Human recombination rates are increased around accelerated conserved regions--evidence for continued selection?

MOTIVATION: We hypothesized that recombination rates might be increased at genetic loci that are subject to more intense selection. Here, we test this hypothesis by using a recently published set of accelerated conserved regions and fine-scale recombination rate estimates provided by the HapMap project. RESULTS: We observed that fine-scale recombination rates are increased around conserved noncoding regions that show accelerated evolution in human or chimp, as compared to noncoding regions showing accelerated evolution in mouse and those being conserved between human and fugu. Recombination rates around hominid accelerated conserved regions (ACRs) are furthermore increased as compared to exonic regions. On the other hand, GC-content is reduced around ACRs, excluding a major confounding influence of GC-content on the observed variation in recombination rate. Conclusion: Our observations indicate that selection intensity could be an important determinant of local recombination rate variation and that continued positive selection might act at many ACR loci. Alternatively, a confounding factor needs to be found that causes a congruent signal in recombination rate estimates based on human polymorphism data and in the comparative genomic data. Researchers who consider the explanation involving selection as more likely may expect more common functional sequence variants at ACRs in genetic association studies.     

The influence of recombination on human genetic diversity

In humans, the rate of recombination, as measured on the megabase scale, is positively associated with the level of genetic variation, as measured at the genic scale. Despite considerable debate, it is not clear whether these factors are causally linked or, if they are, whether this is driven by the repeated action of adaptive evolution or molecular processes such as double-strand break formation and mismatch repair. We introduce three innovations to the analysis of recombination and diversity: fine-scale genetic maps estimated from genotype experiments that identify recombination hotspots at the kilobase scale, analysis of an entire human chromosome, and the use of wavelet techniques to identify correlations acting at different scales. We show that recombination influences genetic diversity only at the level of recombination hotspots. Hotspots are also associated with local increases in GC content and the relative frequency of GC-increasing mutations but have no effect on substitution rates. Broad-scale association between recombination and diversity is explained through covariance of both factors with base composition. To our knowledge, these results are the first evidence of a direct and local influence of recombination hotspots on genetic variation and the fate of individual mutations. However, that hotspots have no influence on substitution rates suggests that they are too ephemeral on an evolutionary time scale to have a strong influence on broader scale patterns of base composition and long-term molecular evolution.     

Male-driven evolution

The strength of male-driven evolution - that is, the magnitude of the sex ratio of mutation rate - has been a controversial issue, particularly in primates. While earlier studies estimated the male-to-female ratio (alpha) of mutation rate to be about 4-6 in higher primates, two recent studies claimed that alpha is only about 2 in humans. However, a more recent comparison of mutation rates between a noncoding fragment on Y and a homologous region on chromosome 3 gave an estimate of alpha = 5.3, reinstating strong male-driven evolution in hominoids. Several studies investigated variation in mutation rates among genomic regions that may not be related to sex differences and found strong evidence for such variation. The causes for regional variation in mutation rate are not clear but GC content and recombination are two possible causes. Thus, while the strong male-driven evolution in higher primates suggests that errors during DNA replication in the germ cells are the major source of mutation, the contribution of some replication-independent factors such as recombination may also be important.     

Significant positive correlation between the recombination rate and GC content in the human pseudoautosomal region.

This paper establishes that recombination drives the evolution of GC content in a significant way. Because the human P-arm pseudoautosomal region (PAR1) has been shown to have a high recombination rate, at least 20-fold more frequent than the genomic average of approximately 1 cM/Mb, this region provides an ideal system to study the role of recombination in the evolution of base composition. Nine non-coding regions of PAR1 are analyzed in this study. We have observed a highly significant positive correlation between the recombination rate and GC content (rho = 0.837, p < or = 0.005). Five regions that lie in the distal part of PAR1 are shown to be significantly higher than genomic average divergence. By comparing the intra- and inter-specific AT->GC -GC->AT ratios, we have detected no fixation bias toward GC alleles except for L254915, which has excessive AT-->GC changes in the human lineage. Thus, we conclude that the high GC content of the PAR1 genes better fits the biased gene conversion (BGC) model.     

