Nonsynonymous Rates Research Articles

The evolution and expression of RBCL in holoparasitic sister‐genera Harveya and Hyobanche (Orobanchaceae)

The evolution of holoparasitism decreases the adaptive value of genes maintaining the photosynthetic apparatus. These may become pseudogenes through insertion or deletion events resulting in frameshift mutations, or by the evolution of premature stop codons. The holoparasitic sister genera Harveya and Hyobanche have undergone alternate pathways of evolution and expression at the plastid locus rbcL. An open reading frame in all but a single species of Harveya is maintained by purifying selection and is expressed. However, the function of Rubisco in this putative holoparasite is unknown. Conversely, Hyobanche has undergone rbcL pseudogene formation, and comparison of synonymous and nonsynonymous rates of evolution indicates that selection has not played a role in its evolution. This is complicated by the following findings: multiple pseudogene copies of rbcL exist in tissues of Hyobanche, rbcL transcripts also encode pseudogenes, and the large subunit is present in some tissues of Hyobanche. We hypothesize that the rbcL operon is in a state of degradation as may be expected in a holoparasite and is not endogenously expressed. Rather, the large subunit may be taken up from the host plants, and accumulate in tissues as a result of transpiration.

A highly unexpected strong correlation between fixation probability of nonsynonymous mutations and mutation rate

Under prevailing theories, the nonsynonymous-to-synonymous substitution ratio (i.e. K(a)/K(s)), which measures the fixation probability of nonsynonymous mutations, is correlated with the strength of selection. In this article, we report that K(a)/K(s) is also strongly correlated with the mutation rate as measured by K(s), and that this correlation appears to have a similar magnitude as the correlation between K(a)/K(s) and selective strength. This finding cannot be reconciled with current theories. It suggests that we should re-evaluate the current paradigms of coding-sequence evolution, and that the wide use of K(a)/K(s) as a measure of selective strength needs reassessment.

Adaptive Diversification of Vomeronasal Receptor 1 Genes in Rodents

The vomeronasal receptor 1 (V1R) are believed to be pheromone receptors in rodents. Here we used computational methods to identify 95 and 62 new putative V1R genes from the draft rat and mouse genome sequence, respectively. The rat V1R repertoire consists of 11 subfamilies, 10 of which are shared with the mouse, while rat appears to lack the H and I subfamilies found in mouse and possesses one unique subfamily (M). The estimations of the relative divergence times suggest that many subfamilies originated after the split of rodents and primates. The analysis also reveals that these clusters underwent an expansion very close to the split of mouse and rat. In addition, maximum likelihood analysis showed that the nonsynonymous and synonymous rate ratio for most of these clusters was much higher than one, suggesting the role of positive selection in the diversification of these duplicated V1R genes. Because V1R are thought to mediate the process of signal transduction in response to pheromone detection, we speculate that the V1R genes have evolved under positive Darwinian selection to maintain the ability to discriminate between large and complex pheromonal mixtures.

Elevated evolutionary rates in the laboratory strain ofSaccharomyces cerevisiae

By using the maximum likelihood method, we made a genome-wide comparison of the evolutionary rates in the lineages leading to the laboratory strain (S288c) and a wild strain (YJM789) of Saccharomyces cerevisiae and found that genes in the laboratory strain tend to evolve faster than in the wild strain. The pattern of elevated evolution suggests that relaxation of selection intensity is the dominant underlying reason, which is consistent with recurrent bottlenecks in the S. cerevisiae laboratory strain population. Supporting this conclusion are the following observations: (i) the increases in nonsynonymous evolutionary rate occur for genes in all functional categories; (ii) most of the synonymous evolutionary rate increases in S288c occur in genes with strong codon usage bias; (iii) genes under stronger negative selection have a larger increase in nonsynonymous evolutionary rate; and (iv) more genes with adaptive evolution were detected in the laboratory strain, but they do not account for the majority of the increased evolution. The present discoveries suggest that experimental and possible industrial manipulations of the laboratory strain of yeast could have had a strong effect on the genetic makeup of this model organism. Furthermore, they imply an evolution of laboratory model organisms away from their wild counterparts, questioning the relevancy of the models especially when extensive laboratory cultivation has occurred. In addition, these results shed light on the evolution of livestock and crop species that have been under human domestication for years.

