RNF168 E3 ligase participates in ubiquitin signaling and recruitment of SLX4 during DNA crosslink repair

Cell Rep. 2021 Oct 26;37(4):109879. doi: 10.1016/j.celrep.2021.109879.

Abstract

SLX4/FANCP is a key Fanconi anemia (FA) protein and a DNA repair scaffold for incision around a DNA interstrand crosslink (ICL) by its partner XPF nuclease. The tandem UBZ4 ubiquitin-binding domains of SLX4 are critical for the recruitment of SLX4 to damage sites, likely by binding to K63-linked polyubiquitin chains. However, the identity of the ubiquitin E3 ligase that mediates SLX4 recruitment remains unknown. Using small interfering RNA (siRNA) screening with a GFP-tagged N-terminal half of SLX4 (termed SLX4-N), we identify the RNF168 E3 ligase as a critical factor for mitomycin C (MMC)-induced SLX4 foci formation. RNF168 and GFP-SLX4-N colocalize in MMC-induced ubiquitin foci. Accumulation of SLX4-N at psoralen-laser ICL tracks or of endogenous SLX4 at Digoxigenin-psoralen/UVA ICL is dependent on RNF168. Finally, we find that RNF168 is epistatic with SLX4 in promoting MMC tolerance. We conclude that RNF168 is a critical component of the signal transduction that recruits SLX4 to ICL damage.

Keywords: Fanconi anemia; RNF168; SLX4; interstrand crosslink repair; ubiquitination.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA Repair*
  • Digoxigenin / pharmacology
  • Ficusin / pharmacology
  • HCT116 Cells
  • Humans
  • MCF-7 Cells
  • Mitomycin / pharmacology
  • Recombinases / genetics
  • Recombinases / metabolism*
  • Signal Transduction*
  • Ubiquitin / genetics
  • Ubiquitin / metabolism*
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*

Substances

  • Recombinases
  • Ubiquitin
  • Mitomycin
  • RNF168 protein, human
  • Ubiquitin-Protein Ligases
  • SLX4 protein, human
  • Ficusin
  • Digoxigenin