Transcription factor binding sites are frequently under accelerated evolution in primates.

Recent comparative genomic studies have identified many human accelerated elements (HARs) with elevated substitution rates in the human lineage. However, it remains unknown to what extent transcription factor binding sites (TFBSs) are under accelerated evolution in humans and other primates. Here, we introduce two pooling-based phylogenetic methods with dramatically enhanced sensitivity to examine accelerated evolution in TFBSs. Using these new methods, we show that more than 6000 TFBSs annotated in the human genome have experienced accelerated evolution in Hominini, apes, and Old World monkeys. Although these TFBSs individually show relatively weak signals of accelerated evolution, they collectively are more abundant than HARs. Also, we show that accelerated evolution in Pol III binding sites may be driven by lineage-specific positive selection, whereas accelerated evolution in other TFBSs might be driven by nonadaptive evolutionary forces. Finally, the accelerated TFBSs are enriched around developmental genes, suggesting that accelerated evolution in TFBSs may drive the divergence of developmental processes between primates.

Chromosome-level genome assembly of a high-altitude-adapted frog (Rana kukunoris) from the Tibetan plateau provides insight into amphibian genome evolution and adaptation.

BACKGROUND: The high-altitude-adapted frog Rana kukunoris, occurring on the Tibetan plateau, is an excellent model to study life history evolution and adaptation to harsh high-altitude environments. However, genomic resources for this species are still underdeveloped constraining attempts to investigate the underpinnings of adaptation. RESULTS: The R. kukunoris genome was assembled to a size of 4.83 Gb and the contig N50 was 1.80 Mb. The 6555 contigs were clustered and ordered into 12 pseudo-chromosomes covering ~ 93.07% of the assembled genome. In total, 32,304 genes were functionally annotated. Synteny analysis between the genomes of R. kukunoris and a low latitude species Rana temporaria showed a high degree of chromosome level synteny with one fusion event between chr11 and chr13 forming pseudo-chromosome 11 in R. kukunoris. Characterization of features of the R. kukunoris genome identified that 61.5% consisted of transposable elements and expansions of gene families related to cell nucleus structure and taste sense were identified. Ninety-five single-copy orthologous genes were identified as being under positive selection and had functions associated with the positive regulation of proteins in the catabolic process and negative regulation of developmental growth. These gene family expansions and positively selected genes indicate regions for further interrogation to understand adaptation to high altitude. CONCLUSIONS: Here, we reported a high-quality chromosome-level genome assembly of a high-altitude amphibian species using a combination of Illumina, PacBio and Hi-C sequencing technologies. This genome assembly provides a valuable resource for subsequent research on R. kukunoris genomics and amphibian genome evolution in general.

Allopatric origin of sympatric whitefish morphs with insights on the genetic basis of their reproductive isolation.

The European whitefish (Coregonus lavaretus) species complex is a classic example of recent adaptive radiation. Here, we examine a whitefish population introduced to northern Finnish Lake Tsahkal in the late 1960s, where three divergent morphs (viz. littoral, pelagic, and profundal feeders) were found 10 generations after. Using demographic modeling based on genomic data, we show that whitefish morphs evolved during a phase of strict isolation, refuting a rapid sympatric divergence scenario. The lake is now an artificial hybrid zone between morphs originated in allopatry. Despite their current syntopy, clear genetic differentiation remains between two of the three morphs. Using admixture mapping, we identify five SNPs associated with gonad weight variation, a proxy for sexual maturity and spawning time. We suggest that ecological adaptations in spawning time evolved in allopatry are currently maintaining partial reproductive isolation in the absence of other barriers to gene flow.

Population Structure Limits Parallel Evolution in Sticklebacks.

