Genetic robustness and selection at the protein level for synonymous codons

Synonymous codons are neutral at the protein level, therefore natural selection at the protein level should have no effect on their frequencies. Synonymous codons, however, differ in their capacity to reduce the effects of errors: after mutation, certain codons keep on coding for the same amino acid or for amino acids with similar properties, while other synonymous codons produce very different amino acids. Therefore, the impact of errors on a coding sequence (genetic robustness) can be measured by analysing its codon usage. I analyse the codon usage of sequenced nuclear and cytoplasmic genomes and I show that there is an extensive variation in genetic robustness at the DNA sequence level, both among genomes and among genes of the same genome. I also show theoretically that robustness can be adaptive, that is natural selection may lead to a preference for codons that reduce the impact of errors. If selection occurs only among the mutants of a codon (e.g. among the progeny before the adult phase), however, the codons that are more sensitive to the effects of mutations may increase in frequency because they manage to get rid more easily of deleterious mutations. I also suggest other possible explanations for the evolution of genetic robustness at the codon level.     

Comparative Genomics of Trypanosomatid Pathogens using Codon Usage Bias

It is well known that an amino acid can be encoded by more than one codon, called synonymous codons. The preferential use of one particular codon for coding an amino acid is referred to as codon usage bias (CUB). A quantitative analytical method, CUB and a related tool, Codon Adaptative Index have been applied to comparatively study whole genomes of a few pathogenic Trypanosomatid species. This quantitative attempt is of direct help in the comparison of qualitative features like mutational and translational selection. Pathogens of the Leishmania and Trypanosoma genus cause debilitating disease and suffering in human beings and animals. Of these, whole genome sequences are available for only five species. The complete coding sequences (CDS), highly expressed, essential and low expressed genes have all been studied for their CUB signature. The codon usage bias of essential genes and highly expressed genes show distribution similar to codon usage bias of all CDSs in Trypanosomatids. Translational selection is the dominant force selecting the preferred codon, and selection due to mutation is negligible. In contrast to an earlier study done on these pathogens, it is found in this work that CUB and CAI may be used to distinguish the Trypanosomatid genomes at the sub-genus level. Further, CUB may effectively be used as a signature of the species differentiation by using Principal Component Analysis (PCA).

Abbreviations

CUB - Codon Usage Bias, CAI - Codon Adaptative Index, CDS - Coding sequences, t-RNA - Transfer RNA, PCA - Principal Component Analysis.     

Effect of codon message on xylanase thermal activity

Because the genetic codon is known for degeneracy, its effect on enzyme thermal property is seldom investigated. A dataset was constructed for GH10 xylanase coding sequences and optimal temperatures for activity (T(opt)). Codon contents and relative synonymous codon usages were calculated and respectively correlated with the enzyme T(opt) values, which were used to describe the xylanase thermophilic tendencies without dividing them into two thermophilic and mesophilic groups. After analyses of codon content and relative synonymous codon usages were checked by the Bonferroni correction, we found five codons, with three (AUA, AGA, and AGG) correlating positively and two (CGU and AGC) correlating negatively with the T(opt) value. The three positive codons are purine-rich codons, and the two negative codons have A-ends. The two negative codons are pyridine-rich codons, and one has a C-end. Comparable with the codon C- and A-ending features, C- and A-content within mRNA correlated negatively and positively with the T(opt) value, respectively. Thereby, codons have effects on enzyme thermal property. When the issue is analyzed at the residual level, the effect of codon message is lost. The codons relating to enzyme thermal property are selected by thermophilic force at nucleotide level.     

Selection on Synonymous Sites for Increased Accessibility around miRNA Binding Sites in Plants

Synonymous codons are widely selected for various biological mechanisms in both prokaryotes and eukaryotes. Recent evidence suggests that microRNA (miRNA) function may affect synonymous codon choices near miRNA target sites. To better understand this, we perform genome-wide analysis on synonymous codon usage around miRNA target sites in four plant genomes. We observed a general trend of increased site accessibility around miRNA target sites in plants. Guanine-cytosine (GC)-poor codons are preferred in the flank region of miRNA target sites. Within-genome analyses show significant variation among miRNA targets in species. GC content of the target gene can partly explain the variation of site accessibility among miRNA targets. miRNA targets in GC-rich genes show stronger selection signals than those in GC-poor genes. Gene's codon usage bias and the conservation level of miRNA and its target also have some effects on site accessibility, but the expression level of miRNA or its target and the mechanism of miRNA activity do not contribute to site accessibility differences among miRNA targets. We suggest that synonymous codons near miRNA targets are selected for efficient miRNA binding and proper miRNA function. Our results present a new dimension of natural selection on synonymous codons near miRNA target sites in plants, which will have important implications of coding sequence evolution.     

