Control of Meiotic Recombination at a Human Crossover Hotspot
thesisposted on 29.08.2013, 10:40 by Mahmut Cerkez Ergoren
Meiotic recombination plays a key role in reshuffling haplotypes in human populations, thus profoundly affecting evolution. However, our understanding of recombination dynamics is largely limited to descriptions of variation in populations and families. Higher resolution analysis (0.0001 cM or less) of de novo recombination events in human sperm DNA has revealed clustering into very narrow hotspots (1-2 kb) that generally coincide with abrupt breakdown of linkage disequilibrium (LD). Myers et al. (2008) used population genetics approaches to survey HapMap Phase II genotype data across the whole genome to infer sites of historical recombination. Around 40% of ~30,000 hotspots so identified contain a 13-bp motif CCNCCNTNNCCNC which appears to be involved in hotspot specification and which may also drive modes of genome instability such as disease-causing genomic rearrangements, minisatellite mutation and common mitochondrial deletions. In addition, recent studies identified that the trans-regulator PRDM9 of meiotic recombination hotspots in humans and mice, contains a zinc finger array that can recognize the 13-bp sequence motif associated with hotspots, with binding to this motif possibly triggering hotspot activity via chromatin re-modelling in humans. Berg et al., (2010) reported that PRDM9 is a major global regulator of hotspots in humans, even hotspots lacking the sequence motif, and influences aspects of genome instability. To examine the effect of the 13-bp motif on crossover frequencies and distribution, four candidate putative hotspots located on different chromosomes and identified from HapMap data were studied. These hotspots had the motif at their centre, and a SNP that disrupts the motif. LDU analysis confirmed the locations of the putative hotspots to a 1-2-kb interval. However, only the first candidate LD hotspot, ‘Hotspot DA’ could be analysed in detailed high-resolution analyses for understanding the influences of cis-regulation on hotspot activity. The other candidate LD hotspots were eliminated because of their disrupting SNP locations on the hotspot or because of a lack of suitable donors in our sperm donor panel. Comparing the rates and distributions of sperm crossover events between donors heterozygous for the disrupting SNP showed that there is a huge asymmetry between the two alleles with the derived and motif-disrupting allele suppressing hotspot activity. This direct study to understand the effect of the disrupting allele on crossover initiation revealed that Hotspot DA is the first to show very strong direct cis-regulation for hotspot activity, and despite being influenced by the trans-factor PRDM9, it is not the major regulator for this hotspot.