Esc2 orchestrates substrate-specific sumoylation by acting as a SUMO E2 cofactor in genome maintenance

Abstract

SUMO modification regulates diverse cellular processes by targeting hundreds of proteins. However, the limited number of sumoylation enzymes raises the question of how such a large number of substrates are efficiently modified. Specifically, how genome maintenance factors are dynamically sumoylated at DNA replication and repair sites to modulate their functions is poorly understood. Here, we demonstrate a role for the conserved yeast Esc2 protein in this process by acting as a SUMO E2 cofactor. Esc2 is required for genome stability and binds to Holliday junctions and replication fork structures. Our targeted screen found that Esc2 promotes the sumoylation of a Holliday junction dissolution complex and specific replisome proteins. Esc2 does not elicit these effects via stable interactions with substrates or their common SUMO E3. Rather, we show that a SUMO-like domain of Esc2 stimulates sumoylation by exploiting a noncovalent SUMO binding site on the E2 enzyme. This role of Esc2 in sumoylation is required for Holliday junction clearance and genome stability. Our findings thus suggest that Esc2 acts as a SUMO E2 cofactor at distinct DNA structures to promote the sumoylation of specific substrates and genome maintenance.

Integrative analysis reveals unique structural and functional features of the Smc5/6 complex

“…Given that other SUMO ligases bind to SUMO to support SUMO transfer to substrates, the apparent lack of SUMO binding by Nse2 suggests that Nse5 may substitute for it during specific sumoylation events. Interestingly, we recently found that the Esc2 protein acts as another cofactor that helps Nse2-mediated sumoylation of the STR complex and Pol2 (48). It is thus possible that Nse2 may collaborate with different cofactors to efficiently sumoylate distinct substrates, and that the Smc5/6 complex may act as a "composite SUMO ligase."…”

Multifaceted regulation of the sumoylation of the Sgs1 DNA helicase

“…We have previously established an in vitro sumoylation system that can robustly sumoylate Sgs1 when the purified STR complex is incubated with purified sumoylation enzymes (Fig. S1A) (18). As seen for other sumoylation reactions, the addition of SUMO E1, SUMO E2, and SUMO in the presence of ATP are sufficient to support basal level of Sgs1 sumoylation, since the SUMO E2 can transfer SUMO to substrates upon its activation by the SUMO E1 at the consumption of ATP (14,18,22).…”

“…S1A) (18). As seen for other sumoylation reactions, the addition of SUMO E1, SUMO E2, and SUMO in the presence of ATP are sufficient to support basal level of Sgs1 sumoylation, since the SUMO E2 can transfer SUMO to substrates upon its activation by the SUMO E1 at the consumption of ATP (14,18,22). For simplicity, this reaction system is referred to as basal sumoylation reaction hereafter.…”

“…Similarly, BLM sumoylation depends on the Mms21 homolog in human cells (17). In yeast, STR sumoylation is additionally promoted by the conserved scaffold protein Esc2 (18). Esc2 contains two SUMO-like domains (SLDs) and only the C-terminal SLD2 binds to the SUMO E2 enzyme (Ubc9) and promotes sumoylation (18).…”

“…In yeast, STR sumoylation is additionally promoted by the conserved scaffold protein Esc2 (18). Esc2 contains two SUMO-like domains (SLDs) and only the C-terminal SLD2 binds to the SUMO E2 enzyme (Ubc9) and promotes sumoylation (18). In addition to Esc2, in vivo STR sumoylation shows a dependency on HJ structures that are present at low levels in cells (14).…”