7. The Ruv system resolves Holliday junctions

14.7 The Ruv system resolves Holliday junctions

Figure 14.12 RuvAB is an asymmetric complex that promotes branch migration of a Holliday junction.

A group of three genes in E. coli codes for functions involved later in recombination. The products of ruvA and ruvB increase the formation of heteroduplex structures. RuvA recognizes the structure of the Holliday junction. RuvA binds to all four strands of DNA at the crossover point and forms two tetramers that sandwich the DNA. RuvB is an ATPase that functions as a hexamer; it has helicase activity that provides the motor for branch migration. Hexameric rings of RuvB bind around each duplex of DNA upstream of the crossover point. A diagram of the function of the complex is shown in Figure 14.12.

The RuvAB complex can cause the branch to migrate as fast as 10 V20 bp/sec. A similar activity is provided by the RecG helicase. RuvAB displaces RecA from DNA during its action. The RuvAB and RecG activities both can act on Holliday junctions, but if both are mutant, E. coli is completely defective in recombination activity.

The third gene, ruvC, codes for an endonuclease that specifically recognizes Holliday junctions. It can cleave such junctions in vitro to resolve recombination intermediates. A common tetranucleotide sequence provides a hotspot for RuvC to resolve the Holliday junction. The tetranucleotide (ATTG) is asymmetric, and thus may direct resolution with regard to which pair of strands is nicked. This determines whether the outcome is patch recombinant formation (no overall recombination) or splice recombinant formation (recombination between flanking markers).

Figure 14.13 Bacterial enzymes can catalyze all stages of recombination in the repair pathway following the production of suitable substrate DNA molecules.

We may now account for the stages of recombination in E. coli in terms of individual proteins. Figure 14.13 shows the events that are involved in using recombination to repair a gap in one duplex by retrieving material from the other duplex. The major caveat in applying these conclusions to recombination in eukaryotes is that bacterial recombination generally involves interaction between a fragment of DNA and a whole chromosome. It occurs as a repair reaction that is stimulated by damage to DNA, and this is not entirely equivalent to recombination between genomes at meiosis. Nonetheless, similar molecular activities are involved in manipulating DNA.

Homologs of RecA are ubiquitous among prokaryotes, and related proteins have been found in eukaryotes. Two genes in S. cerevisiae, DMC1 and rad51, code for proteins that are related to RecA. Mutations in these genes cause a similar phenotype; they accumulate double Vstrand breaks and fail to form normal synaptonemal complexes. This reinforces the idea that exchange of strands between DNA duplexes is involved in formation of the synaptonemal complex, and raises the possibility that chromosome synapsis is related to the bacterial strand assimilation reaction. However, eukaryotic homologs of RecA do not form filaments, so the mechanics of the reaction are likely to be different in eukaryotes.