Molecular triage and isoform protection safeguard ubiquitin chain editing on DNA

In an accompanying study, we identify the RPA-ZUP1 complex as a ubiquitin chain-editing machine that promotes removal of RPA from ssDNA. Because loss of ZUP1 causes pathological RPA trapping on ssDNA, cells must preserve a functional RPA-ZUP1 pool to sustain RPA turnover. How this homeostasis is achieved is unknown. Here, we show that ZUP1 is subject to ubiquitin-dependent fate control. ZUP1 ubiquitination promotes recruitment of p97/VCP and ATXN3, defining a molecular triage pathway that protects ZUP1 from premature proteasomal degradation. Opposing this, the E3 ligases UBR5 and HUWE1 define a degradative branch that limits ZUP1 abundance, with HUWE1 restricting its accumulation on damaged chromatin. We further show that cells express a naturally occurring ZUP1 isoform resistant to efficient ubiquitination and turnover, suggesting that isoform diversity provides a protected pool of chain-editing activity. Together, these findings define a homeostatic system that preserves ZUP1 while limiting its persistence on damaged chromatin.