Cell cycle–dependent chromatin loading of the Fanconi anemia core complex by FANCM/FAAP24

Abstract

IntroductionThe 13 identified Fanconi anemia (FA) proteins cooperate in a common cellular pathway regulating the cellular response to DNA cross-linking agents, such as cisplatin (CDDP), diepoxybutane (DEB), and mitomycin C (MMC). 1 Of these FA proteins, 8 (A, B, C, E, F, G, L, and M) are assembled into a core complex, 2,3 which contains a ubiquitin E3 ligase activity (FANCL subunit) 4 and a DNA translocase activity (FANCM). 5 In response to DNA damage, or during S-phase progression, the FA core complex coordinately monoubiquitinates 2 downstream substrates, FANCD2 6,7 and FANCI. [8][9][10] These monoubiquitinated proteins subsequently translocate to the chromatin where they are believed to interact with additional downstream FA proteins, including FANCJ/BRIP1, [11][12][13] FANCD1/BRCA2, 14 and FANCN/PALB2. 15,16 These downstream proteins then regulate homologous recombination (HR) repair. Disruption of any of the proteins in the FA pathway accounts for the common cellular and clinical phenotype of FA patients. 17 How the pathway participates in the process of DNA cross-link repair remains unknown. 18 Some FA complementation groups also exhibit additional phenotypic variation, suggesting that some FA genes have functions outside a simple linear FA pathway. [19][20][21][22] The FA core complex may have additional functions beyond the monoubiquitination of FANCD2 and FANCI. 8 A FANCD2-Ub linear fusion protein complements the MMC hypersensitivity of Fancd2-deficient chicken cells, but fails to correct the phenotype of FA core complex-deficient cells. 23 The FA core complex may therefore have additional activities, such as the recognition of specific DNA substrates, the regulation of the DNA replication machinery, and/or the monoubiquitination of additional (unknown) substrates. These additional functions may be explained, at least in part, by the presence of FA core subcomplexes with variable sizes and variable subcellular distributions. 3 When a replication fork encounters an interstrand DNA crosslink during replication, the replication fork arrests near the lesion, resulting in aberrant DNA structures. These abnormal structures activate checkpoint and repair pathways. FA cells, carrying mutations in FA genes, are highly sensitive to DNA crosslinking agents, compared with other DNA-damaging agents, such as ionizing radiation (IR), ultraviolet (UV), and hydroxyurea (HU). This hypersensitivity suggests that some components of the FA core complex may be involved in detecting and binding the DNA lesions caused by treatment of DNA crosslinking agents.The recently identified FANCM 5 and FANCJ 11-13 proteins suggest a direct involvement of FA proteins at sites of DNA repair. FANCM is homologous to the archaeal protein Hef (helicaseassociated endonuclease for fork-structured DNA), and is a member of the XP-F superfamily. 24 The N-terminal region of FANCM is able to bind to single-stranded DNA. 25 Moreover, FANCM has a DNA-dependent ATPase activity, and it can dissociate DNA triple helices in vitro. 5,25 FANCJ/BRIP1, whi...

The Fanconi Anemia Protein FANCM Is Controlled by FANCD2 and the ATR/ATM Pathways

“…xFANCM Is a Chromatin-binding Phosphoprotein-Recent studies showed that human FANCM binds to chromatin and is phosphorylated during the S phase of the cell cycle and in response to DNA-damaging agents like mitomycin C or hydroxyurea (8,12). To analyze xFANCM chromatin recruitment during a synchronous S phase, we incubated Xenopus sperm chromatin in egg extracts that allow for nuclear assembly and DNA replication under natural cell cycle control.…”

“…In human cells, FANCM localizes to chromatin and is required for chromatin recruitment of other FA core complex proteins (8,12). FANCM is phosphorylated during both the M and S phases and in response to DNA-damaging agents (8,12,13). Interestingly, DNA damage-induced phosphorylation of FANCM is independent of the FA core complex (8), suggesting that FANCM is controlled by other, as yet unknown upstream components of the DNA damage response.…”

“…Moreover, FANCM binds Holliday junctions and DNA replication fork structures in vitro and promotes ATP-dependent branch point migration, suggesting that FANCM might be involved in DNA processing at stalled replication forks (10,11). In human cells, FANCM localizes to chromatin and is required for chromatin recruitment of other FA core complex proteins (8,12). FANCM is phosphorylated during both the M and S phases and in response to DNA-damaging agents (8,12,13).…”

Protein monoubiquitylation: targets and diverse functions

“…Another two of these proteins, FANCD2 (at K561) and FANCI (at K523), collectively referred to as ID2, are monoubiquitylated in response to ICL (Table 2), which is detected on the basis of the association of FANCM with a stalled replication fork in S phase (Kim et al . 2008). FANCL contains a RING finger domain and is the catalytic subunit of the FA core complex.…”

Papel das proteínas XPD e DNA polimerase eta nas respostas de células humanas a danos no genoma

“…Entretanto, enquanto o reparo por HR demora aproximadamente sete horas, o reparo por NHEJ demora em média trinta minutos (Mao et al, 2008) (Räschle et al, 2008). O reparo dessas lesões é iniciado pelo reconhecimento da lesão, através da parada na forquilha de replicação, pela proteína FANCM (Kim et al, 2008) (Wu e Hickson, 2003). As proteínas BRCA1 e BRCA2 (FANCD1) também são recrutadas pelas proteínas FANC (Garcia-Higuera et al, 2001) e estão envolvidas na coordenação dos processos de reparo por recombinação necessários para a estabilização da forquilha bloqueada e continuação do reparo na fase S, também dependentes de RAD51 e das polimerases REV1 (Mirchandani et al, 2008),  (Moldovan et al, 2010) e  (REV7/REV3L).…”