Disruption of muREC2/RAD51L1 in mice results in early embryonic lethality which can Be partially rescued in a p53(-/-) background.

In the past few years, an increasing number of genes in the recA/RAD51 recombination-repair family have been cloned, including REC2/RAD51L1, RAD51C, RAD51D, R51H3, XRCC2, and XRCC3 (1, 3, 4, 24, 25, 31, 32). Among them, REC2/RAD51L1, which was first cloned in our lab and subsequently at two other labs, encodes a 350-amino-acid protein exhibiting significant homology to the Escherichia coli RECA and Ustilago maydis REC2 and RAD51 genes (25, 26). The regions of similarity include the nucleotide-binding A and B motifs and a DNA binding domain. Overexpression of hREC2/RAD51L1 in Chinese hamster ovary (CHO) cells causes a G1 delay in the cell cycle and hypersensitivity to UV irradiation (10). Overexpression of hREC2/RAD51L1 in T cells of transgenic mice results in partial blockage of T-cell differentiation and hypersensitivity of T cells to ionizing radiation (27a). Although the human Rec2/Rad51L1 protein has not been shown to catalyze recombinase reactions such as DNA pairing and strand transfer, amino acid alignment classifies the gene as a RAD51 ortholog. Some similarities in function have been noted, however. The REC2/RAD51L1 gene is induced by both ionizing radiation (25) and UV radiation (22). The hsRad51 protein appears to be recruited to the nucleus in response to DNA damage (8) and has been shown to interact with p53 directly, suggesting a role in cell cycle regulation and perhaps apoptosis (2, 28). Finally, disruption of both the muRAD51 gene and the muREC2/RAD51L1 gene results in early embryonic lethality (16, 34) (see below). The role of muRec2/Rad51L1 in DNA repair may not involve direct interaction with the damaged site. Thus far, we have been unable to demonstrate a significant level of DNA recognition by this protein (24a), and no ATP-hydrolytic activity has been observed (6a). Furthermore, recent results indicate that this gene may form fusion products upon translocation with an HMGC1 gene in leiomyoma. Potentiating the response to DNA damage may involve an indirect association with the proteins that regulate the cell cycle. Thus, in terms of DNA repair, the properties of this homolog of Rad51 are likely to be very different from those of the prototype protein. To investigate the biological function of muREC2/RAD51L1 and create a mouse model to study DNA repair mechanisms, we disrupted muREC2/RAD51L1 in embryonic stem (ES) cells via homologous recombination. These experiments resulted in the creation of mice bearing a single allelic copy of the mREC2/RAD51L1 gene. We were unable to obtain a homozygous knockout mouse because the targeting process resulted in early embryonic lethality. Heterozygous mice appear normal after 12 months, while homozygous mutants die during early embryonic development. Analysis of mutant embryos in vivo and in vitro indicated that REC2/RAD51L1 is essential for cell proliferation. Although breeding of REC2/RAD51L1-heterozygous mice with p53-knockout mice failed to generate double-knockout pups, the double-mutant embryos survived longer, indicating a partial rescue by p53. In this report, we outline the details of our efforts.