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This study identifies the degradation pathway for the F-box protein Dia2, which plays an important role in maintaining genomic integrity. The Hect domain E3 ligase Tom1 recognizes a stretch of positively charged residues in Dia2, leading to Dia2 degradation by the ubiquitin proteasome system. Failure to degrade Dia2 disrupts cell cycle dynamics.

Ubiquitin-mediated proteolysis plays a key role in many pathways inside the cell and is particularly important in regulating cell cycle transitions. SCF (Skp1/Cul1/F-box protein) complexes are modular ubiquitin ligases whose specificity is determined by a substrate-binding F-box protein. Dia2 is a Saccharomyces cerevisiaeF-box protein previously described to play a role in invasive growth and pheromone response pathways. We find that deletion of DIA2renders cells cold-sensitive and subject to defects in cell cycle progression, including premature S-phase entry. Consistent with a role in regulating DNA replication, the Dia2 protein binds replication origins. Furthermore, the dia2mutant accumulates DNA damage in both S and G2/M phases of the cell cycle. These defects are likely a result of the absence of SCFDia2activity, as a Dia2 ΔF-box mutant shows similar phenotypes. Interestingly, prolonging G1-phase in dia2cells prevents the accumulation of DNA damage in S-phase. We propose that Dia2 is an origin-binding protein that plays a role in regulating DNA replication.

Ubiquitin-mediated destruction of regulatory proteins is a frequent means of controlling progression through signaling pathways . F-box proteins are components of modular E3 ubiquitin protein ligases called SCFs, which function in phosphorylation-dependent ubiquitination (, reviewed in ). F-box proteins contain a carboxy-terminal domain that interacts with substrates and a 42–48 amino-acid F-box motif which binds to the protein Skp1 . Skp1 binding links the F-box protein with a core ubiquitin ligase composed of the proteins Cdc53/Cul1, Rbx1 (also called Hrt1 and Roc1) and the E2 ubiquitin-conjugating enzyme Cdc34 . The genomes of the budding yeast Saccharomyces cerevisiaeand the nematode worm Caenorhabditis eleganscontain, respectively, 16 and more than 60 F-box proteins ; in S. cerevisiae, the F-box proteins Cdc4, Grr1 and Met30 target cyclin-dependent kinase inhibitors, G1 cyclins and transcriptional regulators for ubiquitination (, reviewed in ). Only four mammalian F-box proteins (Cyclin F, Skp1, β-TRCP and NFB42) have been identified so far . Here, we report the identification of a family of 33 novel mammalian F-box proteins. The large number of these proteins in mammals suggests that the SCF system controls a correspondingly large number of regulatory pathways in vertebrates. Four of these proteins contain a novel conserved motif, the F-box-associated (FBA) domain, which may represent a new protein–protein interaction motif. The identification of these genes will help uncover pathways controlled by ubiquitin-mediated proteolysis in mammals.