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)5–7. 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 essential for centrosome duplication, as an interactor and substrate of Cyclin F. Utilizing 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 via 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 is unable to 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.

Cyclin F suppresses B-Myb activity to promote cell cycle checkpoint control

“…Cyclin F does not bind to CDKs but has a cyclin box domain, which contains a hydrophobic patch motif 8,30,31 . In cyclin F, this hydrophobic patch with the amino-acid sequence MRYIL (MetArg-Tyr-Ile-Leu) associates with the Cy-motif (arginine-Xleucine) in its substrates 6,8 . To further investigate the interplay between cyclin F and B-Myb, we performed immunoprecipitation (IP) experiments to probe for an interaction between the two proteins, and we were able to co-immunoprecipitate cyclin F with Flag-tagged B-Myb (Fig.…”

“…Furthermore, cyclin F was described to regulate mitotic progression through control of centrosomal function by degrading CP110 (ref. 8). However, a role in checkpoint control has not yet been reported.…”

Fidgetin-like 1 is a ciliogenesis-inhibitory centrosome protein

“…Given that FIGL-1 has a ubiquitination site, 29 it is likely that FIGL-1 level is regulated via the ubiquitination-proteasome pathway, which regulates the levels of some other centrosome proteins, such as CP110. 30,31 We also noticed that the FIGL-1 level was lower in G2 phase than in other phases (Fig. 4E).…”

FBH1 protects melanocytes from transformation and is deregulated in melanomas

“…Extract preparation and immunoblotting have previously been described. 12 Whole-cell lysates were generated by lysing 1 volume of cells with three volumes of SDS lysis buffer [50 mM TRIS pH 6.8, 2% SDS, 10% glycerol, 1 mM Na 3 VO 4 , 1 mM NaF and protease inhibitor mix (Roche)]. For chromatin fractionation, cells were extracted with CSK buffer 13 for 5 min on ice and centrifuged for 3 min at 1,300 g. The insoluble pellets were digested with Turbo nuclease to generate the chromatin fraction.…”