Aminoacyl-tRNA Synthetases, the Genetic Code, and the Evolutionary Process

The aminoacyl-tRNA synthetases (AARSs) and their relationship to the genetic code are examined from the evolutionary perspective. Despite a loose correlation between codon assignments and AARS evolutionary relationships, the code is far too highly structured to have been ordered merely through the evolutionary wanderings of these enzymes. Nevertheless, the AARSs are very informative about the evolutionary process. Examination of the phylogenetic trees for each of the AARSs reveals the following. (i) Their evolutionary relationships mostly conform to established organismal phylogeny: a strong distinction exists between bacterial- and archaeal-type AARSs. (ii) Although the evolutionary profiles of the individual AARSs might be expected to be similar in general respects, they are not. It is argued that these differences in profiles reflect the stages in the evolutionary process when the taxonomic distributions of the individual AARSs became fixed, not the nature of the individual enzymes. (iii) Horizontal transfer of AARS genes between Bacteria and Archaea is asymmetric: transfer of archaeal AARSs to the Bacteria is more prevalent than the reverse, which is seen only for the “gemini group.” (iv) The most far-ranging transfers of AARS genes have tended to occur in the distant evolutionary past, before or during formation of the primary organismal domains. These findings are also used to refine the theory that at the evolutionary stage represented by the root of the universal phylogenetic tree, cells were far more primitive than their modern counterparts and thus exchanged genetic material in far less restricted ways, in effect evolving in a communal sense.     

Aminoacyl-tRNA synthetases database

Aminoacyl-tRNA synthetases (AARSs) are at the center of the question of the origin of life. They constitute a family of enzymes integrating the two levels of cellular organization: nucleic acids and proteins. AARSs arose early in evolution and are believed to be a group of ancient proteins. They are responsible for attaching amino acid residues to their cognate tRNA molecules, which is the first step in the protein synthesis. The role they play in a living cell is essential for the precise deciphering of the genetic code. The analysis of AARSs evolutionary history was not possible for a long time due to a lack of a sufficiently large number of their amino acid sequences. The emerging picture of synthetases' evolution is a result of recent achievements in genomics [Woese,C., Olsen,G.J., Ibba,M. and S?ll,D. (2000) Microbiol. Mol. Biol. Rev., 64, 202-236]. In this paper we present a short introduction to the AARSs database. The updated database contains 1047 AARS primary structures from archaebacteria, eubacteria, mitochondria, chloroplasts and eukaryotic cells. It is the compilation of amino acid sequences of all AARSs known to date, which are available as separate entries via the WWW at http://biobases.ibch.poznan.pl/aars/.     

Intraphylum diversity and complex evolution of cyanobacterial aminoacyl-tRNA synthetases

A comparative genomic analysis of 35 cyanobacterial strains has revealed that the gene complement of aminoacyl-tRNA synthetases (AARSs) and routes for aminoacyl-tRNA synthesis may differ among the species of this phylum. Several genes encoding AARS paralogues were identified in some genomes. In-depth phylogenetic analysis was done for each of these proteins to gain insight into their evolutionary history. GluRS, HisRS, ArgRS, ThrRS, CysRS, and Glu-Q-RS showed evidence of a complex evolutionary course as indicated by a number of inconsistencies with our reference tree for cyanobacterial phylogeny. In addition to sequence data, support for evolutionary hypotheses involving horizontal gene transfer or gene duplication events was obtained from other observations including biased sequence conservation, the presence of indels (insertions or deletions), or vestigial traces of ancestral redundant genes. We present evidences for a novel protein domain with two putative transmembrane helices recruited independently by distinct AARS in particular cyanobacteria.     

Aminoacyl-tRNA synthetases database Y2K

The aminoacyl-tRNA synthetases (AARS) are a diverse group of enzymes that ensure the fidelity of transfer of genetic information from DNA into protein. They catalyse the attachment of amino acids to transfer RNAs and thereby establish the rules of the genetic code by virtue of matching the nucleotide triplet of the anticodon with its cognate amino acid. Currently, 818 AARS primary structures have been reported from archaebacteria, eubacteria, mitochondria, chloro-plasts and eukaryotic cells. The database is a compilation of the amino acid sequences of all AARSs, known to date, which are available as separate entries or alignments of related proteins via the WWW at http://rose.man.poznan.pl/aars/index.html     

MIST,a Novel Approach to Reveal Hidden Substrate Specificity in Aminoacyl-tRNA Synthetases

