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.[Keywords: Recql5 helicase; DNA repair; homologous recombination; tumor suppressor; Rad51 recombinase] Supplemental material is available at http://www.genesdev.org.

Type II Rothmund-Thomson syndrome (Type II RTS) is a rare autosomal recessive genetic disorder characterized by a congenital skin rash, birth defects of the skeleton, genomic instability and cancer predisposition. It is caused by mutations in the RECQL4 gene and thus represents one of the three cancer-prone genetic diseases that are caused by mutations in a RecQ helicase-encoding gene. Genomic instability has been suspected as a major underlying cause of this disease, and analyses of Type II RTS patient-derived cells demonstrate unusually high frequencies of chromosomal aberrations, suggesting the involvement of chromosomal instability. However, the nature of the instability induced by RECQL4 mutations has not been clearly defined. We created a viable Recql4 mutant mouse model. These mice exhibit a distinctive skin abnormality, birth defects of the skeletal system, genomic instability and increased cancer susceptibility in a sensitized genetic background. Thus, they provide a useful model for studying Type II RTS. In addition, we demonstrate that cells from these mutant mice have high frequencies of premature centromere separation and aneuploidy. Thus, our observations provide evidence for a previously unsuspected role for Recql4 in sister-chromatid cohesion, and suggest that the chromosomal instability may be the underlying cause of cancer predisposition and birth defects in these mutant mice.

In eukaryotes, crossovers in mitotic cells can have deleterious consequences and therefore must be suppressed. Mutations in BLM give rise to Bloom syndrome, a disease that is characterized by an elevated rate of crossovers and increased cancer susceptibility. However, simple eukaryotes such as Saccharomyces cerevisiae have multiple pathways for suppressing crossovers, suggesting that mammals also have multiple pathways for controlling crossovers in their mitotic cells. We show here that in mouse embryonic stem (ES) cells, mutations in either the Bloom syndrome homologue (Blm) or the Recql5 genes result in a significant increase in the frequency of sister chromatid exchange (SCE), whereas deleting both Blm and Recql5 lead to an even higher frequency of SCE. These data indicate that Blm and Recql5 have nonredundant roles in suppressing crossovers in mouse ES cells. Furthermore, we show that mouse embryonic fibroblasts derived from Recql5 knockout mice also exhibit a significantly increased frequency of SCE compared with the corresponding wild-type control. Thus, this study identifies a previously unknown Recql5-dependent, Blm-independent pathway for suppressing crossovers during mitosis in mice.

High-molecular-weight kininogen (HK) plays an important role in the assembly of the plasma kallikrein-kinin system. While the human genome contains a single copy of the kininogen gene, 3 copies exist in the rat (1 encoding K-kininogen and 2 encoding T-kininogen). Here, we confirm that the mouse genome contains 2 homologous kininogen genes, mKng1 and mKng2, and demonstrate that these genes are expressed in a tissue-specific manner. To determine the roles of these genes in murine development and physiology, we disrupted mKng1, which is expressed primarily in the liver. mKng1 Ϫ/Ϫ mice were viable, but lacked plasma HK and low-molecular-weight kininogen (LK), as well as ⌬mHK-D5, a novel kininogen isoform that lacks kininogen domain 5. Moreover, despite normal tail vein bleeding times, mKng1 Ϫ/Ϫ mice displayed a significantly prolonged time to carotid artery occlusion following Rose Bengal administration and laser-induced arterial injury. These results suggest that a single gene, mKng1, is responsible for production of plasma kininogen, and that plasma HK contributes to induced arterial thrombosis in mice. (Blood. 2008;111:1274-1281)