Fanconi Anemia FANCG Protein in Mitigating Radiation- and Enzyme-Induced DNA Double-Strand Breaks by Homologous Recombination in Vertebrate Cells

Abstract

The rare hereditary disorder Fanconi anemia (FA) is characterized by progressive bone marrow failure, congenital skeletal abnormality, elevated susceptibility to cancer, and cellular hypersensitivity to DNA crosslinking chemicals and sometimes other DNA-damaging agents. Molecular cloning identified six causative genes (FANCA, -C, -D2, -E, -F, and -G) encoding a multiprotein complex whose precise biochemical function remains elusive. Recent studies implicate this complex in DNA damage responses that are linked to the breast cancer susceptibility proteins BRCA1 and BRCA2. Mutations in BRCA2, which participates in homologous recombination (HR), are the underlying cause in some FA patients. To elucidate the roles of FA genes in HR, we disrupted the FANCG/XRCC9 locus in the chicken B-cell line DT40. FANCG-deficient DT40 cells resemble mammalian fancg mutants in that they are sensitive to killing by cisplatin and mitomycin C (MMC) and exhibit increased MMC and radiation-induced chromosome breakage. We find that the repair of I-SceI-induced chromosomal double-strand breaks (DSBs) by HR is decreased ϳ9-fold in fancg cells compared with the parental and FANCG-complemented cells. In addition, the efficiency of gene targeting is mildly decreased in FANCGdeficient cells, but depends on the specific locus. We conclude that FANCG is required for efficient HRmediated repair of at least some types of DSBs.

The Fanconi Anemia/BRCA pathway: new faces in the crowd

“…Although the purpose of FANCD2 monoubiquitination remains unknown, it seems likely that it has a role in HR as BRCA1, BRCA2, RAD51, and NBS1 have all been implicated in this process (Moynahan et al 1999;Tutt et al 2001;Kobayashi et al 2004). In support of this hypothesis, cell lines deficient in the FA pathway have been reported to have defective HR (Donahue et al 2003;Yamamoto et al 2003;Niedzwiedz et al 2004;Nakanishi et al 2005). In general, disruption of upstream FA proteins, such as FANCA and FANCG (Yamamoto et al 2003) result in milder HR defects than disruption of BRCA2/FANCD1 (Moynahan et al 2001b), suggesting that the FA chromatin-associated complex 2 is largely responsible for the DNA repair functions of the FA pathway.…”

“…In support of this hypothesis, cell lines deficient in the FA pathway have been reported to have defective HR (Donahue et al 2003;Yamamoto et al 2003;Niedzwiedz et al 2004;Nakanishi et al 2005). In general, disruption of upstream FA proteins, such as FANCA and FANCG (Yamamoto et al 2003) result in milder HR defects than disruption of BRCA2/FANCD1 (Moynahan et al 2001b), suggesting that the FA chromatin-associated complex 2 is largely responsible for the DNA repair functions of the FA pathway. Interestingly, recent evidence suggests that DNA cross-links may be preferentially repaired by the singlestrand annealing subset of HR (Jonnalagadda et al 2005).…”

Fanconi anaemia genes and susceptibility to cancer

“…However, much remains to be done to define the functions of these proteins and to understand how they contribute to DNA recombination and repair. The use of proteomics to dissect the components of FA protein complexes (Meetei et al, 2005) and model organisms such as chicken DT40 cells to knockout combinations of FA genes (Yamamoto et al, 2003;Bridge et al, 2005;Mosedale et al, 2005) are beginning to yield valuable insights into how these proteins combine to recognize or repair DNA damage.…”

Identification of novel substrates for human checkpoint kinase Chk1 and Chk2 through genome-wide screening using a consensus Chk phosphorylation motif

“…For example, Rad51D is required for recombinational repair of damaged DNA (Takata et al, 2001). FANCG/XRCC9 is one of the Fanconi anemia genes and is involved in DNA double strand break (DSB) repair in S phase (Yamamoto et al, 2003).…”