Genomic consequences of selection on self-incompatibility genes

Frequency-dependent selection at plant self-incompatibility systems is inherent and well understood theoretically. A self-incompatibility locus leads to a strong peak of diversity in the genome, to a unique distribution of diversity across the species and possibly to increased introgression between closely related species. We review recent empirical studies demonstrating these features and relate the empirical findings to theoretical predictions. We show how these features are being exploited in searches for other genes under multi-allelic balancing selection and for inference on recent breakdown of self-incompatibility.     

Effect of balancing selection on spatial genetic structure within populations: theoretical investigations on the self-incompatibility locus and empirical studies in Arabidopsis halleri

The effect of selection on patterns of genetic structure within and between populations may be studied by contrasting observed patterns at the genes targeted by selection with those of unlinked neutral marker loci. Local directional selection on target genes will produce stronger population genetic structure than at neutral loci, whereas the reverse is expected for balancing selection. However, theoretical predictions on the intensity of this signal under precise models of balancing selection are still lacking. Using negative frequency-dependent selection acting on self-incompatibility systems in plants as a model of balancing selection, we investigated the effect of such selection on patterns of spatial genetic structure within a continuous population. Using numerical simulations, we tested the effect of the type of self-incompatibility system, the number of alleles at the self-incompatibility locus and the dominance interactions among them, the extent of gene dispersal, and the immigration rate on spatial genetic structure at the selected locus and at unlinked neutral loci. We confirm that frequency-dependent selection is expected to reduce the extent of spatial genetic structure as compared to neutral loci, particularly in situations with low number of alleles at the self-incompatibility locus, high frequency of codominant interactions among alleles, restricted gene dispersal and restricted immigration from outside populations. Hence the signature of selection on spatial genetic structure is expected to vary across species and populations, and we show that empirical data from the literature as well as data reported here on three natural populations of the herb Arabidopsis halleri confirm these theoretical results.     

S-allele diversity in Sorbus aucuparia and Crataegus monogyna (Rosaceae: Maloideae)

RT-PCR was used to obtain the first estimates from natural populations of allelic diversity at the RNase-based gametophytic self-incompatibility locus in the Rosaceae. A total of 20 alleles were retrieved from 20 Sorbus aucuparia individuals, whereas 17 alleles were found in 13 Crataegus monogyna samples. Estimates of population-level allele numbers fall within the range observed in the Solanaceae, the only other family with RNase-based incompatibility for which estimates are available. The nucleotide diversity of S-allele sequences was found to be much lower in the two Rosaceae species as compared with the Solanaceae. This was not due to a lower sequence divergence among most closely related alleles. Rather, it is the depth of the entire genealogy that differs markedly in the two families, with Rosaceae S-alleles exhibiting more recent apparent coalescence. We also investigated patterns of selection at the molecular level by comparing nucleotide diversity at synonymous and nonsynonymous sites. Stabilizing selection was inferred for the 5' region of the molecule, while evidence of diversifying selection was present elsewhere.     

Insight into the roles of selection in speciation from genomic patterns of divergence and introgression in secondary contact in venomous rattlesnakes

Investigating secondary contact of historically isolated lineages can provide insight into how selection and drift influence genomic divergence and admixture. Here, we studied the genomic landscape of divergence and introgression following secondary contact between lineages of the Western Diamondback Rattlesnake (Crotalus atrox) to determine whether genomic regions under selection in allopatry also contribute to reproductive isolation during introgression. We used thousands of nuclear loci to study genomic differentiation between two lineages that have experienced recent secondary contact following isolation, and incorporated sampling from a zone of secondary contact to identify loci that are resistant to gene flow in hybrids. Comparisons of patterns of divergence and introgression revealed a positive relationship between allelic differentiation and resistance to introgression across the genome, and greater‐than‐expected overlap between genes linked to lineage‐specific divergence and loci that resist introgression. Genes linked to putatively selected markers were related to prominent aspects of rattlesnake biology that differ between populations of Western Diamondback rattlesnakes (i.e., venom and reproductive phenotypes). We also found evidence for selection against introgression of genes that may contribute to cytonuclear incompatibility, consistent with previously observed biased patterns of nuclear and mitochondrial alleles suggestive of partial reproductive isolation due to cytonuclear incompatibilities. Our results provide a genome‐scale perspective on the relationships between divergence and introgression in secondary contact that is relevant for understanding the roles of selection in maintaining partial isolation of lineages, causing admixing lineages to not completely homogenize.     

