Comparative Analysis of Codon Usage Bias and Codon Context Patterns between Dipteran and Hymenopteran Sequenced Genomes

Background

Codon bias is a phenomenon of non-uniform usage of codons whereas codon context generally refers to sequential pair of codons in a gene. Although genome sequencing of multiple species of dipteran and hymenopteran insects have been completed only a few of these species have been analyzed for codon usage bias.

Methods and Principal Findings

Here, we use bioinformatics approaches to analyze codon usage bias and codon context patterns in a genome-wide manner among 15 dipteran and 7 hymenopteran insect species. Results show that GAA is the most frequent codon in the dipteran species whereas GAG is the most frequent codon in the hymenopteran species. Data reveals that codons ending with C or G are frequently used in the dipteran genomes whereas codons ending with A or T are frequently used in the hymenopteran genomes. Synonymous codon usage orders (SCUO) vary within genomes in a pattern that seems to be distinct for each species. Based on comparison of 30 one-to-one orthologous genes among 17 species, the fruit fly Drosophila willistoni shows the least codon usage bias whereas the honey bee (Apis mellifera) shows the highest bias. Analysis of codon context patterns of these insects shows that specific codons are frequently used as the 3′- and 5′-context of start and stop codons, respectively.

Conclusions

Codon bias pattern is distinct between dipteran and hymenopteran insects. While codon bias is favored by high GC content of dipteran genomes, high AT content of genes favors biased usage of synonymous codons in the hymenopteran insects. Also, codon context patterns vary among these species largely according to their phylogeny.     

Comparative analysis of codon usage patterns in chloroplast genomes of the Asteraceae family

Codon usage bias (CUB) is an important evolutionary feature in a genome and has been widely documented from prokaryotes to eukaryotes. However, the significance of CUB in the Asteraceae family has not been well understood, with no Asteraceae species having been analyzed for this characteristic. Here, we use bioinformatics approaches to comparatively analyze the general patterns and influencing factors of CUB in five Asteraceae chloroplast (cp) genomes. The results indicated that the five genomes had similar codon usage patterns, showing a strong bias towards a high representation of NNA and NNT codons. Neutrality analysis showed that these cp genomes had a narrow GC distribution and no significant correlation was observed between GC₁₂ and GC₃. Parity Rule 2 (PR2) plot analysis revealed that purines were used more frequently than pyrimidines. Effective number of codons (ENc)-plot analysis showed that most genes followed the parabolic line of trajectory, but several genes with low ENc values lying below the expected curve were also observed. Furthermore, correspondence analysis of relative synonymous codon usage (RSCU) yielded a first axis that explained only a partial amount of variation of codon usage. These findings suggested that both natural selection and mutational bias contributed to codon bias, while selection was the major force to shape the codon usage in these Asteraceae cp genomes. Our study, which is the first to investigate codon usage patterns in Asteraceae plastomes, will provide helpful information about codon distribution and variation in these species, and also shed light on the genetic and evolutionary mechanisms of codon biology within this family.     

Codon usage bias: causative factors,quantification methods and genome‐wide patterns: with emphasis on insect genomes

Codon usage bias refers to the phenomenon where specific codons are used more often than other synonymous codons during translation of genes, the extent of which varies within and among species. Molecular evolutionary investigations suggest that codon bias is manifested as a result of balance between mutational and translational selection of such genes and that this phenomenon is widespread across species and may contribute to genome evolution in a significant manner. With the advent of whole‐genome sequencing of numerous species, both prokaryotes and eukaryotes, genome‐wide patterns of codon bias are emerging in different organisms. Various factors such as expression level, GC content, recombination rates, RNA stability, codon position, gene length and others (including environmental stress and population size) can influence codon usage bias within and among species. Moreover, there has been a continuous quest towards developing new concepts and tools to measure the extent of codon usage bias of genes. In this review, we outline the fundamental concepts of evolution of the genetic code, discuss various factors that may influence biased usage of synonymous codons and then outline different principles and methods of measurement of codon usage bias. Finally, we discuss selected studies performed using whole‐genome sequences of different insect species to show how codon bias patterns vary within and among genomes. We conclude with generalized remarks on specific emerging aspects of codon bias studies and highlight the recent explosion of genome‐sequencing efforts on arthropods (such as twelve Drosophila species, species of ants, honeybee, Nasonia and Anopheles mosquitoes as well as the recent launch of a genome‐sequencing project involving 5000 insects and other arthropods) that may help us to understand better the evolution of codon bias and its biological significance.     

