Across a species range, islands of stressful habitats impose similar selection pressures on isolated populations. It is as yet unclear, when populations respond to these selective pressures, the extent to which this results in convergent genetic evolution and whether convergence is due to independent mutations or shared ancestral variation. We address these questions investigating a classic example of adaptation by natural selection – the colonization of plant species to heavy metal contaminated soils. We use field-based reciprocal transplant experiments to demonstrate that mine alleles at a major copper tolerance QTL, Tol1, are strongly selected in the mine environment, but are neutral, or nearly so, in the off-mine environment. To identify scaffolds in genetic linkage with this locus, we assemble the genome of a mine adapted M. guttatus genotype and sequence near isogenic lines (NILs) homozygous for tolerant or non-tolerant alleles at Tol1. We identify genes with differential expression between NILs and differences in allele frequency between independent pairs of mine and off-mine populations to identify Tol1 candidate genes. We identify a single gene, a multicopper oxidase, with large differences in expression between NILs and allele frequency between populations. Furthermore, we find patterns of genetic variation at Tol1, and four additional candidate adaptation loci, are consistent with selection acting upon beneficial haplotypes that predates the existence of the copper mine habitat. We estimate the age of selected Tol1 haplotype to be at least 1700 years old and was at a frequency of 0.4-0.6% in the ancestral population when mining was initiated 150 years ago. These results suggest that adaptation to the mine habitat routinely occurs via selection on ancestral variation, rather than independent de-novo mutations or migration between populations.

Thousands of candidate human-specific regulatory sequences (HSRS) have been identified, supporting the idea that unique to human phenotypes result from human-specific alterations of genomic regulatory networks. Here, conservation patterns analysis of 18,364 regulatory DNA segments comprising candidate HSRS was carried out using the most recent releases of the reference genomes’ databases of humans and nonhuman primates (NHP) and defining the sequence conservation threshold as the minimum ratio of bases that must remap of 1.00. Present analyses identified 5,535 candidate HSRS defined by either the acceleration of mutation rates on the human lineage or the functional divergence from chimpanzee that are highly conserved in NHP and appear to evolve by the exaptation of ancestral DNA pathway. This pathway seems mechanistically distinct from the evolution of regulatory DNA driven by the species-specific expansion of transposable elements. It is proposed that phenotypic divergence of Homo sapiens is driven by the evolution of human-specific genomic regulatory networks via at least two mechanistically distinct pathways of creation of divergent sequences of regulatory DNA: i) exaptation of the highly conserved ancestral regulatory DNA segments; ii) human-specific insertions of transposable elements.

The extent to which selection and demography impact patterns of genetic diversity in natural populations of Drosophila melanogaster is yet to be fully understood. We previously observed that the pattern of LD at scales of ~10 kb in the Drosophila Genetic Reference Panel (DGRP), consisting of 145 inbred strains from Raleigh, North Carolina, measured both between pairs of sites and as haplotype homozygosity, is elevated above neutral demographic expectations. Further, we demonstrated that signatures of strong and recent soft sweeps are abundant. However, the extent to which this pattern is specific to this derived and admixed population is unknown. Neither is it clear whether such a pattern may have arisen as a consequence of the extensive inbreeding performed to generate the DGRP data. Here we analyze > 100 fully sequenced strains from Zambia, an ancestral population to the Raleigh population, that has experienced little to no admixture and was generated by sequencing haploid embryos rather than inbred strains. This data set allows us to determine whether patterns of elevated LD and signatures of abundant soft sweeps are generic to multiple populations of D. melanogaster or whether they are generated either by inbreeding, bottlenecks or admixture in the DGRP dataset. We find an elevation in long-range LD and haplotype homozygosity in the Zambian dataset, confirming the result from the DGRP data set. This elevation in LD and haplotype structure remains even after controlling for many sources of LD in the data including genomic inversions, admixture, population substructure, close relatedness of individual strains, and recombination rate variation. Furthermore, signatures of partial soft sweeps similar to those found in the DGRP are common in Zambia. These results suggest that while the selective forces and sources of adaptive mutations may differ in Zambia and Raleigh, elevated long-range LD and signatures of soft sweeps are generic in D. melanogaster.