Rate variation in parasitic plants: correlated and uncorrelated patterns among plastid genes of different function

BackgroundThe analysis of synonymous and nonsynonymous rates of DNA change can help in the choice among competing explanations for rate variation, such as differences in constraint, mutation rate, or the strength of genetic drift. Nonphotosynthetic plants of the Orobanchaceae have increased rates of DNA change. In this study 38 taxa of Orobanchaceae and relatives were used and 3 plastid genes were sequenced for each taxon.ResultsPhylogenetic reconstructions of relative rates of sequence evolution for three plastid genes (rbcL, matK and rps2) show significant rate heterogeneity among lineages and among genes. Many of the non-photosynthetic plants have increases in both synonymous and nonsynonymous rates, indicating that both (1) selection is relaxed, and (2) there has been a change in the rate at which mutations are entering the population in these species. However, rate increases are not always immediate upon loss of photosynthesis. Overall there is a poor correlation of synonymous and nonsynonymous rates. There is, however, a strong correlation of synonymous rates across the 3 genes studied and the lineage-speccific pattern for each gene is strikingly similar. This indicates that the causes of synonymous rate variation are affecting the whole plastid genome in a similar way. There is a weaker correlation across genes for nonsynonymous rates. Here the picture is more complex, as could be expected if there are many causes of variation, differing from taxon to taxon and gene to gene.ConclusionsThe distinctive pattern of rate increases in Orobanchaceae has at least two causes. It is clear that there is a relaxation of constraint in many (though not all) non-photosynthetic lineages. However, there is also some force affecting synonymous sites as well. At this point, it is not possible to tell whether it is generation time, speciation rate, mutation rate, DNA repair efficiency or some combination of these factors.

Statistical analysis on adaptive evolution of SQUA genes in angiosperms

SQUAMOSA (SQUA) subfamily includes important perianth identity genes of MADS-box gene family. SQUA genes of Dendrocalamus latiflorus were sequenced, and phylogenetic form on SQUA genes in angiosperms was analyzed. Relative rate and adaptive evolution after SQUA gene duplication in recent common ancestor of monocots and eudicots were analyzed using the methods of relative rate test, statistic on synonymous and non-synonymous coden substitution sites and likelihood rate test. The results show that both of relative rate and synonymous and non-synonymous coden substitution in eudicot clade are significantly higher than those in monocot clade, and the value of d(N)/d(S) uncovered possible positive selective pressure in eudicot clade.

Genomic organization and expression of 23 new genes from MATα locus of Cryptococcus neoformans var. gattii

The pathogenic yeast Cryptococcus neoformans ( Cn) causes cryptococcosis, a life-threatening disease of the brain. Molecular studies of Cn variety gattii have lagged behind other two varieties (var. grubii and var. neoformans) although they have distinct biology and disease patterns. We focused on gene discovery in MATα locus because it predominates in clinical strains. A var. gattii cosmid library was screened with DNA probes from other two varieties. Two positive clones were sequenced to identify ORFs based on similarities to known proteins, and to ESTs using bioinformatics, and manually by a curator. Approximately 76 kb sequenced DNA revealed 23 genes and ORFs. The existence of predicted genes was verified by RT-PCR analyses designed to amplify spliced sequences. The results confirmed that the transcripts were expressed both at 30 and 37 °C. The var. gattii MATα locus genes showed rearrangements in order and orientation vis-à-vis other two varieties. Mating-specific genes showed higher nonsynonymous mutation rates, and gene trees showed var. gattii strains in a distinct clade. The identification of the largest number, thus far, of var. gattii structural genes should set the stage for future molecular pathogenesis studies.