The impact of recombination on nucleotide substitutions in the human genome

Unraveling the evolutionary forces responsible for variations of neutral substitution patterns among taxa or along genomes is a major issue for detecting selection within sequences. Mammalian genomes show large-scale regional variations of GC-content (the isochores), but the substitution processes at the origin of this structure are poorly understood. We analyzed the pattern of neutral substitutions in 1 Gb of primate non-coding regions. We show that the GC-content toward which sequences are evolving is strongly negatively correlated to the distance to telomeres and positively correlated to the rate of crossovers (R²=47%). This demonstrates that recombination has a major impact on substitution patterns in human, driving the evolution of GC-content. The evolution of GC-content correlates much more strongly with male than with female crossover rate, which rules out selectionist models for the evolution of isochores. This effect of recombination is most probably a consequence of the neutral process of biased gene conversion (BGC) occurring within recombination hotspots. We show that the predictions of this model fit very well with the observed substitution patterns in the human genome. This model notably explains the positive correlation between substitution rate and recombination rate. Theoretical calculations indicate that variations in population size or density in recombination hotspots can have a very strong impact on the evolution of base composition. Furthermore, recombination hotspots can create strong substitution hotspots. This molecular drive affects both coding and non-coding regions. We therefore conclude that along with mutation, selection and drift, BGC is one of the major factors driving genome evolution. Our results also shed light on variations in the rate of crossover relative to non-crossover events, along chromosomes and according to sex, and also on the conservation of hotspot density between human and chimp.     

Compositional evolution of noncoding DNA in the human and chimpanzee genomes

We have examined the compositional evolution of noncoding DNA in the primate genome by comparison of lineage-specific substitutions observed in 1.8 Mb of genomic alignments of human, chimpanzee, and baboon with 6542 human single-nucleotide polymorphisms (SNPs) rooted using chimpanzee sequence. The pattern of compositional evolution, measured in terms of the numbers of GC-->AT and AT-->GC changes, differs significantly between fixed and polymorphic sites, and indicates that there is a bias toward fixation of AT-->GC mutations, which could result from weak directional selection or biased gene conversion in favor of high GC content. Comparison of the frequency distributions of a subset of the SNPs revealed no significant difference between GC-->AT and AT-->GC polymorphisms, although AT-->GC polymorphisms in regions of high GC segregate at slightly higher frequencies on average than GC-->AT polymorphisms, which is consistent with a fixation bias favoring high GC in these regions. However, the substitution data suggest that this fixation bias is relatively weak, because the compositional structure of the human and chimpanzee genomes is becoming homogenized, with regions of high GC decreasing in GC content and regions of low GC increasing in GC content. The rate and pattern of nucleotide substitution in 333 Alu repeats within the human-chimpanzee-baboon alignments are not significantly affected by the GC content of the region in which they are inserted, providing further evidence that, since the time of the human-chimpanzee ancestor, there has been little or no regional variation in mutation bias.     

Evolutionary rate of immunoglobulin alpha noncoding region is greater in hominoids than in Old World monkeys.

Recent studies on the molecular evolution of primates show that the evolutionary rate among hominoids is considerably slower than that among nonhominoid primates. However, this observation at the nucleotide-sequence level is restricted to the beta-globin family region. In this study, we sequenced orthologous immunoglobulin alpha (C alpha) genes of chimpanzee, gorilla, orangutan, and crab-eating macaque (an Old World monkey) and compared them with that of the human by using noncoding regions for analysis. Since significant differences in rates among hominoids were not found by using the relative rate test, we evaluated the ratio (R) of the evolutionary distance between Old World monkey and human to the distance between orangutan and human. The R value (1.12) for the C alpha gene was much smaller than the expected value (1.38-2.33), showing that the nucleotide substitution rate (= mutation rate per year under selective neutrality) of the C alpha gene is greater in the human lineage than in the Old World monkey lineage. We also did a similar analysis for the gamma 1-, gamma 2-, psi eta-, and delta-globin genes and found a considerable heterogeneity (1.12-2.37) among the R values, including that for the C alpha gene. This indicates that the hominoid slowdown of the evolutionary rate is not a universal phenomenon in primate evolution.     