Aminoacyl-tRNA synthetases (AARSs) constitute a family of RNA-binding proteins, that participate in the translation of the genetic code, by covalently linking amino acids to appropriate tRNAs. Due to their fundamental importance for cell life, AARSs are likely to be one of the most ancient families of enzymes and have therefore been characterized extensively. Paradoxically, little is known about their capacity to discriminate tRNAs mainly because of the practical challenges that represent precise and systematic tRNA identification. This work describes a new technical and conceptual approach named MIST (Microarray Identification of Shifted tRNAs) designed to study the formation of tRNA/AARS complexes independently from the aminoacylation reaction. MIST combines electrophoretic mobility shift assays with microarray analyses. Although MIST is a non-cellular assay, it fully integrates the notion of tRNA competition. In this study we focus on yeast cytoplasmic Arginyl-tRNA synthetase (yArgRS) and investigate in depth its ability to discriminate cellular tRNAs. We report that yArgRS in submicromolar concentrations binds cognate and non-cognate tRNAs with a wide range of apparent affinities. In particular, we demonstrate that yArgRS binds preferentially to type II tRNAs but does not support their misaminoacylation. Our results reveal important new trends in tRNA/AARS complex formation and potential deep physiological implications.     

Ancient gene duplications and the root(s) of the tree of life

Summary. Tracing organismal histories on the timescale of the tree of life remains one of the challenging tasks in evolutionary biology. The hotly debated questions include the evolutionary relationship between the three domains of life (e.g., which of the three domains are sister domains, are the domains para-, poly-, or monophyletic) and the location of the root within the universal tree of life. For the latter, many different points of view have been considered but so far no consensus has been reached. The only widely accepted rationale to root the universal tree of life is to use anciently duplicated paralogous genes that are present in all three domains of life. To date only few anciently duplicated gene families useful for phylogenetic reconstruction have been identified. Here we present results from a systematic search for ancient gene duplications using twelve representative, completely sequenced, archaeal and bacterial genomes. Phylogenetic analyses of identified cases show that the majority of datasets support a root between Archaea and Bacteria; however, some datasets support alternative hypotheses, and all of them suffer from a lack of strong phylogenetic signal. The results are discussed with respect to the impact of horizontal gene transfer on the ability to reconstruct organismal evolution. The exchange of genetic information between divergent organisms gives rise to mosaic genomes, where different genes in a genome have different histories. Simulations show that even low rates of horizontal gene transfer dramatically complicate the reconstruction of organismal evolution, and that the different most recent common molecular ancestors likely existed at different times and in different lineages. Correspondence and reprints: Department of Molecular and Cell Biology, University of Connecticut, Storrs, CT 06269-3125, U.S.A. Present address: Genome Atlantic, Department of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada.     

Evolution of arginine deiminase (ADI) pathway genes

We have analyzed the evolution of the three genes encoding structural enzymes of the arginine deiminase (ADI) pathway, arginine deiminase (ADI), ornithine transcarbamoylase (OTC), and carbamate kinase (CK) in a wide range of organisms, including Archaea, Bacteria, and Eukarya. This catabolic route was probably present in the last common ancestor to all the domains of life. The results obtained indicate that these genes have undergone a complex evolutionary history, including horizontal transfer events, duplications, and losses. Therefore, these genes are not adequate to infer organismal relationships at deep branching levels, but they provide an insight into how catabolic genes evolved and were assembled into metabolic pathways. Our results suggest that the three genes evolved independently and were later assembled into a single cluster with functional interdependence, thus, providing support for the gene recruitment hypothesis. Furthermore, the molecular phylogenetic analysis of OTC suggests a new classification of these genes into three subfamilies.     

Negative catalysis by the editing domain of class I aminoacyl-tRNA synthetases

Aminoacyl-tRNA synthetases (AARS) translate the genetic code by loading tRNAs with the cognate amino acids. The errors in amino acid recognition are cleared at the AARS editing domain through hydrolysis of misaminoacyl-tRNAs. This ensures faithful protein synthesis and cellular fitness. Using Escherichia coli isoleucyl-tRNA synthetase (IleRS) as a model enzyme, we demonstrated that the class I editing domain clears the non-cognate amino acids well-discriminated at the synthetic site with the same rates as the weakly-discriminated fidelity threats. This unveiled low selectivity suggests that evolutionary pressure to optimize the rates against the amino acids that jeopardize translational fidelity did not shape the editing site. Instead, we propose that editing was shaped to safeguard cognate aminoacyl-tRNAs against hydrolysis. Misediting is prevented by the residues that promote negative catalysis through destabilisation of the transition state comprising cognate amino acid. Such powerful design allows broad substrate acceptance of the editing domain along with its exquisite specificity in the cognate aminoacyl-tRNA rejection. Editing proceeds by direct substrate delivery to the editing domain (in cis pathway). However, we found that class I IleRS also releases misaminoacyl-tRNA^Ile and edits it in trans. This minor editing pathway was up to now recognized only for class II AARSs.     