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.     

A general model of codon bias due to GC mutational bias

Background

In spite of extensive research on the effect of mutation and selection on codon usage, a general model of codon usage bias due to mutational bias has been lacking. Because most amino acids allow synonymous GC content changing substitutions in the third codon position, the overall GC bias of a genome or genomic region is highly correlated with GC3, a measure of third position GC content. For individual amino acids as well, G/C ending codons usage generally increases with increasing GC bias and decreases with increasing AT bias. Arginine and leucine, amino acids that allow GC-changing synonymous substitutions in the first and third codon positions, have codons which may be expected to show different usage patterns.

Principal Findings

In analyzing codon usage bias in hundreds of prokaryotic and plant genomes and in human genes, we find that two G-ending codons, AGG (arginine) and TTG (leucine), unlike all other G/C-ending codons, show overall usage that decreases with increasing GC bias, contrary to the usual expectation that G/C-ending codon usage should increase with increasing genomic GC bias. Moreover, the usage of some codons appears nonlinear, even nonmonotone, as a function of GC bias. To explain these observations, we propose a continuous-time Markov chain model of GC-biased synonymous substitution. This model correctly predicts the qualitative usage patterns of all codons, including nonlinear codon usage in isoleucine, arginine and leucine. The model accounts for 72%, 64% and 52% of the observed variability of codon usage in prokaryotes, plants and human respectively. When codons are grouped based on common GC content, 87%, 80% and 68% of the variation in usage is explained for prokaryotes, plants and human respectively.

Conclusions

The model clarifies the sometimes-counterintuitive effects that GC mutational bias can have on codon usage, quantifies the influence of GC mutational bias and provides a natural null model relative to which other influences on codon bias may be measured.     

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.     

Analysis of synonymous codon usage patterns in different plant mitochondrial genomes

Codon usage in mitochondrial genome of the six different plants was analyzed to find general patterns of codon usage in plant mitochondrial genomes. The neutrality analysis indicated that the codon usage patterns of mitochondrial genes were more conserved in GC content and no correlation between GC12 and GC3. T and A ending codons were detected as the preferred codons in plant mitochondrial genomes. The Parity Rule 2 plot analysis showed that T was used more frequently than A. The EN_C-plot showed that although a majority of the points with low EN_C values were lying below the expected curve, a few genes lied on the expected curve. Correspondence analysis of relative synonymous codon usage yielded a first axis that explained only a partial amount of variation of codon usage. These findings suggest that natural selection is likely to be playing a large role in codon usage bias in plant mitochondrial genomes, but not only natural selection but also other several factors are likely to be involved in determining the selective constraints on codon bias in plant mitochondrial genomes. Meantime, 1 codon (P. patens), 6 codons (Z. mays), 9 codons (T. aestivum), 15 codons (A. thaliana), 15 codons (M. polymorpha) and 15 codons (N. tabacum) were defined as the preferred codons of the six plant mitochondrial genomes.     

Patterns of codon usage in two ciliates that reassign the genetic code: Tetrahymena thermophila and Paramecium tetraurelia

We used the recently sequenced genomes of the ciliates Tetrahymena thermophila and Paramecium tetraurelia to analyze the codon usage patterns in both organisms; we have analyzed codon usage bias, Gln codon usage, GC content and the nucleotide contexts of initiation and termination codons in Tetrahymena and Paramecium. We also studied how these trends change along the length of the genes and in a subset of highly expressed genes. Our results corroborate some of the trends previously described in Tetrahymena, but also negate some specific observations. In both genomes we found a strong bias toward codons with low GC content; however, in highly expressed genes this bias is smaller and codons ending in GC tend to be more frequent. We also found that codon bias increases along gene segments and in highly expressed genes and that the context surrounding initiation and termination codons are always AT rich. Our results also suggest differences in the efficiency of translation of the reassigned stop codons between the two species and between the reassigned codons. Finally, we discuss some of the possible causes for such translational efficiency differences.     