Selected codon usage bias in members of the class Mollicutes

Mollicutes are parasitic microorganisms mainly characterized by small cell sizes, reduced genomes and great A and T mutational bias. We analyzed the codon usage patterns of the completely sequenced genomes of bacteria that belong to this class. We found that for many organisms not only mutational bias but also selection has a major effect on codon usage. Through a comparative perspective and based on three widely used criteria we were able to classify Mollicutes according to the effect of selection on codon usage. We found conserved optimal codons in many species and study the tRNA gene pool in each genome. Previous results are reinforced by the fact that, when selection is operative, the putative optimal codons found match the respective cognate tRNA. Finally, we trace selection effect backwards to the common ancestor of the class and estimate the phylogenetic inertia associated with this character. We discuss the possible scenarios that explain the observed evolutionary patterns.     

GC-biased gene conversion and selection affect GC content in the Oryza genus (rice)

Base composition varies among and within eukaryote genomes. Although mutational bias and selection have initially been invoked, more recently GC-biased gene conversion (gBGC) has been proposed to play a central role in shaping nucleotide landscapes, especially in yeast, mammals, and birds. gBGC is a kind of meiotic drive in favor of G and C alleles, associated with recombination. Previous studies have also suggested that gBGC could be at work in grass genomes. However, these studies were carried on third codon positions that can undergo selection on codon usage. As most preferred codons end in G or C in grasses, gBGC and selection can be confounded. Here we investigated further the forces that might drive GC content evolution in the rice genus using both coding and noncoding sequences. We found that recombination rates correlate positively with equilibrium GC content and that selfing species (Oryza sativa and O. glaberrima) have significantly lower equilibrium GC content compared with more outcrossing species. As recombination is less efficient in selfing species, these results suggest that recombination drives GC content. We also detected a positive relationship between expression levels and GC content in third codon positions, suggesting that selection favors codons ending with G or C bases. However, the correlation between GC content and recombination cannot be explained by selection on codon usage alone as it was also observed in noncoding positions. Finally, analyses of polymorphism data ruled out the hypothesis that genomic variation in GC content is due to mutational processes. Our results suggest that both gBGC and selection on codon usage affect GC content in the Oryza genus and likely in other grass species.     

Mutation and Selection Cause Codon Usage and Bias in Mitochondrial Genomes of Ribbon Worms (Nemertea)

The phenomenon of codon usage bias is known to exist in many genomes and it is mainly determined by mutation and selection. To understand the patterns of codon usage in nemertean mitochondrial genomes, we use bioinformatic approaches to analyze the protein-coding sequences of eight nemertean species. Neutrality analysis did not find a significant correlation between GC12 and GC3. ENc-plot showed a few genes on or close to the expected curve, but the majority of points with low-ENc values are below it. ENc-plot suggested that mutational bias plays a major role in shaping codon usage. The Parity Rule 2 plot (PR2) analysis showed that GC and AT were not used proportionally and we propose that codons containing A or U at third position are used preferentially in nemertean species, regardless of whether corresponding tRNAs are encoded in the mitochondrial DNA. Context-dependent analysis indicated that the nucleotide at the second codon position slightly affects synonymous codon choices. These results suggested that mutational and selection forces are probably acting to codon usage bias in nemertean mitochondrial genomes.     

