| Autor: |
Camilla S. Colding-Christensen, Ellen S. Kakulidis, Javier Arroyo-Gomez, Ivo A. Hendriks, Connor Arkinson, Zita Fábián, Agnieszka Gambus, Niels Mailand, Julien P. Duxin, Michael L. Nielsen |
| Rok vydání: |
2023 |
| Popis: |
Ubiquitin widely modifies proteins, thereby regulating most cellular functions. The complexity of ubiquitin signalling necessitates unbiased methods enabling global detection of dynamic protein ubiquitylation. Here, we describe UBIMAX (UBiquitin target Identification byMass spectrometry inXenopusegg extracts), which enriches ubiquitin-conjugated proteins and quantifies regulation of protein ubiquitylation under precise and adaptable conditions. We benchmark UBIMAX by investigating DNA double-strand break-responsive ubiquitylation events, identifying previously known targets and revealing the actin-organising protein Dbn1 as a novel major target of DNA damage-induced ubiquitylation. We find that Dbn1 is targeted for proteasomal degradation by the SCFβ-Trcp1ubiquitin ligase, in a conserved mechanism driven by ATM-mediated phosphorylation of a previously uncharacterized β-Trcp1 degron containing an SQ motif. We further show that this degron is sufficient to induce DNA-damage dependent protein degradation of a model substrate. Collectively, we demonstrate UBIMAX’s ability to identify novel targets of stimulus-regulated ubiquitylation and reveal an SCFβ-Trcp1-mediated ubiquitylation mechanism controlled directly by the apical DNA damage response kinases. |
| Databáze: |
OpenAIRE |
| Externí odkaz: |
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