Bayesian Inference of Shared Recombination Hotspots Between Humans and Chimpanzees

Recombination generates variation and facilitates evolution. Recombination (or lack thereof) also contributes to human genetic disease. Methods for mapping genes influencing complex genetic diseases via association rely on linkage disequilibrium (LD) in human populations, which is influenced by rates of recombination across the genome. Comparative population genomic analyses of recombination using related primate species can identify factors influencing rates of recombination in humans. Such studies can indicate how variable hotspots for recombination may be both among individuals (or populations) and over evolutionary timescales. Previous studies have suggested that locations of recombination hotspots are not conserved between humans and chimpanzees. We made use of the data sets from recent resequencing projects and applied a Bayesian method for identifying hotspots and estimating recombination rates. We also reanalyzed SNP data sets for regions with known hotspots in humans using samples from the human and chimpanzee. The Bayes factors (BF) of shared recombination hotspots between human and chimpanzee across regions were obtained. Based on the analysis of the aligned regions of human chromosome 21, locations where the two species show evidence of shared recombination hotspots (with high BFs) were identified. Interestingly, previous comparative studies of human and chimpanzee that focused on the known human recombination hotspots within the β-globin and HLA regions did not find overlapping of hotspots. Our results show high BFs of shared hotspots at locations within both regions, and the estimated locations of shared hotspots overlap with the locations of human recombination hotspots obtained from sperm-typing studies.     

Mating system and recombination affect molecular evolution in four Triticeae species.

Mating systems and recombination are thought to have a deep impact on the organization and evolution of genomes. Because of the decline in effective population size and the interference between linked loci, the efficacy of selection is expected to be reduced in regions with low recombination rates and in the whole genome of self-fertilizing species. At the molecular level, relaxed selection is expected to result in changes in the rate of protein evolution and the pattern of codon bias. It is increasingly recognized that recombination also affects non-selective processes such as the biased gene conversion towards GC alleles (bGC). Like selection, this kind of meiotic drive in favour of GC over AT alleles is expected to be reduced in weakly recombining regions and genomes. Here, we investigated the effect of mating system and recombination on molecular evolution in four Triticeae species: two outcrossers (Secale cereale and Aegilops speltoides) and two selfers (Triticum urartu and Triticum monococcum). We found that GC content, possibly driven by bGC, is affected by mating system and recombination as theoretically predicted. Selection efficacy, however, is only weakly affected by mating system and recombination. We investigated the possible reasons for this discrepancy. A surprising one is that, in outcrossing lineages, selection efficacy could be reduced because of high substitution rates in favour of GC alleles. Outcrossers, but not selfers, would thus suffer from a 'GC-induced' genetic load. This result sheds new light on the evolution of mating systems.     

The Red Queen Model of Recombination Hotspots Evolution in the Light of Archaic and Modern Human Genomes