Estimating absolute rates of synonymous and nonsynonymous nucleotide substitution in order to characterize natural selection and date species divergences.

The rate of molecular evolution can vary among lineages. Sources of this variation have differential effects on synonymous and nonsynonymous substitution rates. Changes in effective population size or patterns of natural selection will mainly alter nonsynonymous substitution rates. Changes in generation length or mutation rates are likely to have an impact on both synonymous and nonsynonymous substitution rates. By comparing changes in synonymous and nonsynonymous rates, the relative contributions of the driving forces of evolution can be better characterized. Here, we introduce a procedure for estimating the chronological rates of synonymous and nonsynonymous substitutions on the branches of an evolutionary tree. Because the widely used ratio of nonsynonymous and synonymous rates is not designed to detect simultaneous increases or simultaneous decreases in synonymous and nonsynonymous rates, the estimation of these rates rather than their ratio can improve characterization of the evolutionary process. With our Bayesian approach, we analyze cytochrome oxidase subunit I evolution in primates and infer that nonsynonymous rates have a greater tendency to change over time than do synonymous rates. Our analysis of these data also suggests that rates have been positively correlated.

Mammalian housekeeping genes evolve more slowly than tissue-specific genes.

Do housekeeping genes, which are turned on most of the time in almost every tissue, evolve more slowly than genes that are turned on only at specific developmental times or tissues? Recent large-scale gene expression studies enable us to have a better definition of housekeeping genes and to address the above question in detail. In this study, we examined 1581 human-mouse orthologous gene pairs for their patterns of sequence evolution, contrasting housekeeping genes with tissue-specific genes. Our results show that, in comparison to tissue-specific genes, housekeeping genes on average evolve more slowly and are under stronger selective constraints as reflected by significantly smaller values of Ka/Ks. Besides stronger purifying selection, we explored several other factors that can possibly slow down nonsynonymous rates in housekeeping genes. Although mutational bias might slightly slow the nonsynonymous rates in housekeeping genes, it is unlikely to be the major cause of the rate difference between the two types of genes. The codon usage pattern of housekeeping genes does not seem to differ from that of tissue-specific genes. Moreover, contrary to the old textbook concept, we found that approximately 74% of the housekeeping genes in our study belong to multigene families, not significantly different from that of the tissue-specific genes ( approximately 70%). Therefore, the stronger selective constraints on housekeeping genes are not due to a lower degree of genetic redundancy.

Identification of genes with fast-evolving regions in microbial genomes

Complete sequences of multiple strains of the same microbial species provide an invaluable source for studying the evolutionary dynamics between orthologous genes over a relatively short time scale. Usually the intensity of the selection pressure is inferred from a comparison between the nonsynonymous substitution rate and the synonymous substitution rate. In this paper, we propose an alternative method for detecting genes with one or more fast-evolving regions from pairwise comparisons of orthologous genes. Our method looks for regions with overrepresented nonsynonymous mutations along the alignment, and requires a higher nonsynonymous evolution rate in those regions than the neutral evolution rate. It identifies gene targets under intensive selection pressure that are not detected from the conventional rate comparison analysis. For those identified genes with known annotations, most of them have a clear role in processes such as bacterial defense and host-pathogen interactions. Gene sets reported from our method provide a measure of the phenotypic divergence between two closely related genomes.

Sex-linked mammalian sperm proteins evolve faster than autosomal ones.

