ATP-dependent and ATP-independent Roles for the Rad54 Chromatin Remodeling Enzyme during Recombinational Repair of a DNA Double Strand Break
18 Mar 2005-Journal of Biological Chemistry (American Society for Biochemistry and Molecular Biology)-Vol. 280, Iss: 11, pp 10855-10860
TL;DR: A novel Rad54-dependent chromatin remodeling event that occurs in vivo during the DNA strand invasion step of HR is identified, and this ATP-dependent remodeling activity of Rad54 appears to control subsequent steps in the HR process.
About: This article is published in Journal of Biological Chemistry.The article was published on 2005-03-18 and is currently open access. It has received 86 citations till now. The article focuses on the topics: Strand invasion & Replication protein A.
Citations
More filters
TL;DR: Mechanistic aspects of HR relating to DSB and ICL repair as well as replication fork support related to DNA double-stranded breaks and interstrand crosslinks are reviewed.
Abstract: Homologous recombination (HR) comprises a series of interrelated pathways that function in the repair of DNA double-stranded breaks (DSBs) and interstrand crosslinks (ICLs). In addition, recombination provides critical support for DNA replication in the recovery of stalled or broken replication forks, contributing to tolerance of DNA damage. A central core of proteins, most critically the RecA homolog Rad51, catalyzes the key reactions that typify HR: homology search and DNA strand invasion. The diverse functions of recombination are reflected in the need for context-specific factors that perform supplemental functions in conjunction with the core proteins. The inability to properly repair complex DNA damage and resolve DNA replication stress leads to genomic instability and contributes to cancer etiology. Mutations in the BRCA2 recombination gene cause predisposition to breast and ovarian cancer as well as Fanconi anemia, a cancer predisposition syndrome characterized by a defect in the repair of DNA interstrand crosslinks. The cellular functions of recombination are also germane to DNA-based treatment modalities of cancer, which target replicating cells by the direct or indirect induction of DNA lesions that are substrates for recombination pathways. This review focuses on mechanistic aspects of HR relating to DSB and ICL repair as well as replication fork support.
932 citations
TL;DR: It is reported that both the RSC and Swi/Snf ATP-dependent chromatin-remodeling complexes play key roles in double-strand break (DSB) repair, specifically by homologous recombination (HR).
Abstract: The failure of cells to repair damaged DNA can result in genomic instability and cancer. To efficiently repair chromosomal DNA lesions, the repair machinery must gain access to the damaged DNA in the context of chromatin. Here we report that both the RSC and Swi/Snf ATP-dependent chromatin-remodeling complexes play key roles in double-strand break (DSB) repair, specifically by homologous recombination (HR). RSC and Swi/Snf are each recruited to an in vivo DSB site but with distinct kinetics. We show that Swi/Snf is required earlier, at or preceding the strand invasion step of HR, while RSC is required following synapsis for completion of the recombinational repair event.
312 citations
Cites background from "ATP-dependent and ATP-independent R..."
...ATP-dependent nucleosome-remodeling factors are also implicated in repair: The Rad54p enzyme functions in multiple steps of DSB repair by HR (Alexiadis and Kadonaga 2002; Alexeev et al. 2003; Wolner and Peterson 2005), INO80 has recently been linked to DSB repair (Morrison et al....
[...]
TL;DR: It is proposed that sequestration of unrepaired or slowly repaired DSBs to the nuclear periphery reflects a competition between alternative repair pathways.
Abstract: DNA double-strand breaks (DSBs) are among the most deleterious forms of DNA lesions in cells. Here we induced site-specific DSBs in yeast cells and monitored chromatin dynamics surrounding the DSB using Chromosome Conformation Capture (3C). We find that formation of a DSB within G1 cells is not sufficient to alter chromosome dynamics. However, DSBs formed within an asynchronous cell population result in large decreases in both intra- and interchromosomal interactions. Using live cell microscopy, we find that changes in chromosome dynamics correlate with relocalization of the DSB to the nuclear periphery. Sequestration to the periphery requires the nuclear envelope protein, Mps3p, and Mps3p-dependent tethering delays recombinational repair of a DSB and enhances gross chromosomal rearrangements. Furthermore, we show that components of the telomerase machinery are recruited to a DSB and that telomerase recruitment is required for its peripheral localization. Based on these findings, we propose that sequestration of unrepaired or slowly repaired DSBs to the nuclear periphery reflects a competition between alternative repair pathways.