How Mitochondria Redefine the Code

Annotated, complete DNA sequences are available for 213 mitochondrial genomes from 132 species. These provide an extensive sample of evolutionary adjustment of codon usage and meaning spanning the history of this organelle. Because most known coding changes are mitochondrial, such data bear on the general mechanism of codon reassignment. Coding changes have been attributed variously to loss of codons due to changes in directional mutation affecting the genome GC content (Osawa and Jukes 1988), to pressure to reduce the number of mitochondrial tRNAs to minimize the genome size (Anderson and Kurland 1991), and to the existence of transitional coding mechanisms in which translation is ambiguous (Schultz and Yarus 1994a). We find that a succession of such steps explains existing reassignments well. In particular, (1) Genomic variation in the prevalence of a codon's third-position nucleotide predicts relative mitochondrial codon usage well, though GC content does not. This is because A and T, and G and C, are uncorrelated in mitochondrial genomes. (2) Codons predicted to reach zero usage (disappear) do so more often than expected by chance, and codons that do disappear are disproportionately likely to be reassigned. However, codons predicted to disappear are not significantly more likely to be reassigned. Therefore, low codon frequencies can be related to codon reassignment, but appear to be neither necessary nor sufficient for reassignment. (3) Changes in the genetic code are not more likely to accompany smaller numbers of tRNA genes and are not more frequent in smaller genomes. Thus, mitochondrial codons are not reassigned during demonstrable selection for decreased genome size. Instead, the data suggest that both codon disappearance and codon reassignment depend on at least one other event. This mitochondrial event (leading to reassignment) occurs more frequently when a codon has disappeared, and produces only a small subset of possible reassignments. We suggest that coding ambiguity, the extension of a tRNA's decoding capacity beyond its original set of codons, is the second event. Ambiguity can act alone but often acts in concert with codon disappearance, which promotes codon reassignment. Received: 26 October 2000 / Accepted: 19 January 2001     

Translational Selection Is Ubiquitous in Prokaryotes

Codon usage bias in prokaryotic genomes is largely a consequence of background substitution patterns in DNA, but highly expressed genes may show a preference towards codons that enable more efficient and/or accurate translation. We introduce a novel approach based on supervised machine learning that detects effects of translational selection on genes, while controlling for local variation in nucleotide substitution patterns represented as sequence composition of intergenic DNA. A cornerstone of our method is a Random Forest classifier that outperformed previous distance measure-based approaches, such as the codon adaptation index, in the task of discerning the (highly expressed) ribosomal protein genes by their codon frequencies. Unlike previous reports, we show evidence that translational selection in prokaryotes is practically universal: in 460 of 461 examined microbial genomes, we find that a subset of genes shows a higher codon usage similarity to the ribosomal proteins than would be expected from the local sequence composition. These genes constitute a substantial part of the genome—between 5% and 33%, depending on genome size—while also exhibiting higher experimentally measured mRNA abundances and tending toward codons that match tRNA anticodons by canonical base pairing. Certain gene functional categories are generally enriched with, or depleted of codon-optimized genes, the trends of enrichment/depletion being conserved between Archaea and Bacteria. Prominent exceptions from these trends might indicate genes with alternative physiological roles; we speculate on specific examples related to detoxication of oxygen radicals and ammonia and to possible misannotations of asparaginyl–tRNA synthetases. Since the presence of codon optimizations on genes is a valid proxy for expression levels in fully sequenced genomes, we provide an example of an “adaptome” by highlighting gene functions with expression levels elevated specifically in thermophilic Bacteria and Archaea.     

Evolutionary Lability of Context-Dependent Codon Bias in Bacteria

In bacteria, synonymous codon usage can be considerably affected by base composition at neighboring sites. Such context-dependent biases may be caused by either selection against specific nucleotide motifs or context-dependent mutation biases. Here we consider the evolutionary conservation of context-dependent codon bias across 11 completely sequenced bacterial genomes. In particular, we focus on two contextual biases previously identified in Escherichia coli; the avoidance of out-of-frame stop codons and AGG motifs. By identifying homologues of E. coli genes, we also investigate the effect of gene expression level in Haemophilus influenzae and Mycoplasma genitalium. We find that while context-dependent codon biases are widespread in bacteria, few are conserved across all species considered. Avoidance of out-of-frame stop codons does not apply to all stop codons or amino acids in E. coli, does not hold for different species, does not increase with gene expression level, and is not relaxed in Mycoplasma spp., in which the canonical stop codon, TGA, is recognized as tryptophan. Avoidance of AGG motifs shows some evolutionary conservation and increases with gene expression level in E. coli, suggestive of the action of selection, but the cause of the bias differs between species. These results demonstrate that strong context-dependent forces, both selective and mutational, operate on synonymous codon usage but that these differ considerably between genomes. Received: 6 May 1999 / Accepted: 29 October 1999     

SCUMBLE: a method for systematic and accurate detection of codon usage bias by maximum likelihood estimation

The genetic code is degenerate—most amino acids can be encoded by from two to as many as six different codons. The synonymous codons are not used with equal frequency: not only are some codons favored over others, but also their usage can vary significantly from species to species and between different genes in the same organism. Known causes of codon bias include differences in mutation rates as well as selection pressure related to the expression level of a gene, but the standard analysis methods can account for only a fraction of the observed codon usage variation. We here introduce an explicit model of codon usage bias, inspired by statistical physics. Combining this model with a maximum likelihood approach, we are able to clearly identify different sources of bias in various genomes. We have applied the algorithm to Saccharomyces cerevisiae as well as 325 prokaryote genomes, and in most cases our model explains essentially all observed variance.     