Mutation bias is the driving force of codon usage in the Gallus gallus genome

Synonymous codons are used with different frequencies both among species and among genes within the same genome and are controlled by neutral processes (such as mutation and drift) as well as by selection. Up to now, a systematic examination of the codon usage for the chicken genome has not been performed. Here, we carried out a whole genome analysis of the chicken genome by the use of the relative synonymous codon usage (RSCU) method and identified 11 putative optimal codons, all of them ending with uracil (U), which is significantly departing from the pattern observed in other eukaryotes. Optimal codons in the chicken genome are most likely the ones corresponding to highly expressed transfer RNA (tRNAs) or tRNA gene copy numbers in the cell. Codon bias, measured as the frequency of optimal codons (Fop), is negatively correlated with the G + C content, recombination rate, but positively correlated with gene expression, protein length, gene length and intron length. The positive correlation between codon bias and protein, gene and intron length is quite different from other multi-cellular organism, as this trend has been only found in unicellular organisms. Our data displayed that regional G + C content explains a large proportion of the variance of codon bias in chicken. Stepwise selection model analyses indicate that G + C content of coding sequence is the most important factor for codon bias. It appears that variation in the G + C content of CDSs accounts for over 60% of the variation of codon bias. This study suggests that both mutation bias and selection contribute to codon bias. However, mutation bias is the driving force of the codon usage in the Gallus gallus genome. Our data also provide evidence that the negative correlation between codon bias and recombination rates in G. gallus is determined mostly by recombination-dependent mutational patterns.     

The Selective Advantage of Synonymous Codon Usage Bias in Salmonella

The genetic code in mRNA is redundant, with 61 sense codons translated into 20 different amino acids. Individual amino acids are encoded by up to six different codons but within codon families some are used more frequently than others. This phenomenon is referred to as synonymous codon usage bias. The genomes of free-living unicellular organisms such as bacteria have an extreme codon usage bias and the degree of bias differs between genes within the same genome. The strong positive correlation between codon usage bias and gene expression levels in many microorganisms is attributed to selection for translational efficiency. However, this putative selective advantage has never been measured in bacteria and theoretical estimates vary widely. By systematically exchanging optimal codons for synonymous codons in the tuf genes we quantified the selective advantage of biased codon usage in highly expressed genes to be in the range 0.2–4.2 x 10⁻⁴ per codon per generation. These data quantify for the first time the potential for selection on synonymous codon choice to drive genome-wide sequence evolution in bacteria, and in particular to optimize the sequences of highly expressed genes. This quantification may have predictive applications in the design of synthetic genes and for heterologous gene expression in biotechnology.     

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.     

Gene expression,nucleotide composition and codon usage bias of genes associated with human Y chromosome

Analysis of codon usage pattern is important to understand the genetic and evolutionary characteristics of genomes. We have used bioinformatic approaches to analyze the codon usage bias (CUB) of the genes located in human Y chromosome. Codon bias index (CBI) indicated that the overall extent of codon usage bias was low. The relative synonymous codon usage (RSCU) analysis suggested that approximately half of the codons out of 59 synonymous codons were most frequently used, and possessed a T or G at the third codon position. The codon usage pattern was different in different genes as revealed from correspondence analysis (COA). A significant correlation between effective number of codons (ENC) and various GC contents suggests that both mutation pressure and natural selection affect the codon usage pattern of genes located in human Y chromosome. In addition, Y-linked genes have significant difference in GC contents at the second and third codon positions, expression level, and codon usage pattern of some codons like the SPANX genes in X chromosome.     

On Ribosome Load,Codon Bias and Protein Abundance

Different codons encoding the same amino acid are not used equally in protein-coding sequences. In bacteria, there is a bias towards codons with high translation rates. This bias is most pronounced in highly expressed proteins, but a recent study of synthetic GFP-coding sequences did not find a correlation between codon usage and GFP expression, suggesting that such correlation in natural sequences is not a simple property of translational mechanisms. Here, we investigate the effect of evolutionary forces on codon usage. The relation between codon bias and protein abundance is quantitatively analyzed based on the hypothesis that codon bias evolved to ensure the efficient usage of ribosomes, a precious commodity for fast growing cells. An explicit fitness landscape is formulated based on bacterial growth laws to relate protein abundance and ribosomal load. The model leads to a quantitative relation between codon bias and protein abundance, which accounts for a substantial part of the observed bias for E. coli. Moreover, by providing an evolutionary link, the ribosome load model resolves the apparent conflict between the observed relation of protein abundance and codon bias in natural sequences and the lack of such dependence in a synthetic gfp library. Finally, we show that the relation between codon usage and protein abundance can be used to predict protein abundance from genomic sequence data alone without adjustable parameters.     