Ancient origins, profound ecological divergence and extensive hybridization make the fire-bellied toads Bombina bombina and B. variegata (Anura: Bombinatoridae) an intriguing test case of ecological speciation. Narrow Bombina hybrid zones erect barriers to neutral introgression whose strength has been estimated previously. We test this prediction by inferring the rate of gene exchange between pure populations on either side of the intensively studied Krakow transect. We developed a software pipeline to extract high confidence sets of orthologous genes from de novo transcriptome assemblies, fitted a range of divergence models to these data and assessed their relative support with analytic likelihoods calculations. There was clear evidence for post-divergence gene flow, but, as expected, no perceptible signal of recent introgression via the nearby hybrid zone. The analysis of two additional Bombina taxa (B. v. scabra and B. orientalis) validated our parameter estimates against a larger set of prior expectations. Despite substantial cumulative introgression over millions of years, adaptive divergence of the hybridizing taxa is essentially unaffected by their lack of reproductive isolation. Extended distribution ranges also buffer them against small-scale environmental perturbations that have been shown to reverse the speciation process in other, more recent ecotypes.

The mitochondrial (mt) genome is present in several copies in human cells, and intra-individual variation in mtDNA sequences is known as heteroplasmy. A recent study found that heteroplasmies were highly tissue-specific, site-specific, and allele-specific, suggesting that positive selection is acting on such heteroplasmies; however the functional implications have not been explored. This study investigates variation in mtDNA copy numbers (mtCN) in 12 different tissues obtained at autopsy from 152 individuals (ranging in age from 3 days to 96 years). Three different methods to estimate mtCN were compared: shotgun sequencing, capture-enriched sequencing and droplet digital PCR (ddPCR). The highest precision in mtCN estimation was achieved using shotgun sequencing data. However, capture-enrichment data provide reliable estimates of relative (albeit not absolute) mtCNs. Comparisons of mtCN from different tissues of the same individual revealed that mtCNs in different tissues are, with few exceptions, uncorrelated. Hence, each tissue of an individual seems to regulate mtCN in a tissue-related rather than an individual-dependent manner. Skeletal muscle (SM) samples showed an age-related decrease in mtCN that was especially pronounced in males, while there was an age-related increase in mtCN for liver (LIV) samples. MtCN in SM samples was significantly negatively correlated with both the total number of heteroplasmic sites and with minor allele frequency (MAF) at two heteroplasmic sites, 408 and 16327. Heteroplasmies at both sites are highly specific for SM, occur in more than 40 % of the individuals older than 50 years (with MAF up to 28.2 %), and are part of functional elements that regulate mtDNA replication. We hypothesize that positive selection acting on these heteroplasmic sites is reducing mtCN in SM of older individuals.

The Chlamydomonas genome has been sequenced, assembled and annotated to produce a rich resource for genetics and molecular biology in this well-studied model organism. However, the current reference genome contains ~1000 blocks of unknown sequence (‘N-islands’), which are frequently placed in introns of annotated gene models. We developed a strategy, using careful bioinformatics analysis of short-sequence cDNA and genomic DNA reads, to search for previously unknown exons hidden within such blocks, and determine the sequence and exon/intron boundaries of such exons. These methods are based on assembly and alignment completely independent of prior reference assembly or reference annotation. Our evidence indicates that ~one-quarter of the annotated intronic N-islands actually contain hidden exons. For most of these our algorithm recovers full exonic sequence with associated splice junctions and exon-adjacent intron sequence, that can be joined to the reference genome assembly and annotated transcript models. These new exons represent de novo sequence generally present nowhere in the assembled genome, and the added sequence can be shown in many cases to greatly improve evolutionary conservation of the predicted encoded peptides. At the same time, our results confirm the purely intronic status for a substantial majority of N-islands annotated as intronic in the reference annotated genome, increasing confidence in this valuable resource.