Aminoacyl-tRNA synthetases of the multi-tRNA synthetase complex and their role in tumorigenesis

In mammalian cells, 20 aminoacyl-tRNA synthetases (AARS) catalyze the ligation of amino acids to their cognate tRNAs to generate aminoacylated-tRNAs. In higher eukaryotes, 9 of the 20 AARSs, along with 3 auxiliary proteins, join to form the cytoplasmic multi-tRNA synthetase complex (MSC). The complex is absent in prokaryotes, but evolutionary expansion of MSC constituents, primarily by addition of novel interacting domains, facilitates formation of subcomplexes that join to establish the holo-MSC. In some cases, environmental cues direct the release of constituents from the MSC which enables the execution of non-canonical, i.e., “moonlighting”, functions distinct from their essential activities in protein translation. These activities are generally beneficial, but can also be deleterious to the cell. Elucidation of the non-canonical activities of several AARSs residing in the MSC suggest they are potential therapeutic targets for cancer, as well as metabolic and neurologic diseases. Here, we describe the role of MSC-resident AARSs in cancer progression, and the factors that regulate their release from the MSC. Also, we highlight recent developments in therapeutic modalities that target MSC AARSs for cancer prevention and treatment.     

Requirement of aminoacyl-tRNA synthetases for gametogenesis and embryo development in Arabidopsis

Aminoacyl-tRNA synthetases (AARSs) are required for translation in three different compartments of the plant cell: chloroplasts, mitochondria and the cytosol. Elimination of this basal function should result in lethality early in development. Phenotypes of individual mutants may vary considerably, depending on patterns of gene expression, functional redundancy, allele strength and protein localization. We describe here a reverse genetic screen of 50 insertion mutants disrupted in 21 of the 45 predicted AARSs in Arabidopsis. Our initial goal was to find additional EMB genes with a loss-of-function phenotype in the seed. Several different classes of knockouts were discovered, with defects in both gametogenesis and seed development. Three major trends were observed. Disruption of translation in chloroplasts often results in seed abortion at the transition stage of embryogenesis with minimal effects on gametophytes. Disruption of translation in mitochondria often results in ovule abortion before and immediately after fertilization. This early phenotype was frequently missed in prior screens for embryo-defective mutants. Knockout alleles of non-redundant cytosolic AARSs were in general not identified, consistent with the absolute requirement of cytosolic translation for development of male and female gametophytes. These results provide a framework for evaluating redundant functions of AARSs in Arabidopsis, a valuable data set of phenotypes resulting from multiple disruptions of a single basal process, and insights into which genes are required for both gametogenesis and embryo development and might therefore escape detection in screens for embryo-defective mutants.     

The new aspects of aminoacyl-tRNA synthetases

Aminoacyl-tRNA synthetases (AARS) are essential proteins found in all living organisms. They form a diverse group of enzymes that ensure the fidelity of transfer of genetic information from the DNA into the protein. AARS catalyse the attachment of amino acids to transfer RNAs and thereby establish the rules of the genetic code by virtue of matching the nucleotide triplet of the anticodon with its cognate amino acid. Here we summarise the effects of recent studies on this interesting family of multifunctional enzymes.     

