Daniel R. Schrider

Comparative and demographic analysis of orang-utan genomes

‘Orang-utan’ is derived from a Malay term meaning ‘man of the forest’ and aptly describes the southeast Asian great apes native to Sumatra and Borneo. The orang-utan species, Pongo abelii (Sumatran) and Pongo pygmaeus (Bornean), are the most phylogenetically distant great apes from humans, thereby providing an informative perspective on hominid evolution. Here we present a Sumatran orang-utan draft genome assembly and short read sequence data from five Sumatran and five Bornean orang-utan genomes. Our analyses reveal that, compared to other primates, the orang-utan genome has many unique features. Structural evolution of the orang-utan genome has proceeded much more slowly than other great apes, evidenced by fewer rearrangements, less segmental duplication, a lower rate of gene family turnover and surprisingly quiescent Alu repeats, which have played a major role in restructuring other primate genomes. We also describe a primate polymorphic neocentromere, found in both Pongo species, emphasizing the gradual evolution of orang-utan genome structure. Orang-utans have extremely low energy usage for a eutherian mammal, far lower than their hominid relatives. Adding their genome to the repertoire of sequenced primates illuminates new signals of positive selection in several pathways including glycolipid metabolism. From the population perspective, both Pongo species are deeply diverse; however, Sumatran individuals possess greater diversity than their Bornean counterparts, and more species-specific variation. Our estimate of Bornean/Sumatran speciation time, 400,000 years ago, is more recent than most previous studies and underscores the complexity of the orang-utan speciation process. Despite a smaller modern census population size, the Sumatran effective population size (Ne) expanded exponentially relative to the ancestral Ne after the split, while Bornean Ne declined over the same period. Overall, the resources and analyses presented here offer new opportunities in evolutionary genomics, insights into hominid biology, and an extensive database of variation for conservation efforts.

Genomic Variation in Natural Populations of Drosophila melanogaster

This report of independent genome sequences of two natural populations of Drosophila melanogaster (37 from North America and 6 from Africa) provides unique insight into forces shaping genomic polymorphism and divergence. Evidence of interactions between natural selection and genetic linkage is abundant not only in centromere- and telomere-proximal regions, but also throughout the euchromatic arms. Linkage disequilibrium, which decays within 1 kbp, exhibits a strong bias toward coupling of the more frequent alleles and provides a high-resolution map of recombination rate. The juxtaposition of population genetics statistics in small genomic windows with gene structures and chromatin states yields a rich, high-resolution annotation, including the following: (1) 5′- and 3′-UTRs are enriched for regions of reduced polymorphism relative to lineage-specific divergence; (2) exons overlap with windows of excess relative polymorphism; (3) epigenetic marks associated with active transcription initiation sites overlap with regions of reduced relative polymorphism and relatively reduced estimates of the rate of recombination; (4) the rate of adaptive nonsynonymous fixation increases with the rate of crossing over per base pair; and (5) both duplications and deletions are enriched near origins of replication and their density correlates negatively with the rate of crossing over. Available demographic models of X and autosome descent cannot account for the increased divergence on the X and loss of diversity associated with the out-of-Africa migration. Comparison of the variation among these genomes to variation among genomes from D. simulans suggests that many targets of directional selection are shared between these species.

Gene copy-number polymorphism in nature

Differences between individuals in the copy-number of whole genes have been found in every multicellular species examined thus far. Such differences result in unique complements of protein-coding genes in all individuals, and have been shown to underlie adaptive phenotypic differences. Here, we review the evidence for copy-number variants (CNVs), focusing on the methods used to detect them and the molecular mechanisms responsible for generating this type of variation. Although there are multiple technical and computational challenges inherent to these experimental methods, next-generation sequencing technologies are making such experiments accessible in any system with a sequenced genome. We further discuss the connection between copy-number variation within species and copy-number divergence between species, showing that these values are exactly what one would expect from similar comparisons of nucleotide polymorphism and divergence. We conclude by reviewing the growing body of evidence for natural selection on copy-number variants. While it appears that most genic CNVs—especially deletions—are quickly eliminated by selection, there are now multiple studies demonstrating a strong link between copy-number differences at specific genes and phenotypic differences in adaptive traits. We argue that a complete understanding of the molecular basis for adaptive natural selection necessarily includes the study of copy-number variation.

