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.

Homologous recombination (HR) is required for accurate chromosome segregation during the first meiotic division and constitutes a key repair and tolerance pathway for complex DNA damage, including DNA double-strand breaks, interstrand crosslinks, and DNA gaps. In addition, recombination and replication are inextricably linked, as recombination recovers stalled and broken replication forks, enabling the evolution of larger genomes/replicons. Defects in recombination lead to genomic instability and elevated cancer predisposition, demonstrating a clear cellular need for recombination. However, recombination can also lead to genome rearrangements. Unrestrained recombination causes undesired endpoints (translocation, deletion, inversion) and the accumulation of toxic recombination intermediates. Evidently, HR must be carefully regulated to match specific cellular needs. Here, we review the factors and mechanistic stages of recombination that are subject to regulation and suggest that recombination achieves flexibility and robustness by proceeding through metastable, reversible intermediates.

Homologous recombination enables the cell to access and copy intact DNA sequence information in trans, particularly to repair DNA damage affecting both strands of the double helix. Here, we discuss the DNA transactions and enzymatic activities required for this elegantly orchestrated process in the context of the repair of DNA double-strand breaks in somatic cells. This includes homology search, DNA strand invasion, repair DNA synthesis, and restoration of intact chromosomes. Aspects of DNA topology affecting individual steps are highlighted. Overall, recombination is a dynamic pathway with multiple metastable and reversible intermediates designed to achieve DNA repair with high fidelity.

BRCA2 is a tumor suppressor that interacts with RAD51 and functions in homologous recombination, but understanding its precise functions has been hampered by difficulties in purifying such a large protein. Now purified full-length human BRCA2 is shown to bind ∼6 molecules of RAD51 and to promote RAD51 binding to RPA-covered ssDNA in a manner stimulated by DSS1. BRCA2 is a tumor suppressor that functions in homologous recombination, a key genomic integrity pathway. BRCA2 interacts with RAD51, the central protein of recombination, which forms filaments on single-stranded DNA (ssDNA) to perform homology search and DNA strand invasion. We report the purification of full-length human BRCA2 and show that it binds to ∼6 RAD51 molecules and promotes RAD51 binding to ssDNA coated by replication protein A (RPA), in a manner that is stimulated by DSS1.

DNA strand exchange, a process that helps to maintain the integrity of the genome, is promoted in eukaryotes by the Rad51 protein. Eukaryotes also have homologues of Rad51 with functions that are less well defined. Wolf-Dietrich Heyer and colleagues show that in yeast, two of these proteins, Rad55 and Rad57, form a complex that promotes recombination by stabilizing the Rad51 filament against the anti-recombinogenic activity of a helicase, Srs2. These results suggest that the Rad51 filament is a reversible intermediate, the stability of which reflects competing actions of other proteins. This mechanism may have relevance for the human RAD51 paralogues that have important roles in tumour suppression and disease. Homologous recombination is a high-fidelity DNA repair pathway. Besides a critical role in accurate chromosome segregation during meiosis, recombination functions in DNA repair and in the recovery of stalled or broken replication forks to ensure genomic stability. In contrast, inappropriate recombination contributes to genomic instability, leading to loss of heterozygosity, chromosome rearrangements and cell death. The RecA/UvsX/RadA/Rad51 family of proteins catalyses the signature reactions of recombination, homology search and DNA strand invasion1,2. Eukaryotes also possess Rad51 paralogues, whose exact role in recombination remains to be defined3. Here we show that the Saccharomyces cerevisiae Rad51 paralogues, the Rad55–Rad57 heterodimer, counteract the antirecombination activity of the Srs2 helicase. The Rad55–Rad57 heterodimer associates with the Rad51–single-stranded DNA filament, rendering it more stable than a nucleoprotein filament containing Rad51 alone. The Rad51–Rad55–Rad57 co-filament resists disruption by the Srs2 antirecombinase by blocking Srs2 translocation, involving a direct protein interaction between Rad55–Rad57 and Srs2. Our results demonstrate an unexpected role of the Rad51 paralogues in stabilizing the Rad51 filament against a biologically important antagonist, the Srs2 antirecombination helicase. The biological significance of this mechanism is indicated by a complete suppression of the ionizing radiation sensitivity of rad55 or rad57 mutants by concomitant deletion of SRS2, as expected for biological antagonists. We propose that the Rad51 presynaptic filament is a meta-stable reversible intermediate, whose assembly and disassembly is governed by the balance between Rad55–Rad57 and Srs2, providing a key regulatory mechanism controlling the initiation of homologous recombination. These data provide a paradigm for the potential function of the human RAD51 paralogues, which are known to be involved in cancer predisposition and human disease.

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.