Presynaptic filament dynamics in homologous recombination and DNA repair
Departments of Microbiology and of Molecular and Cellular Biology, University of California, Davis, CA, USA.Critical Reviews in Biochemistry and Molecular Biology (Impact Factor: 7.71). 06/2011; 46(3):240-70. DOI: 10.3109/10409238.2011.576007
Homologous recombination (HR) is an essential genome stability mechanism used for high-fidelity repair of DNA double-strand breaks and for the recovery of stalled or collapsed DNA replication forks. The crucial homology search and DNA strand exchange steps of HR are catalyzed by presynaptic filaments-helical filaments of a recombinase enzyme bound to single-stranded DNA (ssDNA). Presynaptic filaments are fundamentally dynamic structures, the assembly, catalytic turnover, and disassembly of which must be closely coordinated with other elements of the DNA recombination, repair, and replication machinery in order for genome maintenance functions to be effective. Here, we reviewed the major dynamic elements controlling the assembly, activity, and disassembly of presynaptic filaments; some intrinsic such as recombinase ATP-binding and hydrolytic activities, others extrinsic such as ssDNA-binding proteins, mediator proteins, and DNA motor proteins. We examined dynamic behavior on multiple levels, including atomic- and filament-level structural changes associated with ATP binding and hydrolysis as evidenced in crystal structures, as well as subunit binding and dissociation events driven by intrinsic and extrinsic factors. We examined the biochemical properties of recombination proteins from four model systems (T4 phage, Escherichia coli, Saccharomyces cerevisiae, and Homo sapiens), demonstrating how their properties are tailored for the context-specific requirements in these diverse species. We proposed that the presynaptic filament has evolved to rely on multiple external factors for increased multilevel regulation of HR processes in genomes with greater structural and sequence complexity.
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- "Human BRCA2 protein performs the classic RMP function of loading RAD51 protein onto RPA-covered ssDNA (Jensen et al. 2010; Liu et al. 2010). It appears that BRCA2 has superseded RAD52 as the major RMP used for presynpatic filament assembly and to promote DNA pairing in humans (Liu et al. 2011a). Unlike RAD52, BRCA2 does not promote the annealing of complementary RPA-covered ssDNA strands (Jensen et al. 2010). "
ABSTRACT: The formation of heteroduplex DNA is a central step in the exchange of DNA sequences via homologous recombination, and in the accurate repair of broken chromosomes via homology-directed repair pathways. In cells, heteroduplex DNA largely arises through the activities of recombination proteins that promote DNA-pairing and annealing reactions. Classes of proteins involved in pairing and annealing include RecA-family DNA-pairing proteins, single-stranded DNA (ssDNA)-binding proteins, recombination mediator proteins, annealing proteins, and nucleases. This review explores the properties of these pairing and annealing proteins, and highlights their roles in complex recombination processes including the double Holliday junction (DhJ) formation, synthesis-dependent strand annealing, and single-strand annealing pathways-DNA transactions that are critical both for genome stability in individual organisms and for the evolution of species. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.
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- "The ADP-bound form of the filament, considered to be the inactive form, has a low helical pitch of 65–85Å85˚85Å. In addition to regulation by the ATPase cycle, presynaptic filament dynamics are regulated by mediator proteins including BRCA2 and RAD51 paralogs, by DNA helicases and motor proteins including BLM and RAD54, and likely by signaling proteins and post-translational modifications (19,22,23). As new genetic determinants of cancer are continually identified, it is striking that many involve mutations affecting proteins that are regulators of RAD51, or even act directly as mediators of RAD51-catalyzed reactions (24,25). "
ABSTRACT: Human RAD51 protein catalyzes DNA pairing and strand exchange reactions that are central to homologous recombination and homology-directed DNA repair. Successful recombination/repair requires the formation of a presynaptic filament of RAD51 on ssDNA. Mutations in BRCA2 and other proteins that control RAD51 activity are associated with human cancer. Here we describe a set of mutations associated with human breast tumors that occur in a common structural motif of RAD51. Tumor-associated D149N, R150Q and G151D mutations map to a Schellman loop motif located on the surface of the RecA homology domain of RAD51. All three variants are proficient in DNA strand exchange, but G151D is slightly more sensitive to salt than wild-type (WT). Both G151D and R150Q exhibit markedly lower catalytic efficiency for adenosine triphosphate hydrolysis compared to WT. All three mutations alter the physical properties of RAD51 nucleoprotein filaments, with G151D showing the most dramatic changes. G151D forms mixed nucleoprotein filaments with WT RAD51 that have intermediate properties compared to unmixed filaments. These findings raise the possibility that mutations in RAD51 itself may contribute to genome instability in tumor cells, either directly through changes in recombinase properties, or indirectly through changes in interactions with regulatory proteins. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.
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ABSTRACT: Across the evolutionary spectrum, living organisms depend on high-fidelity DNA replication and recombination mechanisms to maintain genome stability and thus to avoid mutation and disease. The repair of severe lesions in the DNA such as double-strand breaks or stalled replication forks requires the coordinated activities of both the homologous recombination (HR) and DNA replication machineries. Growing evidence indicates that so-called "accessory proteins" in both systems are essential for the effective coupling of recombination to replication which is necessary to restore genome integrity following severe DNA damage. In this article we review the major processes of homology-directed DNA repair (HDR), including the double Holliday Junction (dHJ), synthesis-dependent strand annealing (SDSA), break-induced replication (BIR), and error-free lesion bypass pathways. Each of these pathways involves the coupling of a HR event to DNA synthesis. We highlight two major classes of accessory proteins in recombination and replication that facilitate HDR: Recombination mediator proteins exemplified by T4 UvsY, Saccharomyces cerevisiae Rad52, and human BRCA2; and DNA helicases/translocases exemplified by T4 Gp41/Gp59, E. coli DnaB and PriA, and eukaryotic Mcm2-7, Rad54, and Mph1. We illustrate how these factors help to direct the flow of DNA and protein-DNA intermediates on the pathway from a double-strand break or stalled replication fork to a high-fidelity recombination-dependent replication apparatus that can accurately repair the damage.
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