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Ca-microsatellites as hotspots of homologous recombination

Keywords:

N.Y. Karpechenko – Junior Research Scientist, Department of Chemical carcinogenesis Blokhin Cancer Research Center RAMS. E-mail: nojadg@mail.ru
V.K. Gasanova – Ph.D. (Med.), Senior Researcher Scientist, Department of Chemical carcinogenesis Blokhin Cancer Research Center RAMS. E-mail: victoriag@inbox.ru
V.I. Popenko – Dr.Sc. (Biol.), Leading Research Scientist, Laboratory Cell Biology, Engelhardt Institute of Molecular Biology, Russian Academy of Sciences. E-mail: popenko@eimb.ru
M.G. Yakubovskaya – Dr.Sc. (Med.), Head of Department of Chemical carcinogenesis Blokhin Cancer Research Center RAMS. E-mail: mgyakubovskaya@mail.ru


The efficient and accurate repair of DNA double-strand breaks is critical for cell survival. Homologous recombination (HR) provides an «error-free» way for double-strand break repair as no loss of genetic information occurs, when homolog or sister chromatid are used for repair. They are mainly involved in HR and the frequency of non-sister chromatid interaction is several orders lower. However, when recombination occurs between non-sister chromatids, half of these cases lead to the loss of heterozygosity. In carcinogenesis of inherited cancer consequences this process exceed impacts of point mutagenesis and non-homologous end joining. Moreover, HR is the way for manifestation of recessive point mutations accumulated during life-time. It is well known that some microsatellites represent «hot-spots» of recombination. In particular, one of the most abundant microsatellites, CA-repeat, enhances HR in different model systems. These sequences are able to form Z-DNA and they are instable both in vitro and in vivo because of the slipped-strand mis-pairing events during replication and repair. Previously, we showed that homologous DNA fragments spontaneously interact by their end nucleation leading to the formation of Holliday junctions (HJ). We took advantage of this phenomenon to study composition of HJ population considering cross-point location as a function of the actual sequence of the fragments: random sequence or CA/TG repetitive tracts. Our investigation was performed using electron microscopy. We demonstrated that the majority of HJs formed by PCR fragments with (CA/GT)31-repeats had their cross-points at the repetitive region, while the cross points of Holliday junctions with entirely random sequences were mainly localized at the ends of the fragments. When DNA fragments with (СА/TG)31-repeats or random sequences of equivalent size were obtained by restriction from plasmids no influence of repetitive sequence was observed. Co-incubation of restricted fragments harboring (СА/TG)29- or (СА/TG)31-repeat cause branch migration impeding at the repetitive sequence. Our results mean that impeding of branch migration at the region of repetitive sequence location can occurs, if microsatellites involved into interaction differ by the number of repetitive units. Taking into consideration CA-microsatellite polymorphism in human population causing high frequency of their heterozygosity, our study results explain, why these sequences may represent «hot-spots» for HR.
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