Ubiquitin and the small ubiquitin-related modifier (SUMO) are small post-translational protein modifiers that alter the properties of their targets by affecting their activities, interactions, stabilities or intracellular localization (1
). Despite non-overlapping conjugation machineries, there is extensive cross-talk between the two modification systems (2
). A growing number of proteins are being identified as targets of both ubiquitin and SUMO. One example is IκBα, an inhibitor of the transcriptional activator NF-κB. IκBα is either ubiquitylated or sumoylated on the same lysine, but with opposite consequences for protein stability (3
). A rather different relationship applies to a class of ubiquitin ligases (E3s) that recognize sumoylated targets as substrates for ubiquitylation and have been implicated in various aspects of DNA repair and genome maintenance (4–9
). These SUMO-targeted ubiquitin ligases (STUbLs) are RING finger E3s harbouring conserved SUMO interaction motifs (SIMs), which consist of a hydrophobic core flanked by several acidic residues (10
). By interacting predominantly with poly-SUMO chains, they mediate ubiquitylation of the SUMO moieties themselves, as well as the proteins to which these are attached. Hence, sumoylation can serve as a signal for subsequent ubiquitylation, often followed by proteasome-mediated degradation. Despite a profound influence on the homeostasis of SUMO conjugates in the cell, few physiological STUbL substrates have been identified to date.
Post-translational modifications of the budding yeast sliding clamp protein, proliferating cell nuclear antigen (PCNA), present a unique example of how ubiquitin and SUMO cooperate in the context of DNA replication and repair (11
). In response to replication-stalling DNA damage, PCNA is monoubiquitylated at a highly conserved lysine, K164, by the E2–E3 complex Rad6–Rad18 (12
). This promotes the recruitment of a class of specialized polymerases capable of using damaged DNA as a template for translesion synthesis (13–15
). Extension to a polyubiquitin chain activates an error-free pathway of damage bypass that likely involves template switching (12
). In contrast to ubiquitylation of PCNA, which is common to all eukaryotes, modification by SUMO appears to be less prevalent. In Saccharomyces cerevisiae
, the SUMO E3 Siz1 promotes attachment of the SUMO homologue Smt3 mainly to K164 (12
). K127 is modified to a lesser degree in a Siz1-independent manner (16
). Sumoylation of budding yeast PCNA during S phase prevents unscheduled recombination events by enhancing the binding of an anti-recombinogenic helicase, Srs2 (17
). Hence, the modification enables ubiquitin-dependent damage bypass by blocking alternative processing pathways.
The cooperation between SUMO and ubiquitin in orchestrating lesion bypass raises the question of how the transition from the S phase-associated sumoylated form of PCNA to the damage-induced ubiquitylated form is accomplished. The ubiquitin E3 Rad18, which is rate limiting for both mono- and polyubiquitylation of PCNA (19
), is likely to play a critical role in this process. Intriguingly, a previous report suggested that Rad18 physically interacts with the SUMO E2 Ubc9 (12
). We have now identified a SIM in Rad18 that strongly stimulates its ubiquitin ligase activity towards the sumoylated form of PCNA. We propose that budding yeast Rad18 is adapted to act primarily on sumoylated PCNA and discuss the implications for the switch between the two modifications in response to DNA damage.