X-linked genes can evolve slower or faster depending on whether most recessive, or at least partially recessive alleles are deleterious or beneficial due to their hemizygous expression in males. Molecular studies of X chromosome divergence have provided conflicting evidence for both a higher and lower rate of nucleotide substitution at both synonymous and nonsynonymous sites, depending on the nucleotide sites sampled. Using human and mouse orthologous genes, we tested the hypothesis that genes encoding male-specific sperm proteins are evolving faster on the X chromosome compared with autosomes. X-linked sperm proteins have an average nonsynonymous mutation rate almost twice as high as sperm genes found on autosomes, unlike other tissue-specific genes, where no significant difference in the nonsynonymous mutation rate between the X chromosome and autosomes was found. However, no difference was found in the average synonymous mutation rate of X-linked versus autosomal sperm proteins, which along with corresponding higher values of Ka/Ks in X-linked sperm proteins suggest that differences in selective forces and not mutation rates are the underlying cause of higher X-linked mammalian sperm protein divergence.

Adaptive evolution of cytochrome c oxidase subunit VIII in anthropoid primates.

Cytochrome c oxidase (COX) is a 13-subunit protein complex that catalyzes the last step in mitochondrial electron transfer in mammals. Of the 10 subunits encoded by nuclear DNA (three are mtDNA products), some are expressed as tissue- and/or development-specific isoforms. For COX subunit VIII, previous work showed that expression of the contractile muscle-specific isoform gene, COX8H, is absent in humans and Old World monkeys, and the other isoform gene, COX8L, is expressed ubiquitously. Here, we show that COX8H is transcribed in most primate clades, but its expression is absent in catarrhines, that is, in Old World monkeys and hominids (apes, including humans), having become a pseudogene in the stem of the catarrhines. The ubiquitously expressed isoform, COX8L, underwent nonsynonymous rate acceleration and elevation in the ratio of nonsynonymous/synonymous changes in the stem of anthropoid primates (New World monkeys and catarrhines), possibly setting the stage for loss of the heart-type (H) isoform. The most rapidly evolving region of VIII-L is one that interacts with COX I, suggesting that the changes are functionally coadaptive. Because accelerated rates of nonsynonymous substitutions in anthropoids such as observed for COX8L are also shown by genes for at least 13 other electron transport chain components, these encoded amino acid replacements may be viewed as part of a series of coadaptive changes that optimized the anthropoid biochemical machinery for aerobic energy metabolism. We argue that these changes were linked to the evolution of an expanded neocortex in anthropoid primates.

Adaptive evolution in the photosensory domain of phytochrome A in early angiosperms.

Flowering plant diversity now far exceeds the combined diversity of all other plant groups. Recently identified extant remnants of the earliest-diverging lines suggest that the first angiosperms may have lived in shady, disturbed, and moist understory habitats, and that the aquatic habit also arose early. This would have required the capacity to begin life in dimly lit environments. If so, evolution in light-sensing mechanisms may have been crucial to their success. The photoreceptor phytochrome A is unique among angiosperm phytochromes in its capacity to serve a transient role under conditions where an extremely high sensitivity is required. We present evidence of altered functional constraints between phytochrome A (PHYA) and its paralog, PHYC. Tests for selection suggest that an elevation in nonsynonymous rates resulted from an episode of selection along the branch leading to all angiosperm PHYA sequences. Most nucleotide sites (95%) are selectively constrained, and the ratio of nonsynonymous to synonymous substitutions on branches within the PHYA clade does not differ from the ratio on the branches in the PHYC clade. Thus, positive selection at a handful of sites, rather than relaxation of selective constraints, apparently has played a major role in the evolution of the photosensory domain of phytochrome A. The episode of selection occurred very early in the history of flowering plants, suggesting that innovation in phyA may have given the first angiosperms some adaptive advantage.

Patterns of nucleotide substitution among simultaneously duplicated gene pairs in Arabidopsis thaliana.

