Activation of the Slx5-Slx8 ubiquitin ligase by poly-small ubiquitin-like modifier conjugates.
Protein sumoylation is a regulated process that is important for the health of human and yeast cells. In budding yeast, a subset of sumoylated proteins is targeted for ubiquitination by a conserved heterodimeric ubiquitin (Ub) ligase, Slx5-Slx8, which is needed to suppress the accumulation of high molecular weight small ubiquitin-like modifier (SUMO) conjugates. Structure-function analysis indicates that the Slx5-Slx8 complex contains multiple SUMO-binding domains that are collectively required for in vivo function. To determine the specificity of Slx5-Slx8, we assayed its Ub ligase activity using sumoylated Siz2 as an in vitro substrate. In contrast to unsumoylated or multisumoylated Siz2, substrates containing poly-SUMO conjugates were efficiently ubiquitinated by Slx5-Slx8. Although Siz2 itself was ubiquitinated, the bulk of the Ub was conjugated to SUMO residues. Slx5-Slx8 primarily mono-ubiquitinated the N-terminal SUMO moiety of the chain. These data indicate that the Slx5-Slx8 Ub ligase is stimulated by poly-SUMO conjugates and that it can ubiquitinate a poly-SUMO chain.
Mullen
Janet R
JR
Department of Molecular Biology and Biochemistry, Rutgers University, Piscataway, NJ 08854, USA.
Brill
Steven J
SJ
D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D025842:SUMO-1 Protein / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D025841:Small Ubiquitin-Related Modifier Proteins; D013379:Substrate Specificity; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D012097:Repressor Proteins; C099362:SMT3 protein, S cerevisiae; D025842:SUMO-1 Protein; D029701:Saccharomyces cerevisiae Proteins; C436836:Siz2 protein, S cerevisiae; D025841:Small Ubiquitin-Related Modifier Proteins; C515574:Slx8 protein, S cerevisiae; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; C515573:Slx5 protein, S cerevisiae