Distribution of S-alleles in island populations of flowering cherry, Prunus lannesiana var. speciosa

We surveyed the distribution of S-alleles in natural island populations of Prunus lannesiana var. speciosa sampled from seven sites on the Izu Peninsula and six Izu islands, Japan. The S-genotypes of sampled individuals were determined by Southern analysis of RFLPs generated by restriction enzyme digestion of genomic DNA, using cDNA of the S-RNase gene as a probe. All individuals were heterozygous, as expected under gametophytic self-incompatibility (GSI). Sixty-three S-alleles were observed in the species, but 12 private to the Izu Peninsula population seemed to be derived from related species, giving a total of 75. The estimated number of S-alleles in each population ranged from 26 to 62, and was inversely correlated with the respective population's distance from the Izu Peninsula, the closest point in the mainland to the islands. This geographical cline in the estimated numbers of S-alleles suggests that gene flow to and from the distant island populations was less frequent, and that the studied species has migrated from the mainland to the Izu islands. The genetic relationship at the S-locus among populations also gave an "isolation by distance" pattern. The genetic differentiation at the S-locus among the populations was very low (F(ST) = 0.014, p < 0.001). The number of S-alleles in the species did not seem to depend on genetic differences associated with population subdivisions. This might be due to the greater effective migration rates of S-alleles, as expected under balancing selection in GSI.     

Massive introgression of major histocompatibility complex (MHC) genes in newt hybrid zones

Variation in the vertebrate major histocompatibility complex (MHC) genes is crucial for fighting pathogen assault. Because new alleles confer a selective advantage, MHC should readily introgress between species, even under limited hybridization. Using replicated transects through two hybrid zones between strongly reproductively isolated European newts, Lissotriton montandoni and L. vulgaris, we demonstrated recent and ongoing MHC class I and II introgression in the Carpathian region. The extent of introgression correlated with the age of contact. In the older zone, MHC similarity between species within transects exceeded similarity between transects within species, implying pervasive introgression ‐ a massive exchange of MHC genes, not limited to specific variants. In simulations, the observed pattern emerged under the combined action of balancing selection and hybridization, but not when these processes acted separately. Thus, massive introgression at advanced stages of divergence can introduce novel and restore previously lost MHC variation, boosting the adaptive potential of hybridizing taxa. In consequence, MHC genes may be the last to stop introgressing between incipient species.     

Plant self-incompatibility in natural populations: a critical assessment of recent theoretical and empirical advances

Self-incompatibility systems in plants are genetic systems that prevent self-fertilization in hermaphrodites through recognition and rejection of pollen expressing the same allelic specificity as that expressed in the pistils. The evolutionary properties of these self-recognition systems have been revealed through a fascinating interplay between empirical advances and theoretical developments. In 1939, Wright suggested that the main evolutionary force driving the genetic and molecular properties of these systems was strong negative frequency-dependent selection acting on pollination success. The empirical observation of high allelic diversity at the self-incompatibility locus in several species, followed by the discovery of very high molecular divergence among alleles in all plant families where the locus has been identified, supported Wright's initial theoretical predictions as well as many of its later developments. In the last decade, however, advances in the molecular characterization of the incompatibility reaction and in the analysis of allelic frequencies and allelic divergence from natural populations have stimulated new theoretical investigations that challenged some important assumptions of Wright's model of gametophytic self-incompatibility. We here review some of these recent empirical and theoretical advances that investigated: (i) the hypothesis that S-alleles are selectively equivalent, and the evolutionary consequences of genetic interactions between alleles; (ii) the occurrence of frequency-dependent selection in female fertility; (iii) the evolutionary genetics of self-incompatibility systems in subdivided populations; (iv) the evolutionary implications of the self-incompatibility locus's genetic architecture; and (v) of its interactions with the genomic environment.     

