(GT) n Repetitive tracts affect several stages of RecA-promoted recombination

Abstract

The effect of GT/CA dinucleotide repeat tracts on RecA-dependent homologous recombination was examined in vitro. By measuring the binding of RecA protein to oligonucleotides containing GT or CA repeats using the surface plasmon resonance (BIAcore), we show that the efficiency of RecA protein binding is sequence dependent: the protein binds to non-repetitive, poly(CA) or poly(GT) sequences with an increasing affinity. This preferential binding to repetitive sequences is specific for RecA protein and is not observed with the single-strand DNA binding (SSB) protein. Despite the fact that RecA filaments formed on GT tracts efficiently bind duplex DNAs, they are unable to promote stable joint formation. Moreover, strand exchange between a duplex DNA and a fully homologous single-stranded DNA (ssDNA) containing dinucleotide repeats, is inhibited as a function of the length of the repetitive tract. The number of molecules which underwent a complete strand exchange decreased from 100% to 80% and 30% for DNA containing seven, 16 and 39 GT repeats, respectively. The inhibition is less pronounced when the ssDNA contains CA instead of GT dinucleotide repeats. We propose that the high affinity of RecA protein for (CA) n or (GT) n tracts prevents strand exchange from progressing across such sequences. Thus, our results suggest that repetitive tracts of dinucleotides CA/GT could influence recombinational activity potentially leading to an increase in genomic rearrangements.