Recombination is an essential process in eukaryotes, which increases diversity by disrupting genetic linkage between loci and ensures the proper segregation of chromosomes during meiosis. In the human genome, recombination events are clustered in hotspots, whose location is determined by the PRDM9 protein. There is evidence that the location of hotspots evolves rapidly, as a consequence of changes in PRDM9 DNA-binding domain. However, the reasons for these changes and the rate at which they occur are not known. In this study, we investigated the evolution of human hotspot loci and of PRDM9 target motifs, both in modern and archaic human lineages (Denisovan) to quantify the dynamic of hotspot turnover during the recent period of human evolution. We show that present-day human hotspots are young: they have been active only during the last 10% of the time since the divergence from chimpanzee, starting to be operating shortly before the split between Denisovans and modern humans. Surprisingly, however, our analyses indicate that Denisovan recombination hotspots did not overlap with modern human ones, despite sharing similar PRDM9 target motifs. We further show that high-affinity PRDM9 target motifs are subject to a strong self-destructive drive, known as biased gene conversion (BGC), which should lead to the loss of the majority of them in the next 3 MYR. This depletion of PRDM9 genomic targets is expected to decrease fitness, and thereby to favor new PRDM9 alleles binding different motifs. Our refined estimates of the age and life expectancy of human hotspots provide empirical evidence in support of the Red Queen hypothesis of recombination hotspots evolution.     

Concerted evolution led to high expression of a prosimian primate delta globin gene locus

The delta globin gene in simian primates is either weakly expressed (in hominoids and New World monkeys) or silent (in Old World monkeys). In prosimian primates, however, an unequal homologous crossover between the psi eta and delta loci of lemurs produced a hybrid psi eta delta pseudogene locus, whereas in tarsier the delta locus encodes a beta-type chain found in 18% of adult tarsier hemoglobin molecules. In the present study, the nucleotide and amino acid sequences of the galago delta and beta globin genes and their encoded peptides were determined, and evidence is provided showing that the galago delta locus encodes a beta-type chain (beta 2) found in 40% of the galago fetal and postnatal hemoglobin molecules, whereas the beta locus encodes the remaining 60% of the beta-type chain (beta 1). Galago beta 1 and beta 2 chains differ from each other by only one amino acid residue. The homology between the galago delta and beta loci extends from 800 base pairs 5' of the proximal CCAAT element to near the end of exon 3 as a result of a recombination event in which beta sequence replaced delta sequence. After this initial recombination event, concerted evolution between the loci continued over their conserved coding, intron 1, and promoter regions but failed to occur between the two loci in their intron 2 and distal 5'-flanking sequences where the two loci have now diverged by 20%. Calculations based on this divergence value and on a rate of noncoding sequence evolution of 4.2 x 10(-9) to 5.5 x 10(-9) substitutions/site/year for the lorisiform lineage to galago yielded a date of 18-24 million years ago for the initial recombination event. The fact that the promoter sequences of the galago delta locus are the same as that of the galago beta locus may account for the high level of expression of the galago delta gene.     

Epigenetic functions enriched in transcription factors binding to mouse recombination hotspots

The regulatory mechanism of recombination is a fundamental problem in genomics, with wide applications in genome-wide association studies, birth-defect diseases, molecular evolution, cancer research, etc. In mammalian genomes, recombination events cluster into short genomic regions called "recombination hotspots". Recently, a 13-mer motif enriched in hotspots is identified as a candidate cis-regulatory element of human recombination hotspots; moreover, a zinc finger protein, PRDM9, binds to this motif and is associated with variation of recombination phenotype in human and mouse genomes, thus is a trans-acting regulator of recombination hotspots. However, this pair of cis and trans-regulators covers only a fraction of hotspots, thus other regulators of recombination hotspots remain to be discovered. In this paper, we propose an approach to predicting additional trans-regulators from DNA-binding proteins by comparing their enrichment of binding sites in hotspots. Applying this approach on newly mapped mouse hotspots genome-wide, we confirmed that PRDM9 is a major trans-regulator of hotspots. In addition, a list of top candidate trans-regulators of mouse hotspots is reported. Using GO analysis we observed that the top genes are enriched with function of histone modification, highlighting the epigenetic regulatory mechanisms of recombination hotspots.     

