RNF4-mediated polyubiquitination regulates the Fanconi anemia/BRCA pathway

J Clin Invest. 2015 Apr;125(4):1523-32. doi: 10.1172/JCI79325. Epub 2015 Mar 9.

Abstract

The Fanconi anemia/BRCA (FA/BRCA) pathway is a DNA repair pathway that is required for excision of DNA interstrand cross-links. The 17 known FA proteins, along with several FA-associated proteins (FAAPs), cooperate in this pathway to detect, unhook, and excise DNA cross-links and to subsequently repair the double-strand breaks generated in the process. In the current study, we identified a patient with FA with a point mutation in FANCA, which encodes a mutant FANCA protein (FANCAI939S). FANCAI939S failed to bind to the FAAP20 subunit of the FA core complex, leading to decreased stability. Loss of FAAP20 binding exposed a SUMOylation site on FANCA at amino acid residue K921, resulting in E2 SUMO-conjugating enzyme UBC9-mediated SUMOylation, RING finger protein 4-mediated (RNF4-mediated) polyubiquitination, and proteasome-mediated degradation of FANCA. Mutation of the SUMOylation site of FANCA rescued the expression of the mutant protein. Wild-type FANCA was also subject to SUMOylation, RNF4-mediated polyubiquitination, and degradation, suggesting that regulated release of FAAP20 from FANCA is a critical step in the normal FA pathway. Consistent with this model, cells lacking RNF4 exhibited interstrand cross-linker hypersensitivity, and the gene encoding RNF4 was epistatic with the other genes encoding members of the FA/BRCA pathway. Together, the results from our study underscore the importance of analyzing unique patient-derived mutations for dissecting complex DNA repair processes.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adult
  • BRCA1 Protein / physiology*
  • Cell Line, Tumor
  • DNA Repair / genetics
  • DNA Repair / physiology*
  • Fanconi Anemia / genetics*
  • Fanconi Anemia Complementation Group A Protein / genetics
  • Fanconi Anemia Complementation Group A Protein / physiology*
  • Fanconi Anemia Complementation Group Proteins / antagonists & inhibitors
  • Fanconi Anemia Complementation Group Proteins / genetics
  • Fanconi Anemia Complementation Group Proteins / physiology
  • Female
  • Genes, BRCA1
  • Humans
  • Mutation, Missense
  • Nuclear Proteins / physiology*
  • Pedigree
  • Phenotype
  • Point Mutation
  • Proteasome Endopeptidase Complex / metabolism
  • Protein Binding
  • Protein Processing, Post-Translational / physiology
  • Proteolysis
  • RNA, Small Interfering / pharmacology
  • Signal Transduction / physiology
  • Sumoylation
  • Transcription Factors / physiology*
  • Triple Negative Breast Neoplasms / genetics
  • Ubiquitination / physiology

Substances

  • BRCA1 Protein
  • BRCA1 protein, human
  • FAAP20 protein, human
  • FANCA protein, human
  • Fanconi Anemia Complementation Group A Protein
  • Fanconi Anemia Complementation Group Proteins
  • Nuclear Proteins
  • RNA, Small Interfering
  • RNF4 protein, human
  • Transcription Factors
  • Proteasome Endopeptidase Complex