title: A requirement of FancL and FancD2 monoubiquitination in DNA repair.
abstract: The rare hereditary disorder Fanconi anemia (FA) can be caused by mutations in components of the FA core complex (FancA/B/C/E/F/G/L/M), a key regulator FancD2, the breast cancer susceptibility protein BRCA2/FancD1, or the newly identified FancJ/BRIP1 helicase. By performing yeast two-hybrid (Y2H) screens using N-terminal chicken (ch) FancD2 as a bait, we have identified chFancL, the likely ubiquitin E3 ligase subunit of the FA core complex. We also found that ectopically expressed FancD2 and FancL co-immunoprecipitated in 293T cells, and this interaction was dependent on the PHD domain of FancL. FANCL-disrupted chicken DT40 cells displayed defects in both FancD2 monoubiquitination and focus formation. Importantly, cell lines lacking the FANCL or FANCD2 genes, or carrying a "knock-in" mutation of the FancD2 monoubiquitination site (where the Lys 563 residue is changed to Arg), displayed quantitatively identical defects in the repair of I-SceI-induced chromosomal breaks by homologous recombination (HR). These data establish the role of FANCL and FancD2 monoubiquitination in HR repair.
authors.affiliation: Department of Immunology and Molecular Genetics, Kawasaki Medical School, Kurashiki, Okayama 701-0192, Japan.
authors.forename: Sohsuke
authors.initials: S
authors.lastname: Seki
authors.forename: Mioko
authors.initials: M
authors.lastname: Ohzeki
authors.forename: Akiko
authors.initials: A
authors.lastname: Uchida
authors.forename: Seiki
authors.initials: S
authors.lastname: Hirano
authors.forename: Nobuko
authors.initials: N
authors.lastname: Matsushita
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kitao
authors.forename: Tsukasa
authors.initials: T
authors.lastname: Oda
authors.forename: Takayuki
authors.initials: T
authors.lastname: Yamashita
authors.forename: Naoki
authors.initials: N
authors.lastname: Kashihara
authors.forename: Akio
authors.initials: A
authors.lastname: Tsubahara
authors.forename: Minoru
authors.initials: M
authors.lastname: Takata
authors.forename: Masamichi
authors.initials: M
authors.lastname: Ishiai
mesh_terms: D000818:Animals; D002460:Cell Line; D002645:Chickens; D004260:DNA Repair / Q000235:genetics / Q000502:physiology*; D005199:Fanconi Anemia / Q000235:genetics / Q000378:metabolism; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D052240:Fanconi Anemia Complementation Group L Protein / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D009154:Mutation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D025801:Ubiquitin / Q000378:metabolism
chemical_list: D052236:Fanconi Anemia Complementation Group D2 Protein; D011994:Recombinant Proteins; D025801:Ubiquitin; D052240:Fanconi Anemia Complementation Group L Protein