Analysis of synonymous codon usage patterns in the genus Rhizobium

The codon usage patterns of rhizobia have received increasing attention. However, little information is available regarding the conserved features of the codon usage patterns in a typical rhizobial genus. The codon usage patterns of six completely sequenced strains belonging to the genus Rhizobium were analysed as model rhizobia in the present study. The relative neutrality plot showed that selection pressure played a role in codon usage in the genus Rhizobium. Spearman’s rank correlation analysis combined with correspondence analysis (COA) showed that the codon adaptation index and the effective number of codons (ENC) had strong correlation with the first axis of the COA, which indicated the important role of gene expression level and the ENC in the codon usage patterns in this genus. The relative synonymous codon usage of Cys codons had the strongest correlation with the second axis of the COA. Accordingly, the usage of Cys codons was another important factor that shaped the codon usage patterns in Rhizobium genomes and was a conserved feature of the genus. Moreover, the comparison of codon usage between highly and lowly expressed genes showed that 20 unique preferred codons were shared among Rhizobium genomes, revealing another conserved feature of the genus. This is the first report of the codon usage patterns in the genus Rhizobium.     

The evolution of biased codon and amino acid usage in nematode genomes

Despite the degeneracy of the genetic code, whereby different codons encode the same amino acid, alternative codons and amino acids are utilized nonrandomly within and between genomes. Such biases in codon and amino acid usage have been demonstrated extensively in prokaryote genomes and likely reflect a balance between the action of mutation, selection, and genetic drift. Here, we quantify the effects of selection and mutation drift as causes of codon and amino acid-usage bias in a large collection of nematode partial genomes from 37 species spanning approximately 700 Myr of evolution, as inferred from expressed sequence tag (EST) measures of gene expression and from base composition variation. Average G + C content at silent sites among these taxa ranges from 10% to 63%, and EST counts range more than 100-fold, underlying marked differences between the identities of major codons and optimal codons for a given species as well as influencing patterns of amino acid abundance among taxa. Few species in our sample demonstrate a dominant role of selection in shaping intragenomic codon-usage biases, and these are principally free living rather than parasitic nematodes. This suggests that deviations in effective population size among species, with small effective sizes among parasites, are partly responsible for species differences in the extent to which selection shapes patterns of codon usage. Nevertheless, a consensus set of optimal codons emerges that is common to most taxa, indicating that, with some notable exceptions, selection for translational efficiency and accuracy favors similar sets of codons regardless of the major codon-usage trends defined by base compositional properties of individual nematode genomes.     

Comparison of base composition and codon usage in insect mitochondrial genomes

Insects, the most biodiverse taxonomic group, have high AT content in their mitochondrial genomes. Although codon usage tends to be AT-rich, base composition and codon usage of mitochondrial genomes may vary among taxa. Thus, we compare base composition and codon usage patterns of 49 insect mitochondrial genomes. For protein coding genes, AT content is as high as 80% in the Hymenoptera and Lepidoptera and as low as 72% in the Orthopotera. The AT content is high at positions 1 and 3, but A content is low at position 2. A close correlation occurs between codon usage and tRNA abundance in nuclear genomes. Optimal codons can pair well with the antr codons of the most abundant tRNAs. One tRNA gene translates a synonymous codon family in vertebrate mitochondrial genomes and these tRNA anticodons can pair with optimal codons. However, optimal codons cannot pair with anticodons in mtDNA ofCochiiomyia hominivorax (Dipteral: CaLliphoridae). Ten optimal codons cannot pair with tRNA anticodons in all 49 insect mitochondrial genomes; non-optimal codon-anticodon usage is common and codon usage is not influenced by tRNA abundance.     

