin F complex purified by V.D’A. All authors discussed the results and commented on the manuscript.

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Correspondence to Michele Pagano.

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D’Angiolella, V., Donato, V., Vijayakumar, S. et al. SCFCyclin F controls centrosome homeostasis and mitotic fidelity through CP110 degradation. Nature 466, 138–142 (2010). https://doi.org/10.1038/nature09140

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