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In this study, the formation and repair of large (>1 kb) insertion/deletion (I/D) heterologies during double-strand-break repair (DSBR) was investigated using a gene-targeting assay that permits efficient recovery of sequence insertion events at the haploid chromosomal immunoglobulin (Ig) [micro]-locus in mouse hybridoma cells. The results revealed that (i) large I/D heterologies were generated on one or both sides of the DSB and, in some cases, formed symmetrically in both homology regions; (ii) large I/D heterologies did not negatively affect the gene targeting frequency; and (iii) prior to DNA replication, the large I/D heterologies were rectified.

The importance of the different RecQ homologos in genome maintenance in mammals is discussed. The results show that RECQL5 helicase is very beneficial in suppressing tumor via Rad51 presynaptic filaments disruption.

The conserved BTR complex, composed of the Bloom's syndrome helicase (BLM), topoisomerase IIIα, RMI1, and RMI2, regulates homologous recombination in favor of non-crossover formation via the dissolution of the double Holliday Junction (dHJ). Here we show enhancement of the BTR-mediated dHJ dissolution reaction by the heterotrimeric single-stranded DNA binding protein replication protein A (RPA). Our results suggest that RPA acts by sequestering a single-stranded DNA intermediate during dHJ dissolution. We provide evidence that RPA physically interacts with RMI1. The RPA interaction domain in RMI1 has been mapped, and RMI1 mutants impaired for RPA interaction have been generated. Examination of these mutants ascertains the significance of the RMI1-RPA interaction in dHJ dissolution. Our results thus implicate RPA as a cofactor of the BTR complex in dHJ dissolution. [ABSTRACT FROM AUTHOR]

Nucleolytic processing of DNA double-strand breaks (DSBs) generates 3' ssDNA tails that are essential for the assembly of DNA damage checkpoint signaling and DNA repair protein complexes. Genetic studies have provided evidence that multiple nuclease activities are involved in DSB end resection. Three recent studies, including work by Jackson and colleagues (pp. 2767-2772) in the October 15, 2008, issue of Genes & Development, have begun to shed some light on the intricacy of this process. [ABSTRACT FROM AUTHOR]

Members of the RecQ helicase family play critical roles in genome maintenance. There are five RecQ homologs in mammals, and defects in three of these (BLM, WRN, and RECQL4) give rise to cancer predisposition syndromes in humans. RECQL and RECQL5 have not been associated with a human disease. Here we show that deletion of Recql5 in mice results in cancer susceptibility. Recql5-deficient cells exhibit elevated frequencies of spontaneous DNA double-strand breaks and homologous recombination (HR) as scored using a reporter that harbors a direct repeat, and are prone to gross chromosomal rearrangements in response to replication stress. To understand how RECQL5 regulates HR, we use purified proteins to demonstrate that human RECQL5 binds the Rad51 recombinase and inhibits Rad51-mediated D-loop formation. By biochemical means and electron microscopy, we show that RECQL5 displaces Rad51 from single-stranded DNA (ssDNA) in a reaction that requires ATP hydrolysis and RPA. Together, our results identify RECQL5 as an important tumor suppressor that may act by preventing inappropriate HR events via Rad51 presynaptic filament disruption. [ABSTRACT FROM AUTHOR]

DNA end resection is critical for chromosome break repair by homologous recombination and influences the efficiency of repair by nonhomologous DNA end joining. An elegant study by Sinha and colleagues (pp. 1423-1437) published in the June 15, 2009, issue of Genes & Development identified a novel mycobacterial DNA end resection protein complex, AdnAB, that harbors dual DNA motor and dual nuclease functions. Sinha and colleagues also demonstrated that the DNA end-binding protein complex Ku regulates the activity of AdnAB.

Homologous recombination accomplishes the exchange of genetic information between two similar or identical DNA duplexes. It can occur either by gene conversion, a process of unidirectional genetic exchange, or by reciprocal crossing over. Homologous recombination is well known for its role in generating genetic diversity in meiosis and, in mitosis, as a DNA repair mechanism. In the immune system, the evidence suggests a role for homologous recombination in Ig gene evolution and in the diversification of Ab function. Previously, we reported the occurrence of homologous recombination between repeated, donor and recipient alleles of the Ig H chain mu gene C (Cmu) region residing at the Ig mu locus in mouse hybridoma cells. In this study, we constructed mouse hybridoma cell lines bearing Cmu region heteroalleles to learn more about the intrachromosomal homologous recombination process. A high frequency of homologous recombination (gene conversion) was observed for markers spanning the entire recipient Cmu region, suggesting that recombination might initiate at random sites within the Cmu region. The Cmu region heteroalleles were equally proficient as either conversion donors or recipients. Remarkably, when the same Cmu heteroalleles were tested for recombination in ectopic genomic positions, the mean frequency of gene conversion was reduced by at least 65-fold. These results are consistent with the murine IgH mu locus behaving as a hot spot for intrachromosomal homologous recombination.