Alternative methods for concatenation of core genes indicate a lack of resolution in deep nodes of the prokaryotic phylogeny.

It has recently been proposed that a well-resolved Tree of Life can be achieved through concatenation of shared genes. There are, however, several difficulties with such an approach, especially in the prokaryotic part of this tree. We tackled some of them using a new combination of maximum likelihood-based methods, developed in order to practice as safe and careful concatenations as possible. First, we used the application concaterpillar on carefully aligned core genes. This application uses a hierarchical likelihood-ratio test framework to assess both the topological congruence between gene phylogenies (i.e., whether different genes share the same evolutionary history) and branch-length congruence (i.e., whether genes that share the same history share the same pattern of relative evolutionary rates). We thus tested if these core genes can be concatenated or should be instead categorized into different incongruent sets. Second, we developed a heat map approach studying the evolution of the phylogenetic support for different bipartitions, when the number of sites of different phylogenetic quality in the concatenation increases. These heatmaps allow us to follow which phylogenetic signals increase or decrease as the concatenation progresses and to detect emerging artifactual groupings, that is, groups that are more and more supported when more and more homoplasic sites are thrown in the analysis. We showed that, as far as 7 major prokaryotic lineages are concerned, only 22 core genes can be said to be congruent and can be safely concatenated. This number is even smaller than the number of genes retained to reconstruct a "Tree of One Per Cent." Furthermore, the concatenation of these 22 markers leads to an unresolved tree as the only groupings in the concatenation tree seem to reflect emerging artifacts. Using concatenated core genes as a valid framework to classify uncharacterized environmental sequences can thus be misleading.

Lateral genomics.

More than 20 complete prokaryotic genome sequences are now publicly available, each by itself an unparalleled resource for understanding organismal biology. Collectively, these data are even more powerful: they could force a dramatic reworking of the framework in which we understand biological evolution. It is possible that a single universal phylogenetic tree is not the best way to depict relationships between all living and extinct species. Instead a web- or net-like pattern, reflecting the importance of horizontal or lateral gene transfer between lineages of organisms, might provide a more appropriate visual metaphor. Here, I ask whether this way of thinking is really justified, and explore its implications.

Phylogenetic classification and the universal tree.

From comparative analyses of the nucleotide sequences of genes encoding ribosomal RNAs and several proteins, molecular phylogeneticists have constructed a "universal tree of life," taking it as the basis for a "natural" hierarchical classification of all living things. Although confidence in some of the tree's early branches has recently been shaken, new approaches could still resolve many methodological uncertainties. More challenging is evidence that most archaeal and bacterial genomes (and the inferred ancestral eukaryotic nuclear genome) contain genes from multiple sources. If "chimerism" or "lateral gene transfer" cannot be dismissed as trivial in extent or limited to special categories of genes, then no hierarchical universal classification can be taken as natural. Molecular phylogeneticists will have failed to find the "true tree," not because their methods are inadequate or because they have chosen the wrong genes, but because the history of life cannot properly be represented as a tree. However, taxonomies based on molecular sequences will remain indispensable, and understanding of the evolutionary process will ultimately be enriched, not impoverished.

Molecular biological mechanisms of speciation.

Growing recognition that much of the evolutionary history of eukaryotic genomes reflects the operation of turnover processes involving repetitive DNA sequences has led to the recent formulation of models describing speciation as a consequence of such turnover. These models are of three general kinds: those attributing hybrid infertility to the process of transposition, those attributing hybrid infertility to mispairing between chromosomes of divergent repetitive DNA composition, and those assuming that change in repetitive DNA's can reset coordinated gene regulation. These models are discussed with respect to the kinds of evidence needed for their corroboration and to their significance for questions related to macroevolutionary punctuated equilibria and genetic revolutions.

Testing the exon theory of genes: the evidence from protein structure.

A tendency for exons to correspond to discrete units of protein structure in protein-coding genes of ancient origin would provide clear evidence in favor of the exon theory of genes, which proposes that split genes arose not by insertion of introns into unsplit genes, but from combinations of primordial mini-genes (exons) separated by spacers (introns). Although putative examples of such correspondence have strongly influenced previous debate on the origin of introns, a general correspondence has not been rigorously proved. Objective methods for detecting correspondences were developed and applied to four examples that have been cited previously as evidence of the exon theory of genes. No significant correspondence between exons and units of protein structure was detected, suggesting that the putative correspondence does not exist and that the exon theory of genes is untenable.