A Model-Based Analysis of GC-Biased Gene Conversion in the Human and Chimpanzee Genomes

GC-biased gene conversion (gBGC) is a recombination-associated process that favors the fixation of G/C alleles over A/T alleles. In mammals, gBGC is hypothesized to contribute to variation in GC content, rapidly evolving sequences, and the fixation of deleterious mutations, but its prevalence and general functional consequences remain poorly understood. gBGC is difficult to incorporate into models of molecular evolution and so far has primarily been studied using summary statistics from genomic comparisons. Here, we introduce a new probabilistic model that captures the joint effects of natural selection and gBGC on nucleotide substitution patterns, while allowing for correlations along the genome in these effects. We implemented our model in a computer program, called phastBias, that can accurately detect gBGC tracts about 1 kilobase or longer in simulated sequence alignments. When applied to real primate genome sequences, phastBias predicts gBGC tracts that cover roughly 0.3% of the human and chimpanzee genomes and account for 1.2% of human-chimpanzee nucleotide differences. These tracts fall in clusters, particularly in subtelomeric regions; they are enriched for recombination hotspots and fast-evolving sequences; and they display an ongoing fixation preference for G and C alleles. They are also significantly enriched for disease-associated polymorphisms, suggesting that they contribute to the fixation of deleterious alleles. The gBGC tracts provide a unique window into historical recombination processes along the human and chimpanzee lineages. They supply additional evidence of long-term conservation of megabase-scale recombination rates accompanied by rapid turnover of hotspots. Together, these findings shed new light on the evolutionary, functional, and disease implications of gBGC. The phastBias program and our predicted tracts are freely available.     

Live hot, die young: transmission distortion in recombination hotspots

There is strong evidence that hotspots of meiotic recombination in humans are transient features of the genome. For example, hotspot locations are not shared between human and chimpanzee. Biased gene conversion in favor of alleles that locally disrupt hotspots is a possible explanation of the short lifespan of hotspots. We investigate the implications of such a bias on human hotspots and their evolution. Our results demonstrate that gene conversion bias is a sufficiently strong force to produce the observed lack of sharing of intense hotspots between species, although sharing may be much more common for weaker hotspots. We investigate models of how hotspots arise, and find that only models in which hotspot alleles do not initially experience drive are consistent with observations of rather hot hotspots in the human genome. Mutations acting against drive cannot successfully introduce such hotspots into the population, even if there is direct selection for higher recombination rates, such as to ensure correct segregation during meiosis. We explore the impact of hotspot alleles on patterns of haplotype variation, and show that such alleles mask their presence in population genetic data, making them difficult to detect.     

Correlation of Genetic and Physical Maps at the a Mating-Type Locus of Coprinus Cinereus

The A mating type locus of Coprinus cinereus is remarkable for its extreme diversity, with over 100 different alleles in natural populations. Classical genetic studies have demonstrated that this hypervariability arises in part from recombination between two subloci of A, alpha and beta, although more recent population genetic data have indicated a third segregating sublocus. In this study, we characterized the molecular basis by which recombination generates nonparental A mating types. We mapped the frequency and location of all recombination events in two crosses and correlated the genetic and physical maps of A. We found that all recombination events were located in 6 kb of noncoding DNA between the alpha and beta subloci and that the rate of recombination in this noncoding region matched that generally observed for this genome. No recombination within gene clusters or within coding regions was observed, and the two alpha and beta subloci described in genetic analyses correlated with the previously characterized alpha and beta gene clusters. We propose that pairs of genes constitute both the sex determining and the hereditary unit of A.     

GC-biased gene conversion and selection affect GC content in the Oryza genus (rice)