We characterized rates and patterns of synonymous and nonsynonymous substitution in 242 duplicated gene pairs on chromosomes 2 and 4 of Arabidopsis thaliana. Based on their collinear order along the two chromosomes, the gene pairs were likely duplicated contemporaneously, and therefore comparison of genetic distances among gene pairs provides insights into the distribution of nucleotide substitution rates among plant nuclear genes. Rates of synonymous substitution varied 13.8-fold among the duplicated gene pairs, but 90% of gene pairs differed by less than 2.6-fold. Average nonsynonymous rates were approximately fivefold lower than average synonymous rates; this rate difference is lower than that of previously studied nonplant lineages. The coefficient of variation of rates among genes was 0.65 for nonsynonymous rates and 0.44 for synonymous rates, indicating that synonymous and nonsynonymous rates vary among genes to roughly the same extent. The causes underlying rate variation were explored. Our analyses tentatively suggest an effect of physical location on synonymous substitution rates but no similar effect on nonsynonymous rates. Nonsynonymous substitution rates were negatively correlated with GC content at synonymous third codon positions, and synonymous substitution rates were negatively correlated with codon bias, as observed in other systems. Finally, the 242 gene pairs permitted investigation of the processes underlying divergence between paralogs. We found no evidence of positive selection, little evidence that paralogs evolve at different rates, and no evidence of differential codon usage or third position GC content.

Synonymous-non-synonymous mutation rates between sequences containing ambiguous nucleotides (Syn-SCAN).

Direct PCR sequencing on genetic material containing allelic mixtures results in sequences containing ambiguous nucleotides. Because codons exhibiting allelic mixtures present evidence of evolutionary pressure, it is important to include this information in the assessment of codon synonymy. We developed a program, 'Synonymous-Nonsynonymous Mutation Rates between Sequences Containing Ambiguous Nucleotides' (Syn-SCAN), that calculates synonymous and non-synonymous substitution rates using a model that includes allelic mixtures. Syn-SCAN is implemented on the web and can be downloaded from http://hivdb.stanford.edu.

Phylogenetic analysis of Japanese encephalitis virus: envelope gene based analysis reveals a fifth genotype, geographic clustering, and multiple introductions of the virus into the Indian subcontinent.

We report the analysis of the complete nucleotide sequence for the Indian isolate (P20778; Genbank Accession number AF080251) of Japanese encephalitis virus (JEV). The phylogenetic tree topology obtained using thirteen complete genome sequences of JEV was reproduced with the envelope, NS1, NS3, and NS5 genes and revealed extensive divergence between the two Indian strains included. A more exhaustive analysis of JEV evolution using 107 envelope sequences available for isolates from different geographic locations worldwide revealed five distinct genotypes of JEV, displaying a minimum nucleotide divergence of 7% with high bootstrap support values. The tree also revealed overall clustering of strains based on geographic location, as well as multiple introductions of JEV into the Indian subcontinent. Nonsynonymous nucleotide divergence rates of the envelope gene estimated that the ancestor common to all JEV genotypes arose within the last three hundred years.

A Role for Selection in Regulating the Evolutionary Emergence of Disease-Causing and Other Coding CAG Repeats in Humans and Mice

The evolutionary expansion of CAG repeats in human triplet expansion disease genes is intriguing because of their deleterious phenotype. In the past, this expansion has been suggested to reflect a broad genomewide expansion of repeats, which would imply that mutational and evolutionary processes acting on repeats differ between species. Here, we tested this hypothesis by analyzing repeat- and flanking-sequence evolution in 28 repeat-containing genes that had been sequenced in humans and mice and by considering overall lengths and distributions of CAG repeats in the two species. We found no evidence that these repeats were longer in humans than in mice. We also found no evidence for preferential accumulation of CAG repeats in the human genome relative to mice from an analysis of the lengths of repeats identified in sequence databases. We then investigated whether sequence properties, such as base and amino acid composition and base substitution rates, showed any relationship to repeat evolution. We found that repeat-containing genes were enriched in certain amino acids, presumably as the result of selection, but that this did not reflect underlying biases in base composition. We also found that regions near repeats showed higher nonsynonymous substitution rates than the remainder of the gene and lower nonsynonymous rates in genes that contained a repeat in both the human and the mouse. Higher rates of nonsynonymous mutation in the neighborhood of repeats presumably reflect weaker purifying selection acting in these regions of the proteins, while the very low rate of nonsynonymous mutation in proteins containing a CAG repeat in both species presumably reflects a high level of purifying selection. Based on these observations, we propose that the mutational processes giving rise to polyglutamine repeats in human and murine proteins do not differ. Instead, we propose that the evolution of polyglutamine repeats in proteins results from an interplay between mutational processes and selection.