Population genetic theory predicts that small effective population sizes (Ne) and restricted gene flow limit the potential for local adaptation. In particular, the probability of evolving similar phenotypes based on shared genetic mechanisms (i.e., parallel evolution), is expected to be reduced. We tested these predictions in a comparative genomic study of two ecologically similar and geographically codistributed stickleback species (viz. Gasterosteus aculeatus and Pungitius pungitius). We found that P. pungitius harbors less genetic diversity and exhibits higher levels of genetic differentiation and isolation-by-distance than G. aculeatus. Conversely, G. aculeatus exhibits a stronger degree of genetic parallelism across freshwater populations than P. pungitius: 2,996 versus 379 single nucleotide polymorphisms located within 26 versus 9 genomic regions show evidence of selection in multiple freshwater populations of G. aculeatus and P. pungitius, respectively. Most regions involved in parallel evolution in G. aculeatus showed increased levels of divergence, suggestive of selection on ancient haplotypes. In contrast, haplotypes involved in freshwater adaptation in P. pungitius were younger. In accordance with theory, the results suggest that connectivity and genetic drift play crucial roles in determining the levels and geographic distribution of standing genetic variation, providing evidence that population subdivision limits local adaptation and therefore also the likelihood of parallel evolution.

Genetic population structure constrains local adaptation in sticklebacks.

Repeated and independent adaptation to specific environmental conditions from standing genetic variation is common. However, if genetic variation is limited, the evolution of similar locally adapted traits may be restricted to genetically different and potentially less optimal solutions or prevented from happening altogether. Using a quantitative trait locus (QTL) mapping approach, we identified the genomic regions responsible for the repeated pelvic reduction (PR) in three crosses between nine-spined stickleback populations expressing full and reduced pelvic structures. In one cross, PR mapped to linkage group 7 (LG7) containing the gene Pitx1, known to control pelvic reduction also in the three-spined stickleback. In the two other crosses, PR was polygenic and attributed to 10 novel QTL, of which 90% were unique to specific crosses. When screening the genomes from 27 different populations for deletions in the Pitx1 regulatory element, these were only found in the population in which PR mapped to LG7, even though the morphological data indicated large-effect QTL for PR in several other populations as well. Consistent with the available theory and simulations parameterized on empirical data, we hypothesize that the observed variability in genetic architecture of PR is due to heterogeneity in the spatial distribution of standing genetic variation caused by >2× stronger population structuring among freshwater populations and >10× stronger genetic isolation by distance in the sea in nine-spined sticklebacks as compared to three-spined sticklebacks.

On the causes of geographically heterogeneous parallel evolution in sticklebacks.

The three-spined stickleback (Gasterosteus aculeatus) is an important model system for the study of parallel evolution in the wild, having repeatedly colonized and adapted to freshwater from the sea throughout the northern hemisphere. Previous studies identified numerous genomic regions showing consistent genetic differentiation between freshwater and marine ecotypes but these had typically limited geographic sampling and mostly focused on the Eastern Pacific region. We analysed population genomic data from global samples of the three-spined stickleback marine and freshwater ecotypes to detect loci involved in parallel evolution at different geographic scales. Most signatures of parallel evolution were unique to the Eastern Pacific and trans-oceanic marine-freshwater differentiation was restricted to a limited number of shared genomic regions, including three chromosomal inversions. On the basis of simulations and empirical data, we demonstrate that this could result from the stochastic loss of freshwater-adapted alleles during the invasion of the Atlantic basin and selection against freshwater-adapted variants in the sea, both of which can reduce standing genetic variation available for freshwater adaptation outside the Eastern Pacific region. Moreover, the elevated linkage disequilibrium associated with marine-freshwater differentiation in the Eastern Pacific is consistent with secondary contact between marine and freshwater populations that evolved in isolation from each other during past glacial periods. Thus, contrary to what earlier studies from the Eastern Pacific region have led us to believe, parallel marine-freshwater differentiation in sticklebacks is far less prevalent and pronounced in all other parts of the species global distribution range.

Estimating uncertainty in divergence times among three-spined stickleback clades using the multispecies coalescent.

