title: The FA Core Complex Contains a Homo-dimeric Catalytic Module for the Symmetric Mono-ubiquitination of FANCI-FANCD2.
abstract: Activation of the main DNA interstrand crosslink repair pathway in higher eukaryotes requires mono-ubiquitination of FANCI and FANCD2 by FANCL, the E3 ligase subunit of the Fanconi anemia core complex. FANCI and FANCD2 form a stable complex; however, the molecular basis of their ubiquitination is ill defined. FANCD2 mono-ubiquitination by FANCL is stimulated by the presence of the FANCB and FAAP100 core complex components, through an unknown mechanism. How FANCI mono-ubiquitination is achieved remains unclear. Here, we use structural electron microscopy, combined with crosslink-coupled mass spectrometry, to find that FANCB, FANCL, and FAAP100 form a dimer of trimers, containing two FANCL molecules that are ideally poised to target both FANCI and FANCD2 for mono-ubiquitination. The FANCC-FANCE-FANCF subunits bridge between FANCB-FANCL-FAAP100 and the FANCI-FANCD2 substrate. A transient interaction with FANCC-FANCE-FANCF alters the FANCI-FANCD2 configuration, stabilizing the dimerization interface. Our data provide a model to explain how equivalent mono-ubiquitination of FANCI and FANCD2 occurs.
authors.affiliation: Macromolecular Machines Laboratory, Clare Hall Laboratory, The Francis Crick Institute, Blanche Lane, South Mimms, EN6 3LD, UK.
authors.forename: Paolo
authors.initials: P
authors.lastname: Swuec
authors.affiliation: Macromolecular Machines Laboratory, Clare Hall Laboratory, The Francis Crick Institute, Blanche Lane, South Mimms, EN6 3LD, UK.
authors.forename: Ludovic
authors.initials: L
authors.lastname: Renault
authors.affiliation: Mass Spectrometry Proteomics and Metabolomics, Clare Hall Laboratory, The Francis Crick Institute, Blanche Lane, South Mimms, EN6 3LD, UK.
authors.forename: Aaron
authors.initials: A
authors.lastname: Borg
authors.affiliation: Genome Stability Unit, St. Vincent's Institute of Medical Research, 9 Princes St Fitzroy, Victoria, VIC 3065, Australia.
authors.forename: Fenil
authors.initials: F
authors.lastname: Shah
authors.affiliation: Genome Stability Unit, St. Vincent's Institute of Medical Research, 9 Princes St Fitzroy, Victoria, VIC 3065, Australia.
authors.forename: Vincent J
authors.initials: VJ
authors.lastname: Murphy
authors.affiliation: Genome Stability Unit, St. Vincent's Institute of Medical Research, 9 Princes St Fitzroy, Victoria, VIC 3065, Australia.
authors.forename: Sylvie
authors.initials: S
authors.lastname: van Twest
authors.affiliation: Mass Spectrometry Proteomics and Metabolomics, Clare Hall Laboratory, The Francis Crick Institute, Blanche Lane, South Mimms, EN6 3LD, UK.
authors.forename: Ambrosius P
authors.initials: AP
authors.lastname: Snijders
authors.affiliation: Genome Stability Unit, St. Vincent's Institute of Medical Research, 9 Princes St Fitzroy, Victoria, VIC 3065, Australia.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Deans
authors.affiliation: Macromolecular Machines Laboratory, Clare Hall Laboratory, The Francis Crick Institute, Blanche Lane, South Mimms, EN6 3LD, UK. Electronic address: alessandro.costa@crick.ac.uk.
authors.forename: Alessandro
authors.initials: A
authors.lastname: Costa
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051856:Fanconi Anemia Complementation Group Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D013058:Mass Spectrometry; D008854:Microscopy, Electron; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity; D054875:Ubiquitination
chemical_list: D004268:DNA-Binding Proteins; C523207:FAAP100 protein, human; C515775:FANCB protein, human; C499905:FANCD2 protein, human; C519921:FANCI protein, human; D052236:Fanconi Anemia Complementation Group D2 Protein; D051856:Fanconi Anemia Complementation Group Proteins
keywords: DNA repair; Fanconi anemia; cryo-electron microscopy; interstrand crosslink; mono-ubiquitination; ubiquitin ligase