299 citations
Cites methods from "ATP-dependent and ATP-independent R..."
...ChIP was carried out essentially as described previously (Wolner and Peterson 2005)....
[...]
...Total genomic DNA prepared by glass bead lysis (Wolner and Peterson 2005) was loaded onto 1% agarose gels and electrophoresed for 3 h at 120 V before capillary transfer to a Nylon membrane (Stratagene)....
[...]
...Hence, PCR with a radioactive nucleotide was used to detect formation of this product as described (Wolner and Peterson 2005)....
[...]
TL;DR: A wealth of genetic, cytological, biochemical and structural data suggests that Rad54 is a core factor of HR, possibly acting at multiple stages during HR in concert with the central homologous pairing protein Rad51.
Abstract: Homologous recombination (HR) is a ubiquitous cellular pathway that mediates transfer of genetic information between homologous or near homologous (homeologous) DNA sequences. During meiosis it ensures proper chromosome segregation in the first division. Moreover, HR is critical for the tolerance and repair of DNA damage, as well as in the recovery of stalled and broken replication forks. Together these functions preserve genomic stability and assure high fidelity transmission of the genetic material in the mitotic and meiotic cell divisions. This review will focus on the Rad54 protein, a member of the Snf2-family of SF2 helicases, which translocates on dsDNA but does not display strand displacement activity typical for a helicase. A wealth of genetic, cytological, biochemical and structural data suggests that Rad54 is a core factor of HR, possibly acting at multiple stages during HR in concert with the central homologous pairing protein Rad51.
246 citations
Cites background or methods from "ATP-dependent and ATP-independent R..."
...Chromatin immunoprecipitation (ChIP) experiments have been used to monitor the recruitment of Rad51 protein to an HO endonuclease-induced DSB at the MAT locus and the HML donor locus in wild-type and rad54 mutant cells (53,57,83)....
[...]
...However, monitoring a positioned nucleosome on the HML donor site with micrococcal nuclease showed no difference between wildtype and rad54 cells (57), suggesting that Rad54 does not act by moving or removing this positioned nucleosome at the HML donor site....
[...]
...It is inferred from these experiments that Rad51 forms functional filaments in rad54 cells, because Rad51 was found targeted to the duplex donor locus (HML) by ChIP (57,83)....
[...]
...The noted difference in Rad51 localization to the HML donor site between wild-type and rad54 cells (53,57) may be a function of defects in filament assembly during pre-synapsis or defects in synapsis/ post-synapsis, which may affect Rad51–DNA complexes during these phases of recombination....
[...]
...Although Rad51 is targeted to the donor locus in rad54 cells, it is unclear if D-loops are formed and several studies were unable to detect DNA synthesis from an invading 30 end at the donor locus (53,57,83)....
[...]
TL;DR: In this paper, the main pathways of eukaryotic DNA double-strand break repair with the focus on homologous recombination and its various subpathways are reviewed.
Abstract: DNA double-strand breaks arise accidentally upon exposure of DNA to radiation and chemicals or result from faulty DNA metabolic processes. DNA breaks can also be introduced in a programmed manner, such as during the maturation of the immune system, meiosis, or cancer chemo- or radiotherapy. Cells have developed a variety of repair pathways, which are fine-tuned to the specific needs of a cell. Accordingly, vegetative cells employ mechanisms that restore the integrity of broken DNA with the highest efficiency at the lowest cost of mutagenesis. In contrast, meiotic cells or developing lymphocytes exploit DNA breakage to generate diversity. Here, we review the main pathways of eukaryotic DNA double-strand break repair with the focus on homologous recombination and its various subpathways. We highlight the differences between homologous recombination and end-joining mechanisms including non-homologous end-joining and microhomology-mediated end-joining and offer insights into how these pathways are regulated. Finally, we introduce noncanonical functions of the recombination proteins, in particular during DNA replication stress.
213 citations
References
More filters
[...]
TL;DR: The genomic sequencing procedures are applicable to the analysis of genetic polymorphisms, DNA methylation at deoxycytidines, and nucleic acid-protein interactions at single nucleotide resolution.