A kingdom-level phylogeny of eukaryotes based on combined protein data.

Current understanding of the higher order systematics of eukaryotes relies largely on analyses of the small ribosomal subunit RNA (SSU rRNA). Independent testing of these results is still limited. We have combined the sequences of four of the most broadly taxonomically sampled proteins available to create a roughly parallel data set to that of SSU rRNA. The resulting phylogenetic tree shows a number of striking differences from SSU rRNA phylogeny, including strong support for most major groups and several major supergroups.

Archaea and the prokaryote-to-eukaryote transition.

Since the late 1970s, determining the phylogenetic relationships among the contemporary domains of life, the Archaea (archaebacteria), Bacteria (eubacteria), and Eucarya (eukaryotes), has been central to the study of early cellular evolution. The two salient issues surrounding the universal tree of life are whether all three domains are monophyletic (i.e., all equivalent in taxanomic rank) and where the root of the universal tree lies. Evaluation of the status of the Archaea has become key to answering these questions. This review considers our cumulative knowledge about the Archaea in relationship to the Bacteria and Eucarya. Particular attention is paid to the recent use of molecular phylogenetic approaches to reconstructing the tree of life. In this regard, the phylogenetic analyses of more than 60 proteins are reviewed and presented in the context of their participation in major biochemical pathways. Although many gene trees are incongruent, the majority do suggest a sisterhood between Archaea and Eucarya. Altering this general pattern of gene evolution are two kinds of potential interdomain gene transferrals. One horizontal gene exchange might have involved the gram-positive Bacteria and the Archaea, while the other might have occurred between proteobacteria and eukaryotes and might have been mediated by endosymbiosis.

Integron-associated gene cassettes in Halifax Harbour: assessment of a mobile gene pool in marine sediments.

The integron/gene cassette systems identified in bacteria comprise a class of genetic elements that allow adaptation by acquisition of gene cassettes. Integron gene cassettes have been shown to facilitate the spread of drug resistance in human pathogens but their role outside a clinical setting has not been explored extensively. We sequenced 2145 integron gene cassettes from four marine sediment samples taken from the vicinity of Halifax Nova Scotia, Canada, increasing the number of gene cassettes obtained from environmental microbial communities by 10-fold. Sequence analyses reveals that the majority of these cassettes encode novel proteins and that this study is consistent with previous claims of high cassette diversity as we estimate a Chao1 diversity index of approximately 3000 cassettes from these samples. The functional distribution of environmental cassettes recovered in this study, when compared with that of cassettes from the only other source with significant sampling (Vibrio genomes) suggests that alternate selection regimes might be acting on these two gene pools. The majority of cassettes recovered in this study encode novel, unknown proteins. In instances where we obtained multiple alleles of a novel protein we demonstrate that non-synonymous versus synonymous substitution rates ratios suggest relaxed selection. Cassette-encoded proteins with known homologues represent a variety of functions and prevalent among these are isochorismatases; proteins involved in iron scavenging. Phylogenetic analysis of these isochorismatases as well as of cassette-encoded acetyltransferases reveals a patchy distribution, suggesting multiple sources for the origin of these cassettes. Finally, the two most environmentally similar sample sites considered in this study display the greatest overlap of cassette types, consistent with the hypothesis that cassette genes encode adaptive proteins.

Archaebacterial genomes: eubacterial form and eukaryotic content.

Since the recognition of the uniqueness and coherence of the archaebacteria (sometimes called Archaea), our perception of their role in early evolution has been modified repeatedly. The deluge of sequence data and rapidly improving molecular systematic methods have combined with a better understanding of archaebacterial molecular biology to describe a group that in some ways appears to be very similar to the eubacteria, though in others is more like the eukaryotes. The structure and contents of archaebacterial genomes are examined here, with an eye to their meaning in terms of the evolution of cell structure and function.