Substitution rates in Drosophila nuclear genes: implications for translational selection.

The relationships between synonymous and nonsynonymous substitution rates and between synonymous rate and codon usage bias are important to our understanding of the roles of mutation and selection in the evolution of Drosophila genes. Previous studies used approximate estimation methods that ignore codon bias. In this study we reexamine those relationships using maximum-likelihood methods to estimate substitution rates, which accommodate the transition/transversion rate bias and codon usage bias. We compiled a sample of homologous DNA sequences at 83 nuclear loci from Drosophila melanogaster and at least one other species of Drosophila. Our analysis was consistent with previous studies in finding that synonymous rates were positively correlated with nonsynonymous rates. Our analysis differed from previous studies, however, in that synonymous rates were unrelated to codon bias. We therefore conducted a simulation study to investigate the differences between approaches. The results suggested that failure to properly account for multiple substitutions at the same site and for biased codon usage by approximate methods can lead to an artifactual correlation between synonymous rate and codon bias. Implications of the results for translational selection are discussed.

Rates of nucleotide substitution and mammalian nuclear gene evolution. Approximate and maximum-likelihood methods lead to different conclusions.

Rates and patterns of synonymous and nonsynonymous substitutions have important implications for the origin and maintenance of mammalian isochores and the effectiveness of selection at synonymous sites. Previous studies of mammalian nuclear genes largely employed approximate methods to estimate rates of nonsynonymous and synonymous substitutions. Because these methods did not account for major features of DNA sequence evolution such as transition/transversion rate bias and unequal codon usage, they might not have produced reliable results. To evaluate the impact of the estimation method, we analyzed a sample of 82 nuclear genes from the mammalian orders Artiodactyla, Primates, and Rodentia using both approximate and maximum-likelihood methods. Maximum-likelihood analysis indicated that synonymous substitution rates were positively correlated with GC content at the third codon positions, but independent of nonsynonymous substitution rates. Approximate methods, however, indicated that synonymous substitution rates were independent of GC content at the third codon positions, but were positively correlated with nonsynonymous rates. Failure to properly account for transition/transversion rate bias and unequal codon usage appears to have caused substantial biases in approximate estimates of substitution rates.

The effect of tandem substitutions on the correlation between synonymous and nonsynonymous rates in rodents.

Nonsynonymous substitutions in DNA cause amino acid substitutions while synonymous substitutions in DNA leave amino acids unchanged. The cause of the correlation between the substitution rates at nonsynonymous (K(A)) and synonymous (K(S)) sites in mammals is a contentious issue, and one that impacts on many aspects of molecular evolution. Here we use a large set of orthologous mammalian genes to investigate the causes of the K(A)-K(S) correlation in rodents. The strength of the K(A)-K(S) correlation exceeds the neutral theory expectation when substitution rates are estimated using algorithmic methods, but not when substitution rates are estimated by maximum likelihood. Irrespective of this methodological uncertainty the strength of the K(A)-K(S) correlation appears mostly due to tandem substitutions, an excess of which is generated by substitutional nonindependence. Doublet mutations cannot explain the excess of tandem synonymous-nonsynonymous substitutions, and substitution patterns indicate that selection on silent sites is the likely cause. We find no evidence for selection on codon usage. The nature of the relationship between synonymous divergence and base composition is unclear because we find a significant correlation if we use maximum-likelihood methods but not if we use algorithmic methods. Finally, we find that K(S) is reduced at the start of genes, which suggests that selection for RNA structure may affect silent sites in mammalian protein-coding genes.