Rates and Genomic Consequences of Spontaneous Mutational Events in Drosophila melanogaster

Because spontaneous mutation is the source of all genetic diversity, measuring mutation rates can reveal how natural selection drives patterns of variation within and between species. We sequenced eight genomes produced by a mutation-accumulation experiment in Drosophila melanogaster. Our analysis reveals that point mutation and small indel rates vary significantly between the two different genetic backgrounds examined. We also find evidence that ∼2% of mutational events affect multiple closely spaced nucleotides. Unlike previous similar experiments, we were able to estimate genome-wide rates of large deletions and tandem duplications. These results suggest that, at least in inbred lines like those examined here, mutational pressures may result in net growth rather than contraction of the Drosophila genome. By comparing our mutation rate estimates to polymorphism data, we are able to estimate the fraction of new mutations that are eliminated by purifying selection. These results suggest that ∼99% of duplications and deletions are deleterious—making them 10 times more likely to be removed by selection than nonsynonymous mutations. Our results illuminate not only the rates of new small- and large-scale mutations, but also the selective forces that they encounter once they arise.

Extensive error in the number of genes inferred from draft genome assemblies

Current sequencing methods produce large amounts of data, but genome assemblies based on these data are often woefully incomplete. These incomplete and error-filled assemblies result in many annotation errors, especially in the number of genes present in a genome. In this paper we investigate the magnitude of the problem, both in terms of total gene number and the number of copies of genes in specific families. To do this, we compare multiple draft assemblies against higher-quality versions of the same genomes, using several new assemblies of the chicken genome based on both traditional and next-generation sequencing technologies, as well as published draft assemblies of chimpanzee. We find that upwards of 40% of all gene families are inferred to have the wrong number of genes in draft assemblies, and that these incorrect assemblies both add and subtract genes. Using simulated genome assemblies of Drosophila melanogaster, we find that the major cause of increased gene numbers in draft genomes is the fragmentation of genes onto multiple individual contigs. Finally, we demonstrate the usefulness of RNA-Seq in improving the gene annotation of draft assemblies, largely by connecting genes that have been fragmented in the assembly process.

Pervasive multinucleotide mutational events in eukaryotes.

Many aspects of mutational processes are nonrandom, from the preponderance of transitions relative to transversions to the higher rate of mutation at CpG dinucleotides [1]. However, it is still often assumed that single-nucleotide mutations are independent of one another, each being caused by separate mutational events. The occurrence of multiple, closely spaced substitutions appears to violate assumptions of independence and is often interpreted as evidence for the action of adaptive natural selection [2, 3], balancing selection [4], or compensatory evolution [5, 6]. Here we provide evidence of a frequent, widespread multinucleotide mutational process active throughout eukaryotes. Genomic data from mutation-accumulation experiments, parent-offspring trios, and human polymorphisms all show that simultaneous nucleotide substitutions occur within short stretches of DNA. Regardless of species, such multinucleotide mutations (MNMs) consistently comprise ~3% of the total number of nucleotide substitutions. These results imply that previous adaptive interpretations of multiple, closely spaced substitutions may have been unwarranted and that MNMs must be considered when interpreting sequence data.

Very Few RNA and DNA Sequence Differences in the Human Transcriptome

RNA editing is an important cellular process by which the nucleotides in a mature RNA transcript are altered to cause them to differ from the corresponding DNA sequence. While this process yields essential transcripts in humans and other organisms, it is believed to occur at a relatively small number of loci. The rarity of RNA editing has been challenged by a recent comparison of human RNA and DNA sequence data from 27 individuals, which revealed that over 10,000 human exonic sites appear to exhibit RNA-DNA differences (RDDs). Many of these differences could not have been caused by either of the two previously known human RNA editing mechanisms—ADAR-mediated A→G substitutions or APOBEC1-mediated C→U switches—suggesting that a previously unknown mechanism of RNA editing may be active in humans. Here, we reanalyze these data and demonstrate that genomic sequences exist in these same individuals or in the human genome that match the majority of RDDs. Our results suggest that the majority of these RDD events were observed due to accurate transcription of sequences paralogous to the apparently edited gene but differing at the edited site. In light of our results it seems prudent to conclude that if indeed an unknown mechanism is causing RDD events in humans, such events occur at a much lower frequency than originally proposed.