Hitch-hiking to a locus under balancing selection: high sequence diversity and low population subdivision at the S-locus genomic region in Arabidopsis halleri.

Hitch-hiking to a site under balancing selection is expected to produce a local increase in nucleotide polymorphism and a decrease in population differentiation compared with the background genomic level, but empirical evidence supporting these predictions is scarce. We surveyed molecular diversity at four genes flanking the region controlling self-incompatibility (the S-locus) in samples from six populations of the herbaceous plant Arabidopsis halleri, and compared their polymorphism with sequences from five control genes unlinked to the S-locus. As a preliminary verification, the S-locus flanking genes were shown to co-segregate with SRK, the gene involved in the self-incompatibility reaction at the pistil level. In agreement with theory, our results demonstrated a significant peak of nucleotide diversity around the S-locus as well as a significant decrease in population genetic structure in the S-locus region compared with both control genes and a set of seven unlinked microsatellite markers. This is consistent with the theoretical expectation that balancing selection is increasing the effective migration rate in subdivided populations. Although only four S-locus flanking genes were investigated, our results suggest that these two signatures of the hitch-hiking effect are localized in a very narrow genomic region.     

History cleans up messes: The impact of time in driving divergence and introgression in a tropical suture zone

Contact zones provide an excellent arena in which to address questions about how genomic divergence evolves during lineage divergence. They allow us to both infer patterns of genomic divergence in allopatric populations isolated from introgression and to characterize patterns of introgression after lineages meet. Thusly motivated, we analyze genome‐wide introgression data from four contact zones in three genera of lizards endemic to the Australian Wet Tropics. These contact zones all formed between morphologically cryptic lineage‐pairs within morphologically defined species, and the lineage‐pairs meeting in the contact zones diverged anywhere from 3.1 to 5.8 million years ago. By characterizing patterns of molecular divergence across an average of 11K genes and fitting geographic clines to an average of 7.5K variants, we characterize how patterns of genomic differentiation and introgression change through time. Across this range of divergences, we find that genome‐wide differentiation increases but becomes no less heterogeneous. In contrast, we find that introgression heterogeneity decreases dramatically, suggesting that time helps isolated genomes “congeal.” Thus, this work emphasizes the pivotal role that history plays in driving lineage divergence.     

Genealogical Structure among Alleles Regulating Self-Incompatibility in Natural Populations of Flowering Plants

A method is proposed for characterizing the structure of genealogies among alleles that regulate selfincompatibility in flowering plants. Expected distributions of ratios of divergence times among alleles, scaled by functions of allele number, were generated by numerical simulation. These distributions appeared relatively insensitive to the particular parameter values assigned in the simulations over a fourfold range in effective population size and a 100-fold range in mutation rate. Generalized leastsquares estimates of the scaled indices were obtained from genealogies reconstructed from nucleotide sequences of self-incompatibility alleles from natural populations of two solanaceous species. Comparison of the observed indices to the expected distributions generated by numerical simulation indicated that the allelic genealogy of one species appeared consistent with the symmetric balancing selection generated by self-incompatibility. However, the allelic genealogy of the second species showed unusually long terminal branches, suggesting the operation of additional evolutionary processes.     

Comparative analysis of the within-population genetic structure in wild cherry (Prunus avium L.) at the self-incompatibility locus and nuclear microsatellites