Base composition varies among and within eukaryote genomes. Although mutational bias and selection have initially been invoked, more recently GC-biased gene conversion (gBGC) has been proposed to play a central role in shaping nucleotide landscapes, especially in yeast, mammals, and birds. gBGC is a kind of meiotic drive in favor of G and C alleles, associated with recombination. Previous studies have also suggested that gBGC could be at work in grass genomes. However, these studies were carried on third codon positions that can undergo selection on codon usage. As most preferred codons end in G or C in grasses, gBGC and selection can be confounded. Here we investigated further the forces that might drive GC content evolution in the rice genus using both coding and noncoding sequences. We found that recombination rates correlate positively with equilibrium GC content and that selfing species (Oryza sativa and O. glaberrima) have significantly lower equilibrium GC content compared with more outcrossing species. As recombination is less efficient in selfing species, these results suggest that recombination drives GC content. We also detected a positive relationship between expression levels and GC content in third codon positions, suggesting that selection favors codons ending with G or C bases. However, the correlation between GC content and recombination cannot be explained by selection on codon usage alone as it was also observed in noncoding positions. Finally, analyses of polymorphism data ruled out the hypothesis that genomic variation in GC content is due to mutational processes. Our results suggest that both gBGC and selection on codon usage affect GC content in the Oryza genus and likely in other grass species.     

Hypervariable noncoding sequences in Saccharomyces cerevisiae

Compared to protein-coding sequences, the evolution of noncoding sequences and the selective constraints placed on these sequences is not well characterized. To compare the evolution of coding and noncoding sequences, we have conducted a survey for DNA polymorphism at five randomly chosen loci among a diverse collection of 81 strains of Saccharomyces cerevisiae. Average rates of both polymorphism and divergence are 40% lower at noncoding sites and 90% lower at nonsynonymous sites in comparison to synonymous sites. Although noncoding and coding sequences show substantial variability in ratios of polymorphism to divergence, two of the loci, MLS1 and PDR10, show a higher rate of polymorphism at noncoding compared to synonymous sites. The high rate of polymorphism is not accompanied by a high rate of divergence and is limited to a few small regions. These hypervariable regions include sites with three segregating bases at a single site and adjacent polymorphic sites. We show that this clustering of polymorphic sites is significantly greater than one would expect on the basis of the spacing between polymorphic fourfold degenerate sites. Although hypervariable noncoding sequences could result from selection on regulatory mutations, they could also result from transient mutational hotspots.     

The analysis of nucleotide substitutions, gaps, and recombination events between RHD and RHCE genes through complete sequencing

We determined the entire nucleotide sequences of all introns within the RHD and RHCE genes by amplifying genomic DNA using long PCR methods. The RHD and RHCE genes were 57,295 and 57,831 bp in length, respectively. Aligning both genes revealed 138 gaps (insertions and deletions) below 100 bp, 1116 substitutions in all introns and all exons (coding region), and 5 gaps of over 100 bp. Homologies (%) between the RH genes were 93.8% over all introns and coding exons and 91.7% over all exons and introns. Various short tandem repeats (STRs) and many interspersed nuclear elements were identified in both genes. The proportions of Alu sequences in the RHD and RHCE genes were 25.9 and 25.7%, respectively and these Alu sequences were concentrated in several regions. We confirmed multiple recombinations in introns 1 and 2. Such multiple recombination, which probably arose due to the concentrations of Alu sequences and the high level of the homology (%), is one of most important factors in the formation and evolution of RH gene. The variability of the Rh system may be generated because of these features of RH genes. Apparent mutational hotspots and regions with low of K values (the numbers of substitutions per nucleotide site) caused by recombinations as well as true mutational hotspots may be found in human genome. Accordingly, in searching for and identifying single nucleotide polymorphisms (SNPs) especially in noncoding regions, apparent mutational hotspots and areas of low K values by recombination should be noted since the unequal distribution of SNPs will reduce the power of SNPs as genetic maker. Combining the complete sequences' data of both RH genes with serological findings will provide beneficial information with which to elucidate the mechanism of recombination, mutation, polymorphism, and evolution of other genes containing the RH gene as well as to analyze Rh variants and develop new methods of Rh genotyping.     

Can a genome change its (hot)spots?

A new study by Jeffreys et al. shows that the rate of recombination in recombination hotspots in humans is not constant through time. This observation adds weight to the idea that hotspots are transient on evolutionary timescales. However, questions remain as to what controls their evolution and how these rapid changes influence broad-scale rates of recombination.