Gene Copy-Number Polymorphism Caused by Retrotransposition in Humans

The era of whole-genome sequencing has revealed that gene copy-number changes caused by duplication and deletion events have important evolutionary, functional, and phenotypic consequences. Recent studies have therefore focused on revealing the extent of variation in copy-number within natural populations of humans and other species. These studies have found a large number of copy-number variants (CNVs) in humans, many of which have been shown to have clinical or evolutionary importance. For the most part, these studies have failed to detect an important class of gene copy-number polymorphism: gene duplications caused by retrotransposition, which result in a new intron-less copy of the parental gene being inserted into a random location in the genome. Here we describe a computational approach leveraging next-generation sequence data to detect gene copy-number variants caused by retrotransposition (retroCNVs), and we report the first genome-wide analysis of these variants in humans. We find that retroCNVs account for a substantial fraction of gene copy-number differences between any two individuals. Moreover, we show that these variants may often result in expressed chimeric transcripts, underscoring their potential for the evolution of novel gene functions. By locating the insertion sites of these duplicates, we are able to show that retroCNVs have had an important role in recent human adaptation, and we also uncover evidence that positive selection may currently be driving multiple retroCNVs toward fixation. Together these findings imply that retroCNVs are an especially important class of polymorphism, and that future studies of copy-number variation should search for these variants in order to illuminate their potential evolutionary and functional relevance.

Genome-wide analysis of retrogene polymorphisms in Drosophila melanogaster

Gene duplication via retrotransposition has been shown to be an important mechanism in evolution, affecting gene dosage and allowing for the acquisition of new gene functions. Although fixed retrotransposed genes have been found in a variety of species, very little effort has been made to identify retrogene polymorphisms. Here, we examine 37 Illumina-sequenced North American Drosophila melanogaster inbred lines and present the first ever data set and analysis of polymorphic retrogenes in Drosophila. We show that this type of polymorphism is quite common, with any two gametes in the North American population differing in the presence or absence of six retrogenes, accounting for ~13% of gene copy-number heterozygosity. These retrogenes were identified by a straightforward method that can be applied using any type of DNA sequencing data. We also use a variant of this method to conduct a genome-wide scan for intron presence/absence polymorphisms, and show that any two chromosomes in the population likely differ in the presence of multiple introns. We show that these polymorphisms are all in fact deletions rather than intron gain events present in the reference genome. Finally, by leveraging the known location of the parental genes that give rise to the retrogene polymorphisms, we provide direct evidence that natural selection is responsible for the excess of fixations of retrogenes moving off of the X chromosome in Drosophila. Further efforts to identify retrogene and intron presence/absence polymorphisms will undoubtedly improve our understanding of the evolution of gene copy number and gene structure.

Soft Shoulders Ahead: Spurious Signatures of Soft and Partial Selective Sweeps Result from Linked Hard Sweeps

Characterizing the nature of the adaptive process at the genetic level is a central goal for population genetics. In particular, we know little about the sources of adaptive substitution or about the number of adaptive variants currently segregating in nature. Historically, population geneticists have focused attention on the hard-sweep model of adaptation in which a de novo beneficial mutation arises and rapidly fixes in a population. Recently more attention has been given to soft-sweep models, in which alleles that were previously neutral, or nearly so, drift until such a time as the environment shifts and their selection coefficient changes to become beneficial. It remains an active and difficult problem, however, to tease apart the telltale signatures of hard vs. soft sweeps in genomic polymorphism data. Through extensive simulations of hard- and soft-sweep models, here we show that indeed the two might not be separable through the use of simple summary statistics. In particular, it seems that recombination in regions linked to, but distant from, sites of hard sweeps can create patterns of polymorphism that closely mirror what is expected to be found near soft sweeps. We find that a very similar situation arises when using haplotype-based statistics that are aimed at detecting partial or ongoing selective sweeps, such that it is difficult to distinguish the shoulder of a hard sweep from the center of a partial sweep. While knowing the location of the selected site mitigates this problem slightly, we show that stochasticity in signatures of natural selection will frequently cause the signal to reach its zenith far from this site and that this effect is more severe for soft sweeps; thus inferences of the target as well as the mode of positive selection may be inaccurate. In addition, both the time since a sweep ends and biologically realistic levels of allelic gene conversion lead to errors in the classification and identification of selective sweeps. This general problem of “soft shoulders” underscores the difficulty in differentiating soft and partial sweeps from hard-sweep scenarios in molecular population genomics data. The soft-shoulder effect also implies that the more common hard sweeps have been in recent evolutionary history, the more prevalent spurious signatures of soft or partial sweeps may appear in some genome-wide scans.