皱边喉毛花及其近缘物种的叶绿体基因组结构与进化分析

为重建喉毛花属下系统发育关系,明晰属下皱边喉毛花及其近缘种之间的物种关系。本研究利用Illumina高通量测序平台对12 个叶绿体基因组进行双末端测序,获得大量高质量的Clean reads用于后续生物信息学分析。结果表明：（1）喉毛花属下物种的基因组差异较小,均在150 kb左右,基因总数为131 个,其中编码基因81 个。IR区核苷酸多态性比SC低,编码区比非编码区更保守。（2）进化分析结果显示,几乎所有的编码基因受到纯化选择的作用。（3）密码子偏好性分析表明有35 个密码子的RSCU值均大于1,说明使用这些密码子的频率较高,各项密码子偏好性衡量指标说明喉毛花属物种的密码子偏好性较弱。（4）系统发育分析表明CDS、密码子位置与基因间隔区数据集构建的系统发育树具有高度一致的拓扑结构,大部分分支的支持率高。这些结果表明皱边喉毛花及其近缘种的叶绿体基因组无明显差异,在系统发育树上无法按物种聚类,也为后续展开喉毛花属下群体遗传学研究提供科学依据。     

Comparative studies on codon usage pattern of chloroplasts and their host nuclear genes in four plant species

A detailed comparison was made of codon usage of chloroplast genes with their host (nuclear) genes in the four angiosperm speciesOryza sativa, Zea mays, Triticum aestivum andArabidopsis thaliana. The average GC content of the entire genes, and at the three codon positions individually, was higher in nuclear than in chloroplast genes, suggesting different genomic organization and mutation pressures in nuclear and chloroplast genes. The results of Nc-plots and neutrality plots suggested that nucleotide compositional constraint had a large contribution to codon usage bias of nuclear genes inO. sativa, Z. mays, andT. aestivum, whereas natural selection was likely to be playing a large role in codon usage bias in chloroplast genomes. Correspondence analysis and chi-test showed that regardless of the genomic environment (species) of the host, the codon usage pattern of chloroplast genes differed from nuclear genes of their host species by their AU-richness. All the chloroplast genomes have predominantly A- and/or U-ending codons, whereas nuclear genomes have G-, C- or U-ending codons as their optimal codons. These findings suggest that the chloroplast genome might display particular characteristics of codon usage that are different from its host nuclear genome. However, one feature common to both chloroplast and nuclear genomes in this study was that pyrimidines were found more frequently than purines at the synonymous codon position of optimal codons.     

Selection on GGU and CGU Codons in the High Expression Genes in Bacteria

The fourfold degenerate site (FDS) in coding sequences is important for studying the effect of any selection pressure on codon usage bias (CUB) because nucleotide substitution per se is not under any such pressure at the site due to the unaltered amino acid sequence in a protein. We estimated the frequency variation of nucleotides at the FDS across the eight family boxes (FBs) defined as Um(g), the unevenness measure of a gene g. The study was made in 545 species of bacteria. In many bacteria, the Um(g) correlated strongly with Nc′—a measure of the CUB. Analysis of the strongly correlated bacteria revealed that the U-ending codons (GGU, CGU) were preferred to the G-ending codons (GGG, CGG) in Gly and Arg FBs even in the genomes with G+C % higher than 65.0. Further evidence suggested that these codons can be used as a good indicator of selection pressure on CUB in genomes with higher G+C %.     

Strong Heterogeneity in Nucleotidic Composition and Codon Bias in the Pea Aphid (<Emphasis Type="Italic">Acyrthosiphon pisum)</Emphasis> Shown by EST-Based Coding Genome Reconstruction

The aim of this study was to analyze patterns of nucleotidic composition and codon usage in the pea aphid genome (Acyrthosiphon pisum). A collection of 60,000 expressed sequence tags (ESTs) in the pea aphid has been used to automatically reconstruct 5809 coding sequences (CDSs), based on similarity with known proteins and on coding style recognition. Reconstructions were manually checked for ribosomal proteins, leading to tentatively reconstruct the nea-complete set of this category. Pea aphid coding sequences showed a shift toward AT (especially at the third codon position) compared to drosophila homologues. Genes with a putative high level of expression (ribosomal and other genes with high EST support) remained more GC3-rich and had a distinct codon usage from bulk sequences: they exhibited a preference for C-ending codons and CGT (for arginine), which thus appeared optimal for translation. However, the discrimination was not as strong as in drosophila, suggesting a reduced degree of translational selection. The space of variation in codon usage for A. pisum appeared to be larger than in drosophila, with a substantial fraction of genes that remained GC3-rich. Some of those (in particular some structural proteins) also showed high levels of codon bias and a very strong preference for C-ending codons, which could be explained either by strong translational selection or by other mechanisms. Finally, genomic traces were analyzed to build 206 fragments containing a full CDS, which allowed studying the correlations between GC contents of coding and those of noncoding (flanking and introns) sequences.