Incomplete lineage sorting (ILS) can lead to biased divergence time estimates. To explore if and how ILS has influenced the results of a recent worldwide phylogeny of three-spined sticklebacks (Gasterosteus aculeatus), we estimated divergence times among major clades by applying both a concatenation approach and the multispecies coalescent (MSC) model to single-nucleotide polymorphisms. To further test the influence of different calibration strategies, we applied different calibrations to the root and to younger nodes in addition to the ones used in the previous study. Both the updated calibrations and the application of the MSC model influenced divergence time estimates, sometimes significantly. The new divergence time estimates were more ancient than in the previous study for older nodes, whereas the estimates of younger nodes were not strongly affected by the re-analyses. However, since the applied MSC method employs a simple substitution model and cannot account for changes in population size, we suggest that different analytical approaches and calibration strategies should be used in order to explore uncertainty in divergence time estimates. This study provides a valuable reference timeline for the ages of worldwide three-spined stickleback populations and emphasizes the need to embrace, rather than obscure, uncertainties around divergence time estimates.

A phylogenomic perspective on diversity, hybridization and evolutionary affinities in the stickleback genus Pungitius.

Hybridization and convergent evolution are phenomena of broad interest in evolutionary biology, but their occurrence poses challenges for reconstructing evolutionary affinities among affected taxa. Sticklebacks in the genus Pungitius are a case in point: evolutionary relationships and taxonomic validity of different species and populations in this circumpolarly distributed species complex remain contentious due to convergent evolution of traits regarded as diagnostic in their taxonomy, and possibly also due to frequent hybridization among taxa. To clarify the evolutionary relationships among different Pungitius species and populations globally, as well as to study the prevalence and extent of introgression among recognized species, genomic data sets of both reference genome-anchored single nucleotide polymorphisms and de novo assembled RAD-tag loci were constructed with RAD-seq data. Both data sets yielded topologically identical and well-supported species trees. Incongruence between nuclear and mitochondrial DNA-based trees was found and suggested possibly frequent hybridization and mitogenome capture during the evolution of Pungitius sticklebacks. Further analyses revealed evidence for frequent nuclear genetic introgression among Pungitius species, although the estimated proportions of autosomal introgression were low. Apart from providing evidence for frequent hybridization, the results challenge earlier mitochondrial and morphology-based hypotheses regarding the number of species and their affinities in this genus: at least seven extant species can be recognized on the basis of genetic data. The results also shed new light on the biogeographical history of the Pungitius-complex, including suggestion of several trans-Arctic invasions of Europe from the Northern Pacific. The well-resolved phylogeny should facilitate the utility of this genus as a model system for future comparative evolutionary studies.

Worldwide phylogeny of three-spined sticklebacks.

Stickleback fishes in the family Gasterosteidae have become model organisms in ecology and evolutionary biology. However, even in the case of the most widely studied species in this family - the three-spined stickleback (Gasterosteus aculeatus) - the worldwide phylogenetic relationships and colonization history of the different populations and lineages remain poorly resolved. Using a large collection of samples covering most parts of the species distribution range, we subjected thousands of single nucleotide polymorphisms to coalescent analyses in order to reconstruct a robust worldwide phylogeny of extant G. aculeatus populations, as well as their ancestral geographic distributions using Statistical-Dispersal Vicariance and Bayesian Binary MCMC analyses. The results suggest that contemporary populations originated from the Pacific Ocean in the Late Pleistocene, and the Atlantic was colonized through the Arctic Ocean by a lineage that diverged from Pacific sticklebacks ca 44.6 Kya. This lineage contains two branches: one that is distributed in the Mediterranean area, from the Iberian Peninsula to the Black Sea ('Southern European Clade'), and another that is comprised of populations from northern Europe and the east coast of North America ('Trans-Atlantic Clade'). Hence, the results suggest that the North American East Coast was colonized by trans-Atlantic migration. Coalescence-based divergence time estimates suggest that divergence among major clades is much more recent than previously estimated.