Abstract: Unique DNA sequences can be determined directly from mouse genomic DNA. A denaturing gel separates by size mixtures of unlabeled DNA fragments from complete restriction and partial chemical cleavages of the entire genome. These lanes of DNA are transferred and UV-crosslinked to nylon membranes. Hybridization with a short 32P-labeled single-stranded probe produces the image of a DNA sequence "ladder" extending from the 3' or 5' end of one restriction site in the genome. Numerous different sequences can be obtained from a single membrane by reprobing. Each band in these sequences represents 3 fg of DNA complementary to the probe. Sequence data from mouse immunoglobulin heavy chain genes from several cell types are presented. The genomic sequencing procedures are applicable to the analysis of genetic polymorphisms, DNA methylation at deoxycytidines, and nucleic acid-protein interactions at single nucleotide resolution.
7,858 citations
TL;DR: This review encompasses different aspects of DSB-induced recombination in Saccharomyces and attempts to relate genetic, molecular biological, and biochemical studies of the processes of DNA repair and recombination.
Abstract: The budding yeast Saccharomyces cerevisiae has been the principal organism used in experiments to examine genetic recombination in eukaryotes. Studies over the past decade have shown that meiotic recombination and probably most mitotic recombination arise from the repair of double-strand breaks (DSBs). There are multiple pathways by which such DSBs can be repaired, including several homologous recombination pathways and still other nonhomologous mechanisms. Our understanding has also been greatly enriched by the characterization of many proteins involved in recombination and by insights that link aspects of DNA repair to chromosome replication. New molecular models of DSB-induced gene conversion are presented. This review encompasses these different aspects of DSB-induced recombination in Saccharomyces and attempts to relate genetic, molecular biological, and biochemical studies of the processes of DNA repair and recombination.
2,175 citations
TL;DR: This work has divided the SNF2 family into multiple subfamilies, each of which represents what it proposes to be a functionally and evolutionarily distinct group, and used the subfamily structure to predict the functions of some of the uncharacterized proteins in the SNf2 family.
Abstract: The SNF2 family of proteins includes representatives from a variety of species with roles in cellular processes such as transcriptional regulation (e.g. MOT1, SNF2 and BRM), maintenance of chromosome stability during mitosis (e.g. lodestar) and various aspects of processing of DNA damage, including nucleotide excision repair (e.g. RAD16 and ERCC6), recombinational pathways (e.g. RAD54) and post-replication daughter strand gap repair (e.g. RAD5). This family also includes many proteins with no known function. To better characterize this family of proteins we have used molecular phylogenetic techniques to infer evolutionary relationships among the family members. We have divided the SNF2 family into multiple subfamilies, each of which represents what we propose to be a functionally and evolutionarily distinct group. We have then used the subfamily structure to predict the functions of some of the uncharacterized proteins in the SNF2 family. We discuss possible implications of this evolutionary analysis on the general properties and evolution of the SNF2 family.
734 citations
TL;DR: Rad52 functions as a co-factor for the Rad51 recombinase, acting specifically to overcome the apparent competition by RPA for binding to single-stranded DNA.
542 citations
TL;DR: In this article, a stable heterodimer of Rad51, Rad55, and Rad57 was found to exist as a stable product with a dissociation constant of <2 x 10−10 M.
Abstract: Saccharomyces cerevisiae RAD51, RAD55, and RAD57 genes, required for genetic recombination and DNA double-strand-break repair, encode proteins homologous to one another and to the Escherichia coli RecA protein. Rad51 protein catalyzes the DNA strand-exchange reaction with a dependence on ATP and on the heterotrimeric single-strand DNA (ssDNA) binding factor replication protein A (RPA). By several independent criteria, RAD55- and RAD57-encoded products are shown here to exist as a stable heterodimer, with a dissociation constant of <2 x 10(-10) M. In strand exchange, the reaction proceeds efficiently if RPA is incorporated after nucleation of Rad51 onto ssDNA, but if RPA is present during the nucleation phase, as is likely the case in vivo, the amount of strand-exchange products becomes relatively insignificant. Inclusion of the Rad55-Rad57 heterodimer with Rad51 and RPA results in a marked stimulation of strand exchange, providing evidence for a role of the Rad55-Rad57 heterodimer in overcoming the inhibitory effect of RPA.
497 citations