Gametophytic self-incompatibility (SI) systems in plants exhibit high polymorphism at the SI controlling S-locus because individuals with rare alleles have a higher probability to successfully pollinate other plants than individuals with more frequent alleles. This process, referred to as frequency-dependent selection, is expected to shape number, frequency distribution, and spatial distribution of self-incompatibility alleles in natural populations. We investigated the genetic diversity and the spatial genetic structure within a Prunus avium population at two contrasting gene loci: nuclear microsatellites and the S-locus. The S-locus revealed a higher diversity (15 alleles) than the eight microsatellites (4-12 alleles). Although the frequency distribution of S-alleles differed significantly from the expected equal distribution, the S-locus showed a higher evenness than the microsatellites (Shannon's evenness index for the S-locus: E = 0.91; for the microsatellites: E = 0.48-0.83). Also, highly significant deviations from neutrality were found for the S-locus whereas only minor deviations were found for two of eight microsatellites. A comparison of the frequency distribution of S-alleles in three age-cohorts revealed no significant differences, suggesting that different levels of selection acting on the S-locus or on S-linked sites might also affect the distribution and dynamics of S-alleles. Autocorrelation analysis revealed a weak but significant spatial genetic structure for the multilocus average of the microsatellites and for the S-locus, but could not ascertain differences in the extent of spatial genetic structure between these locus types. An indirect estimate of gene dispersal, which was obtained to explain this spatial genetic pattern, indicated high levels of gene dispersal within our population (sigma(g) = 106 m). This high gene dispersal, which may be partly due to the self-incompatibility system itself, aids the effective gene flow of the microsatellites, thereby decreasing the contrast between the neutral microsatellites and the S-locus.     

Gametophytic self-incompatibility in Lycium parishii (Solanaceae): allelic diversity, genealogical structure, and patterns of molecular evolution at the S-RNase locus

We characterized allelic diversity at the locus controlling self-incompatibility (SI) for a population of Lycium parishii (Solanaceae) from Organ Pipe National Monument, Arizona. Twenty-four partial sequences of S-RNase alleles were recovered from 25 individuals. Estimates of allelic diversity range from 23 to 27 alleles and, consistent with expectations for SI, individuals are heterozygous. We compare S-RNase diversity, patterns of molecular evolution, and the genealogical structure of alleles from L. parishii to a previously studied population of its congener L. andersonii. Gametophytic SI is well characterized for Solanaceae and although balancing selection is hypothesized to be responsible for high levels of allelic divergence, the pattern of selection varies depending on the portion of the gene considered. Site-specific models investigating patterns of selection for L. parishii and L. andersonii indicate that positive selection occurs in those regions of the S-RNase gene hypothesized as important to the recognition response, whereas positive selection was not detected for any position within regions previously characterized as conserved. A 10-species genealogy including S-RNases from a pair of congeners from each of five genera in Solanaceae reveals extensive transgeneric evolution of L. parishii S-RNases. Further, within Lycium, the Dn/Ds ratios for pairs of closely related alleles for intraspecific versus interspecific comparisons were not significantly different, suggesting that the S-RNase diversity recovered in these two species was present prior to the speciation event separating them. Despite this, two S-RNases from L. parishii are identical to two previously reported alleles for L. andersonii, suggesting gene flow between these species.     

Molecular variation at the self-incompatibility locus in natural populations of the genera Antirrhinum and Misopates

The self-incompatibility system of flowering plants is a classic example of extreme allelic polymorphism maintained by frequency-dependent selection. We used primers designed from three published Antirrhinum hispanicum S-allele sequences in PCR reactions with genomic DNA of plants sampled from natural populations of Antirrhinum and Misopates species. Not surprisingly, given the polymorphism of S-alleles, only a minority of individuals yielded PCR products of the expected size. These yielded 35 genomic sequences, of nine different sequence types of which eight are highly similar to the A. hispanicum S-allele sequences, and one to a very similar unpublished Antirrhinum S-like RNase sequence. The sequence types are well separated from the S-RNase sequences from Solanaceae and Rosaceae, and also from most known "S-like" RNase sequences (which encode proteins not involved in self-incompatibility). An association with incompatibility types has so far been established for only one of the putative S-alleles, but we describe evidence that the other sequences are also S-alleles. Variability in these sequences follows the pattern of conserved and hypervariable regions seen in other S-RNases, but no regions have higher replacement than silent diversity, unlike the results in some other species.     

