Site-specific inhibition of the small ubiquitin-like modifier (SUMO)-conjugating enzyme Ubc9 selectively impairs SUMO chain formation.
Posttranslational modifications by small ubiquitin-like modifiers (SUMOs) regulate many cellular processes, including genome integrity, gene expression, and ribosome biogenesis. The E2-conjugating enzyme Ubc9 catalyzes the conjugation of SUMOs to ϵ-amino groups of lysine residues in target proteins. Attachment of SUMO moieties to internal lysines in Ubc9 itself can further lead ... to the formation of polymeric SUMO chains. Mono- and poly-SUMOylations of target proteins provide docking sites for distinct adapter and effector proteins important for regulating discrete SUMO-regulated pathways. However, molecular tools to dissect pathways depending on either mono- or poly-SUMOylation are largely missing. Using a protein-engineering approach, we generated high-affinity SUMO2 variants by phage display that bind the back side binding site of Ubc9 and function as SUMO-based Ubc9 inhibitors (SUBINs). Importantly, we found that distinct SUBINs primarily inhibit poly-SUMO chain formation, whereas mono-SUMOylation was not impaired. Proof-of-principle experiments demonstrated that in a cellular context, SUBINs largely prevent heat shock-triggered poly-SUMOylation. Moreover, SUBINs abrogated arsenic-induced degradation of promyelocytic leukemia protein. We propose that the availability of the new chain-selective SUMO inhibitors reported here will enable a thorough investigation of poly-SUMO-mediated cellular processes, such as DNA damage responses and cell cycle progression.
Mesh Terms:
Amino Acid Substitution, Arsenic, Binding Sites, Binding, Competitive, Gene Deletion, Gene Library, HEK293 Cells, HeLa Cells, Hot Temperature, Humans, Ligands, Models, Molecular, Peptide Fragments, Point Mutation, Promyelocytic Leukemia Protein, Protein Interaction Domains and Motifs, RNA Interference, Recombinant Fusion Proteins, Recombinant Proteins, Small Ubiquitin-Related Modifier Proteins, Sumoylation, Ubiquitin-Conjugating Enzymes
Amino Acid Substitution, Arsenic, Binding Sites, Binding, Competitive, Gene Deletion, Gene Library, HEK293 Cells, HeLa Cells, Hot Temperature, Humans, Ligands, Models, Molecular, Peptide Fragments, Point Mutation, Promyelocytic Leukemia Protein, Protein Interaction Domains and Motifs, RNA Interference, Recombinant Fusion Proteins, Recombinant Proteins, Small Ubiquitin-Related Modifier Proteins, Sumoylation, Ubiquitin-Conjugating Enzymes
J. Biol. Chem.
Date: Dec. 15, 2016
PubMed ID: 28784659
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