Calculation of Evolutionary Correlation between Individual Genes and Full-Length Genome: A Method Useful for Choosing Phylogenetic Markers for Molecular Epidemiology

Individual genes or regions are still commonly used to estimate the phylogenetic relationships among viral isolates. The genomic regions that can faithfully provide assessments consistent with those predicted with full-length genome sequences would be preferable to serve as good candidates of the phylogenetic markers for molecular epidemiological studies of many viruses. Here we employed a statistical method to evaluate the evolutionary relationships between individual viral genes and full-length genomes without tree construction as a way to determine which gene can match the genome well in phylogenetic analyses. This method was performed by calculation of linear correlations between the genetic distance matrices of aligned individual gene sequences and aligned genome sequences. We applied this method to the phylogenetic analyses of porcine circovirus 2 (PCV2), measles virus (MV), hepatitis E virus (HEV) and Japanese encephalitis virus (JEV). Phylogenetic trees were constructed for comparisons and the possible factors affecting the method accuracy were also discussed in the calculations. The results revealed that this method could produce results consistent with those of previous studies about the proper consensus sequences that could be successfully used as phylogenetic markers. And our results also suggested that these evolutionary correlations could provide useful information for identifying genes that could be used effectively to infer the genetic relationships.     

F-box基因拷贝数目变异的机制研究:以12种果蝇为例

拷贝数目变异是一种对表型变异和生物进化具有重要意义的基因组结构变异.以前的研究表明不同物种中F-box基因的拷贝数目差异较大.为了深入探索拷贝数目变异的式样和机制.我们以12个果蝇近缘种为研究对象,分析了F-box基因的系统发育关系、进化式样以及它们在染色体上的位置.结果发现,虽然各个物种中F-box基因的拷贝数目差别不大(42-47个),但是仍然存在着很多引起拷贝数目变异的基因获得和丢失事件.这说明表面上变化不大的拷贝数目在一定程度上掩盖了频繁发生的基因获得和丢失事件.通过比较这些基因在染色体上的位置,发现只有在亲缘关系很近的物种之间才能鉴定出有明显微共线性关系的基因组区段.我们还发现,造成F-box基因拷贝数目增加的主要机制是散在重复和串联重复,而反转录转座和新基因的非编码区起源也是两种值得注意的机制.此外,序列变异导致的外显子边界变化以及外显子丢失是引起拷贝数目减少的两种机制.在12种果蝇的最近共同祖先中,F-box基因的拷贝数目与现存物种基本相似,但是基因的获得和丢失事件使得现存物种中的F-box基因在构成上已经有了明显的差别.对数目变异的式样及其与基因功能的关系的研究表明,拷贝数目变异是F-box基因家族"生与死"的进化在基因组层面的系统反映,并有可能为表型变异提供了原始材料.     

Predicting sub-cellular localization of tRNA synthetases from their primary structures

Since endo-symbiotic events occur, all genes of mitochondrial aminoacyl tRNA synthetase (AARS) were lost or transferred from ancestral mitochondrial genome into the nucleus. The canonical pattern is that both cytosolic and mitochondrial AARSs coexist in the nuclear genome. In the present scenario all mitochondrial AARSs are nucleus-encoded, synthesized on cytosolic ribosomes and post-translationally imported from the cytosol into the mitochondria in eukaryotic cell. The site-based discrimination between similar types of enzymes is very challenging because they have almost same physico-chemical properties. It is very important to predict the sub-cellular location of AARSs, to understand the mitochondrial protein synthesis. We have analyzed and optimized the distinguishable patterns between cytosolic and mitochondrial AARSs. Firstly, support vector machines (SVM)-based modules have been developed using amino acid and dipeptide compositions and achieved Mathews correlation coefficient (MCC) of 0.82 and 0.73, respectively. Secondly, we have developed SVM modules using position-specific scoring matrix and achieved the maximum MCC of 0.78. Thirdly, we developed SVM modules using N-terminal, intermediate residues, C-terminal and split amino acid composition (SAAC) and achieved MCC of 0.82, 0.70, 0.39 and 0.86, respectively. Finally, a SVM module was developed using selected attributes of split amino acid composition (SA-SAAC) approach and achieved MCC of 0.92 with an accuracy of 96.00%. All modules were trained and tested on a non-redundant data set and evaluated using fivefold cross-validation technique. On the independent data sets, SA-SAAC based prediction model achieved MCC of 0.95 with an accuracy of 97.77%. The web-server 'MARSpred' based on above study is available at http://www.imtech.res.in/raghava/marspred/.     

The bacterial chromosome

Bacteria represent the vast majority of biological diversity found on Earth. In this review, we focus on selected aspects of their genetic material, those providing insight into structural, functional, dynamic, and evolutionary aspects of their genomes. Bacterial chromosomes are far more dynamic than previously realized, and dozens of mechanisms giving rise to genomic plasticity are now understood. Maturation of the genomics era has provided the tools for unraveling the interwoven details of DNA structure/function relationships that provide a basis for organismal diversity. Some of the most throughly understood processes that underlie the dynamics of genomic structure and function in prokaryotes are examined.     