GENOMIC AND MORPHOLOGICAL ANALYSIS OF A SEMIPERMEABLE AVIAN HYBRID ZONE SUGGESTS ASYMMETRICAL INTROGRESSION OF A SEXUAL SIGNAL

Hybrid zones are geographic regions where differentiated taxa meet and potentially exchange genes. Increasingly, genomic analyses have demonstrated that many hybrid zones are semipermeable boundaries across which introgression is highly variable. In some cases, certain alleles penetrate across the hybrid zone in only one direction, recombining into the alternate genome. We investigated this phenomenon using genomic (genotyping‐by‐sequencing) and morphological (plumage reflectance spectrophotometry) analyses of the hybrid zone between two subspecies of the red‐backed fairy‐wren (Malurus melanocephalus) that differ conspicuously in a sexual signal, male back plumage color. Geographic cline analyses revealed a highly variable pattern of differential introgression, with many narrow coincident clines combined with several significantly wider clines, suggesting that the hybrid zone is a semipermeable tension zone. The plumage cline was shifted significantly into the genomic background of the orange subspecies, consistent with sexual selection driving asymmetrical introgression of red plumage alleles across the hybrid zone. This interpretation is supported by previous experimental work demonstrating an extra‐pair mating advantage for red males, but the role of genetic dominance in driving this pattern remains unclear. This study highlights the potential for sexual selection to erode taxonomic boundaries and promote gene flow, particularly at an intermediate stage of divergence.     

Identification and characterization of a polymorphic receptor kinase gene linked to the self-incompatibility locus of Arabidopsis lyrata

We study the segregation of variants of a putative self-incompatibility gene in Arabidopsis lyrata. This gene encodes a sequence that is homologous to the protein encoded by the SRK gene involved in self-incompatibility in Brassica species. We show by diallel pollinations of plants in several full-sib families that seven different sequences of the gene in A. lyrata are linked to different S-alleles, and segregation analysis in further sibships shows that four other sequences behave as allelic to these. The family data on incompatibility provide evidence for dominance classes among the S-alleles, as expected for a sporophytic SI system. We observe no division into pollen-dominant and pollen-recessive classes of alleles as has been found in Brassica, but our alleles fall into at least three dominance classes in both pollen and stigma expression. The diversity among sequences of the A. lyrata putative S-alleles is greater than among the published Brassica SRK sequences, and, unlike Brassica, the alleles do not cluster into groups with similar dominance.     

High‐density interspecific genetic linkage mapping provides insights into genomic incompatibility between channel catfish and blue catfish

Catfish is the leading aquaculture species in the United States. The interspecific hybrid catfish produced by mating female channel catfish with male blue catfish outperform both of their parent species in a number of traits. However, mass production of the hybrids has been difficult because of reproductive isolation. Investigations of genome structure and organization of the hybrids provide insights into the genetic basis for maintenance of species divergence in the face of gene flow, thereby helping develop strategies for introgression and efficient production of the hybrids for aquaculture. In this study, we constructed a high‐density genetic linkage map using the hybrid catfish system with the catfish 250K SNP array. A total of 26 238 SNPs were mapped to 29 linkage groups, with 12 776 unique marker positions. The linkage map spans approximately 3240 cM with an average intermarker distance of 0.25 cM. A fraction of markers (986 of 12 776) exhibited significant deviation from the expected Mendelian ratio of segregation, and they were clustered in major genomic blocks across 15 LGs, most notably LG9 and LG15. The distorted markers exhibited significant bias for maternal alleles among the backcross progenies, suggesting strong selection against the blue catfish alleles. The clustering of distorted markers within genomic blocks should lend insights into speciation as marked by incompatibilities between the two species. Such findings should also have profound implications for understanding the genomic evolution of closely related species as well as the introgression of hybrid production programs in aquaculture.     

