The synaptic activity of HsDmc1, a human recombination protein specific to meiosis

Abstract

Human Dmc1 protein, a meiosis-specific homolog of Escherichia coli RecA protein, has previously been shown to promote DNA homologous pairing and strand-exchange reactions that are qualitatively similar to those of RecA protein and Rad51. Human and yeast Rad51 proteins each form a nucleoprotein filament that is very similar to the filament formed by RecA protein. However, recent studies failed to find a similar filament made by Dmc1 but showed instead that this protein forms octameric rings and stacks of rings. These observations stimulated further efforts to elucidate the mechanism by which Dmc1 promotes the recognition of homology. Dmc1, purified to a state in which nuclease and helicase activities were undetectable, promoted homologous pairing and strand exchange as measured by fluorescence resonance energy transfer (FRET). Observations on the intermediates and products, which can be distinguished by FRET assays, provided direct evidence of a three-stranded synaptic intermediate. The effects of helix stability and mismatched base pairs on the recognition of homology revealed further that human Dmc1, like human Rad51, requires the preferential breathing of A⅐T base pairs for recognition of homology. We conclude that Dmc1, like human Rad51 and E. coli RecA protein, promotes homologous pairing and strand exchange by a ''synaptic pathway'' involving a three-stranded nucleoprotein intermediate, rather than by a ''helicase pathway'' involving the separation and reannealing of DNA strands.T wo kinds of macromolecular protein structures play prominent roles in homologous genetic recombination: toroids and filaments. Among the toroidal structures, the best understanding of the relation of function to structure exists for Escherichia coli RuvB protein, which forms a hexameric ATPase͞helicase. Two such hexameric rings, interacting with RuvA protein, drive the migration of Holliday structures, the 4-fold branched DNA intermediate formed after initial synapsis and processing have occurred. The consequence of this driven migration is the reciprocal exchange of a pair of like strands between two duplex molecules (1). Rings are also formed by the exonuclease of phage (2), the Erf protein of phage P22 (3), and human Rad52 protein (4, 5). On the basis of the crystal structure of toroidal exonuclease, Kovall and Matthews suggested a mechanistic explanation of the high processivity of this enzyme (2).Some recombination proteins form both rings and filaments: the ␤ protein of phage forms large rings and left-handed helical filaments (6). ␤ protein promotes annealing of complementary single strands (7,8) and migration of a single-stranded branch (9). This protein, which does not hydrolyze ATP, binds more strongly to the product of its annealing activity, which may help to explain how it drives branch migration (10), but there are no correlations of structure with function that help us to understand the role of either the rings or the left-handed filaments. RecT protein, the product of a cryptic prophage in E. coli and a functional ...

DNA replication meets genetic exchange: Chromosomal damage and its repair by homologous recombination

“…Rate constants of the reaction between DNAs with mismatches are consistent with this idea, because it is reported that mismatches (i) reduce the rate of the pairing step; (ii) increase the off rate during the pairing step; and (iii) reduce the rate of subsequent strand exchange (57). Recent studies employing oligonucleotides with varying GC-content as recombination substrates revealed that both the E. coli RecA and its human homologs Rad51 and Dmc1 rely on A:T base pairs for recognition of homology (58)(59)(60).…”

Clarifying the mechanics of DNA strand exchange in meiotic recombination

“…4a). The lack of helical filaments was surprising because Dmc1 has high sequence conservation with RecA-related proteins and because biophysical studies revealed that Dmc1 and Rad51 recognise homology in the same way 32 . It was suggested that rings were an artefact caused by lack of essential cofactors 32 , but models for ring-based DNA strand exchange were also proposed 33 .…”

Double-stranded DNA breaks and gene functions in recombination and meiosis

“…Phylogenetic analysis indicates that DMC1 is a RecA-like protein [60]. DMC1 homologs have been identified in a wide range of organisms, including budding and fission yeast, mammals, plants, and fungi [61][62][63][64][65][66][67][68][69][70]. In the budding yeast, the severity of dmc1 mutants varies among different strains.…”