The Bacterial Chromosome

Bacteria represent the vast majority of biological diversity found on Earth. In this review, we focus on selected aspects of their genetic material, those providing insight into structural, functional, dynamic, and evolutionary aspects of their genomes. Bacterial chromosomes are far more dynamic than previously realized, and dozens of mechanisms giving rise to genomic plasticity are now understood. Maturation of the genomics era has provided the tools for unraveling the interwoven details of DNA structure/function relationships that provide a basis for organismal diversity. Some of the most throughly understood processes that underlie the dynamics of genomic structure and function in prokaryotes are examined.     

Eukaryotic origin of glyceraldehyde-3-phosphate dehydrogenase genes in Clostridium thermocellum and Clostridium cellulolyticum genomes and putative fates of the exogenous gene in the subsequent genome evolution

Although lateral gene transfer (LGT) events have been frequently documented in the evolution of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), no eukaryote-to-prokaryote transfer has been reported so far. Here we describe the first case of the GAPDH gene transfer from a eukaryote to a subset of Clostridium species (Bacteria, Firmicutes). A series of phylogenetic analyses of GAPDH homologues revealed that Clostridium thermocellum and Clostridium cellulolyticum homologues have the evolutionary affinity to the eukaryotic homologues, rather than to those of bacterial species closely related to the two Clostridium species in the organismal phylogeny. These results suggest that the GAPDH genes in the two Clostridium species are of eukaryotic origin, which is the first reported case of eukaryote-to-bacterium GAPDH gene transfer. Since a previously published 16S ribosomal DNA phylogeny and our GAPDH phylogeny commonly suggest an intimate evolutionary relationship between C. thermocellum and C. cellulolyticum, a common ancestor of the two species likely acquired the eukaryotic GAPDH gene. In the C. cellulolyticum genome, the exogenous GAPDH gene was physically separated from other glycolytic genes, suggesting that this gene organization was likely achieved by a random insertion of the laterally transferred gene. On the other hand, in the C. thermocellum genome, the laterally transferred GAPDH gene clusters with other bacterial glycolytic genes. We discuss possible scenarios for the evolutionarily chimeric glycolytic gene cluster in the C. thermocellum genome.     

Evolutionary relationships of adenylation domains in fungi

Non-ribosomal peptide synthetases (NRPSs) and NRPS-like enzymes are abundant in microbes as they are involved in the production of primary and secondary metabolites. In contrast to the well-studied NRPSs, known to produce non-ribosomal peptides, NRPS-like enzymes exhibit more diverse activities and their evolutionary relationships are unclear. Here, we present the first in-depth phylogenetic analysis of fungal NRPS-like A domains from functionally characterized pathways, and their relationships to characterized A domains found in fungal NRPSs. This study clearly differentiated amino acid reductases, including NRPSs, from CoA/AMP ligases, which could be divided into 10 distinct phylogenetic clades that reflect their conserved domain organization, substrate specificity and enzymatic activity. In particular, evolutionary relationships of adenylate forming reductases could be refined and explained the substrate specificity difference. Consistent with their phylogeny, the deduced amino acid code of A domains differentiated amino acid reductases from other enzymes. However, a diagnostic code was found for α-keto acid reductases and clade 7 CoA/AMP ligases only. Comparative genomics of loci containing these enzymes revealed that they can be independently recruited as tailoring genes in diverse secondary metabolite pathways. Based on these results, we propose a refined and clear phylogeny-based classification of A domain-containing enzymes, which will provide a robust framework for future functional analyses and engineering of these enzymes to produce new bioactive molecules.     

The evolution of aminoacyl-tRNA synthetases,the biosynthetic pathways of amino acids and the genetic code

In this paper the partition metric is used to compare binary trees deriving from (i) the study of the evolutionary relationships between aminoacyl-tRNA synthetases, (ii) the physicochemical properties of amino acids and (iii) the biosynthetic relationships between amino acids. If the tree defining the evolutionary relationships between aminoacyl-tRNA synthetases is assumed to be a manifestation of the mechanism that originated the organization of the genetic code, then the results appear to indicate the following: the hypothesis that regards the genetic code as a map of the biosynthetic relationships between amino acids seems to explain the organization of the genetic code, at least as plausibly as the hypotheses that consider the physicochemical properties of amino acids as the main adaptive theme that lead to the structuring of the code.