A 15-Myr-old genetic bottleneck

Balancing selection preserves variation at the self-incompatibility locus (S-locus) of flowering plants for tens of millions of years, making it possible to detect demographic events that occurred prior to the origin of extant species. In contrast to other Solanaceae examined, SI species in the sister genera Physalis and Witheringia share restricted variation at the S-locus. This restriction is indicative of an ancient bottleneck that occurred in a common ancestor. We sequenced 14 S-alleles from the subtribe Iochrominae, a group that is sister to the clade containing Physalis and Witheringia. At least 6 ancient S-allele lineages are represented among these alleles, demonstrating that the Iochrominae taxa do not share the restriction in S-locus diversity. Therefore, the bottleneck occurred after the divergence of the Iochrominae from the lineage leading to the most recent common ancestor of Physalis and Witheringia. Using cpDNA sequences, 3 fossil dates, and a Bayesian-relaxed molecular clock approach, the crown group of Solanaceae was estimated to be 51 Myr old and the restriction of variation at the S-locus occurred 14.0-18.4 Myr before present. These results confirm the great age of polymorphism at the S-locus and the utility of loci under balancing selection for deep historical inference.     

Searching the genomes of inbred mouse strains for incompatibilities that reproductively isolate their wild relatives

Identification of the genes that underlie reproductive isolation provides important insights into the process of speciation. According to the Dobzhansky-Muller model, these genes suffer disrupted interactions in hybrids due to independent divergence in separate populations. In hybrid populations, natural selection acts to remove the deleterious heterospecific combinations that cause these functional disruptions. When selection is strong, this process can maintain multilocus associations, primarily between conspecific alleles, providing a signature that can be used to locate incompatibilities. We applied this logic to populations of house mice that were formed by hybridization involving two species that show partial reproductive isolation, Mus domesticus and Mus musculus. Using molecular markers likely to be informative about species ancestry, we scanned the genomes of 1) classical inbred strains and 2) recombinant inbred lines for pairs of loci that showed extreme linkage disequilibria. By using the same set of markers, we identified a list of locus pairs that displayed similar patterns in both scans. These genomic regions may contain genes that contribute to reproductive isolation between M. domesticus and M. musculus. This hypothesis can now be tested using laboratory crosses and surveys of introgression in the wild.     

Molecular cloning of two Solanum chacoense S-alleles and a hypothesis concerning their evolution

The polymerase chain reaction (PCR) has been used to clone two S-alleles (S13 and S14) from Solanum chacoense. The two alleles do not cross-hybridize on genomic Southern blots or on northern blots using stylar RNA. Although the S14 message was not detected in a stylar cDNA library prepared from mature flowers, a full-length copy of the S13 coding sequence was isolated by screening with the PCR fragment. We have analysed the sequences of the S13 cDNA and the S14 PCR fragment (60% of the mature protein coding sequence) in the context of S-RNase evolution, and propose that random point mutations may be sufficient to generate new S-alleles. Based on a phylogenetic tree composed of RNase sequences containing the conserved RNase motifs HGLWP and KHGXC, we suggest that gametophytic self-incompatibility genes are RNase genes that have acquired a new function in the gametophytic self-incompatibility system early in the evolution of flowering plants.     

An Experimental Test of Adaptive Introgression in Locally Adapted Populations of Splash Pool Copepods

As species struggle to keep pace with the rapidly warming climate, adaptive introgression of beneficial alleles from closely related species or populations provides a possible avenue for rapid adaptation. We investigate the potential for adaptive introgression in the copepod, Tigriopus californicus, by hybridizing two populations with divergent heat tolerance limits. We subjected hybrids to strong heat selection for 15 generations followed by whole-genome resequencing. Utilizing a hybridize evolve and resequence (HER) technique, we can identify loci responding to heat selection via a change in allele frequency. We successfully increased the heat tolerance (measured as LT₅₀) in selected lines, which was coupled with higher frequencies of alleles from the southern (heat tolerant) population. These repeatable changes in allele frequencies occurred on all 12 chromosomes across all independent selected lines, providing evidence that heat tolerance is polygenic. These loci contained genes with lower protein-coding sequence divergence than the genome-wide average, indicating that these loci are highly conserved between the two populations. In addition, these loci were enriched in genes that changed expression patterns between selected and control lines in response to a nonlethal heat shock. Therefore, we hypothesize that the mechanism of heat tolerance divergence is explained by differential gene expression of highly conserved genes. The HER approach offers a unique solution to identifying genetic variants contributing to polygenic traits, especially variants that might be missed through other population genomic approaches.