Variation in the strength of selected codon usage bias among bacteria

Among bacteria, many species have synonymous codon usage patterns that have been influenced by natural selection for those codons that are translated more accurately and/or efficiently. However, in other species selection appears to have been ineffective. Here, we introduce a population genetics-based model for quantifying the extent to which selection has been effective. The approach is applied to 80 phylogenetically diverse bacterial species for which whole genome sequences are available. The strength of selected codon usage bias, S, is found to vary substantially among species; in 30% of the genomes examined, there was no significant evidence that selection had been effective. Values of S are highly positively correlated with both the number of rRNA operons and the number of tRNA genes. These results are consistent with the hypothesis that species exposed to selection for rapid growth have more rRNA operons, more tRNA genes and more strongly selected codon usage bias. For example, Clostridium perfringens, the species with the highest value of S, can have a generation time as short as 7 min.     

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.     

Study of Completed Archaeal Genomes and Proteomes： Hypothesis of Strong Mutational AT Pressure Existed in Their Common Predecessor

The number of completely sequenced archaeal genomes has been sufficient for a large-scale bioinformatic study.We have conducted analyses for each coding region from 36 archaeal genomes using the original CGS algorithm by calculating the total GC content(G+C),GC content in first,second and third codon positions as well as in fourfold and twofold degenerated sites from third codon positions,levels of arginine codon usage(Arg2:AGA/G;Arg4:CGX),levels of amino acid usage and the entropy of amino acid content distribution.In archaeal genomes with strong GC pressure,arginine is coded preferably by GC-rich Arg4 codons,whereas in most of archaeal genomes with G+C0.6,arginine is coded preferably by AT-rich Arg2 codons.In the genome of Haloquadratum walsbyi,which is closely related to GC-rich archaea,GC content has decreased mostly in third codon positions,while Arg4Arg2 bias still persists.Proteomes of archaeal species carry characteristic amino acid biases:levels of isoleucine and lysine are elevated,while levels of alanine,histidine,glutamine and cytosine are relatively decreased.Numerous genomic and proteomic biases observed can be explained by the hypothesis of previously existed strong mutational AT pressure in the common predecessor of all archaea.     

Gaining Insights into the Codon Usage Patterns of TP53 Gene across Eight Mammalian Species

TP53 gene is known as the “guardian of the genome” as it plays a vital role in regulating cell cycle, cell proliferation, DNA damage repair, initiation of programmed cell death and suppressing tumor growth. Non uniform usage of synonymous codons for a specific amino acid during translation of protein known as codon usage bias (CUB) is a unique property of the genome and shows species specific deviation. Analysis of codon usage bias with compositional dynamics of coding sequences has contributed to the better understanding of the molecular mechanism and the evolution of a particular gene. In this study, the complete nucleotide coding sequences of TP53 gene from eight different mammalian species were used for CUB analysis. Our results showed that the codon usage patterns in TP53 gene across different mammalian species has been influenced by GC bias particularly GC₃ and a moderate bias exists in the codon usage of TP53 gene. Moreover, we observed that nature has highly favored the most over represented codon CTG for leucine amino acid but selected against the ATA codon for isoleucine in TP53 gene across all mammalian species during the course of evolution.     

Translationally optimal codons associate with aggregation-prone sites in proteins

We analyze the relationship between codon usage bias and residue aggregation propensity in the genomes of four model organisms, Escherichia coli, yeast, fly, and mouse, as well as the archaeon Halobacterium species NRC-1. Using the Mantel-Haenszel procedure, we find that translationally optimal codons associate with aggregation-prone residues. Our results are qualitatively and quantitatively similar to those of an earlier study where we found an association between translationally optimal codons and buried residues. We also combine the aggregation-propensity data with solvent-accessibility data. Although the resulting data set is small, and hence statistical power low, results indicate that the association between optimal codons and aggregation-prone residues exists both at buried and at exposed sites. By comparing codon usage at different combinations of sites (exposed, aggregation-prone sites versus buried, non-aggregation-prone sites; buried, aggregation-prone sites versus exposed, non-aggregation-prone sites), we find that aggregation propensity and solvent accessibility seem to have independent effects of (on average) comparable magnitude on codon usage. Finally, in fly, we assess whether optimal codons associate with sites at which amino acid substitutions lead to an increase in aggregation propensity, and find only a very weak effect. These results suggest that optimal codons may be required to reduce the frequency of translation errors at aggregation-prone sites that coincide with certain functional sites, such as protein-protein interfaces. Alternatively, optimal codons may be required for rapid translation of aggregation-prone regions.