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Review
. 2010 Mar 15;24(6):521-36.
doi: 10.1101/gad.1903510. Epub 2010 Mar 4.

GEN1/Yen1 and the SLX4 complex: Solutions to the problem of Holliday junction resolution

Affiliations
Review

GEN1/Yen1 and the SLX4 complex: Solutions to the problem of Holliday junction resolution

Jennifer M Svendsen et al. Genes Dev. .

Abstract

Chromosomal double-strand breaks (DSBs) are considered to be among the most deleterious DNA lesions found in eukaryotic cells due to their propensity to promote genome instability. DSBs occur as a result of exogenous or endogenous DNA damage, and also occur during meiotic recombination. DSBs are often repaired through a process called homologous recombination (HR), which employs the sister chromatid in mitotic cells or the homologous chromosome in meiotic cells, as a template for repair. HR frequently involves the formation and resolution of four-way DNA structures referred to as the Holliday junction (HJ). Despite extensive study, the machinery and mechanisms used to process these structures in eukaryotes have remained poorly understood. Recent work has identified XPG and UvrC/GIY domain-containing structure-specific endonucleases that can symmetrically cleave HJs in vitro in a manner that allows for religation without additional processing, properties that are reminiscent of the classical RuvC HJ resolvase in bacteria. Genetic studies reveal potential roles for these HJ resolvases in repair after DNA damage and during meiosis. The stage is now set for a more comprehensive understanding of the specific roles these enzymes play in the response of cells to DSBs, collapsed replication forks, telomere dysfunction, and meiotic recombination.

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Figures

Figure 1.
Figure 1.
Pathways for repairing DSBs via HR. (A) GC occurs at DSBs where both ends of the break are available for repair. It is subdivided into three pathways for repair: DSBR, SDSA, and dHJ dissolution. DSBR can generate either crossover or noncrossover products, while SDSA and HJ dissolution generate only noncrossover products. (B) SSA is the primary means of repair when DSBs are in highly repetitive regions. (C) BIR is employed when only one strand of a DSB is available for repair.
Figure 2.
Figure 2.
In vitro and in vivo activities of SSE that process HJs. (A) GEN1/Yen1 and the mammalian SLX1–SLX4 module contain classical HJ resolvase activities and can cleave static HJs symmetrically like the classical HJ resolvase RuvC. Recombinant MUS81–EME1 does not cleave static HJs, but has robust nHJ cleavage activity. (B) During meiosis, HJs are processed to produce crossover products. In budding yeast, Mus81–Mms4 and Yen1 have redundant roles in meiosis, while in fission yeast, Mus81–Eme1 is required for almost all meiotic events. In Drosophila, the SLX4 ortholog MUS312 and the ERCC4/XPF ortholog MEI-9 work in concert to affect meiotic crossovers. In worms, the situation is more complex, with HIM-18, MUS-81, and XPF-1 responsible for subsets of meiotic events. In mammals, it is still unclear what SSEs are required during meiosis, although there is no obvious meiotic phenotype in mice lacking MUS81.
Figure 3.
Figure 3.
Structural features of classical HJ resolvases and the ERCC4 family of SSEs. (A,B) Structure of dimeric E. coli RuvC (PDB: 1HJR) and S. pombe Ydc2 (PDB: 1KCF) HJ resolvases. The positions of critical acidic residues important for binding divalent metal ions are shown. (C) Domain structures of members of the ERCC4 family of SSEs. ERCC4/XPF contains an ERCC4 domain that is conserved in ERCC1, MUS81, and EME1. ERCC4/XPF also contains a DEAH helicase domain not found in ERCC1, MUS81, or EME1. The C-terminal regions of these proteins also contain conserved tandem HhH motifs. (D) Crystal structure of a MUS81–EME1 complex (PDB: 2ZIU), with the ERCC4 domain of the MUS81 (zebrafish) subunit shown in red, and the HhH motifs shown in magenta. The EME1 protein (human) is shown in cyan. Catalytic residues (see the text for details) are shown in blue.
Figure 4.
Figure 4.
Structural analysis of XPG endonucleases. (A) GEN1 domain structure showing elements within the N-terminal XPG homology region. (B,C) Model of the GEN1 catalytic domain (B) based on the structure of FEN1 (C; PDB: 1B43), a flap endonuclease with an XPG-based catalytic site. The GEN1 model was generated using SWISS-MODEL. Color-coding of motifs within GEN1 is as diagrammed in A. Residues responsible for coordination of the primary and secondary catalytic magnesium ions are in blue and red, respectively.
Figure 5.
Figure 5.
Versatility of the SLX4 complex. (A) The NER endonuclease UvrC as well as SLX1 orthologs contain a GIY motif, also referred to as an Uri domain. The structure of the UvrC domain is shown along with conserved residues (red) involved in binding to a hydrated divalent metal ion (magnesium or manganese, beige sphere). While UvrC has a C-terminal HhH motif, SLX1 contains a PHD-like ring finger motif. (B) Structural anatomy of the SLX4 complex in mammals and its relationship with SLX4 complexes in budding yeast. (C) In vitro cleavage specificity of the SLX4 complex. Bars and arrows represent regions of cleavage. (D) Functions of the yeast, Drosophila, C. elegans, and human SLX4 complexes in vivo.
Figure 6.
Figure 6.
Model depicting distinct roles for SLX1 and MUS81–EME1 within the SLX4 complex. SLX4 complexes contain two HJ processing activities. SLX1–SLX4 behaves as a classical HJ resolvase and symmetrically cleaves HJs to generate nicked ligatable DNA duplexes in vitro. Alternatively, SLX1 may cleave HJs asymmetrically to generate a nHJ that is a suitable substrate for MUS81–EME1. This results in flapped and gapped DNA duplex products that cannot be ligated without further processing.
Figure 7.
Figure 7.
Potential roles for the SLX4 complex during DSBR. (A) In an alternate model for DSBR initially proposed by Osman et al. (2003). SLX1–SLX4 may cleave the HJ-like intermediate formed by strand exchange, and MUS81–EME1 may work downstream to cleave the nHJ generated by second end capture. This mechanism would produce crossover products that are desirable during meiosis. (B) Alternatively, SLX1–SLX4 and/or GEN1 complexes may cleave dHJs during traditional DSBR to generate crossover and/or noncrossover products. Arrowheads represent possible sites of cleavage by SSEs.

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