Evolution. Archaea and eukaryotes versus bacteria?

The recent discovery of homologs of the eukaryotic transcription factor TATA-binding protein in archaea has been taken as support for the view that archaea and eukaryotes have a close phylogenetic relationship.

Cytoskeletal proteins: the evolution of cell division.

The prokaryotic cell division protein FtsZ and eukaryotic tubulin have been shown to have very similar structures and are most likely homologs. The evolutionary transition from FtsZ to tubulin could provide a window into the transition from prokaryotic cells to eukaryotic cells.

Lessons from the Aeropyrum pernix genome.

Aeropyrum pernix is the first crenarchaeote and first aerobic member of the Archaea for which the complete genome sequence has been determined. The sequence confirms the distinct nature of crenarchaeotes and provides new insight into the relationships between the three domains: Bacteria, Archaea and Eukaryotes.

Archaeal genomics: do archaea have a mixed heritage?

A third complete archaeal genome sequence, replete with eukaryote-like genes for replication, transcription and translation, has appeared. The sequence also shows bacteria-like features. It is time to come to grips with this evidence for a mixed heritage.

Selfish DNA: the best defense is a good offense.

The recent discovery of novel biochemical activities of intron-encoded endonucleases emphasizes the selfish nature of mobile genetic elements.

Recurrent paralogy in the evolution of archaeal chaperonins.

Chaperonins are multisubunit double-ring complexes that mediate the folding of nascent proteins [1] [2]. In bacteria, chaperonins are homo-oligomeric and are composed of seven-membered rings. Eukaryotic and most archaeal chaperonin rings are eight-membered and exhibit varying degrees of hetero-oligomerism [3] [4]. We have cloned and sequenced seven new genes encoding chaperonin subunits from the crenarchaeotes Sulfolobus solfataricus, S. acidocaldarius, S. shibatae and Desulfurococcus mobilis. Although some archaeal genomes possess a single chaperonin gene, most have two. We describe a third chaperonin-encoding gene (TF55-gamma) from two Sulfolobus species; phylogenetic analyses indicate that the gene duplication producing TF55-gamma occurred within crenarchaeal evolution. The presence of TF55-gamma in Sulfolobus correlates with their unique nine-membered chaperonin rings. Duplicate genes (paralogs) for chaperonins within archaeal genomes very often resemble each other more than they resemble chaperonin genes from other archaea. Our phylogenetic analyses suggest multiple independent gene duplications - at least seven among the archaea examined. The persistence of paralogous genes for chaperonin subunits in multiple archaeal lineages may involve a process of co-evolution, where chaperonin subunit heterogeneity changes independently of selection on function.

Molecular evolution: recent cases of spliceosomal intron gain?

The 'introns-late' theory holds that spliceosomal introns have been added to genes during eukaryotic evolution. Few clear examples of recent intron gains have been well documented, but two such cases have now been reported, one with possible identification of the source of the intron.

You are what you eat: a gene transfer ratchet could account for bacterial genes in eukaryotic nuclear genomes.

Recent phylogenetic analyses reveal that many eukaryotic nuclear genes whose prokaryotic ancestry can be pinned down are of bacterial origin. Among them are genes whose products function exclusively in cytosolic metabolism. The results are surprising: we had come to believe that the eukaryotic nuclear genome shares a most recent common ancestor with archaeal genomes, thus most of its gene should be 'archaeal' (loosely speaking). Some genes of bacterial origin were expected as the result of transfer from mitochondria, of course, but these were thought to be relatively few, and limited to producing proteins reimported into mitochondria. Here, I suggest that the presence of many bacterial genes with many kinds of functions should not be a surprise. The operation of a gene transfer ratchet would inevitably result in the replacement of nuclear genes of early eukaryotes by genes from the bacteria taken by them as food.

The nature of the universal ancestor and the evolution of the proteome.

The past year has seen several attempts to reconstruct the proteome of the universal ancestor of all life on the basis of comparisons of contempory genomes. However, increasing evidence for lateral gene transfer could mean that such attempts are based on an incorrect understanding of evolution.

Microbial genomes: dealing with diversity.

We have now complete genome sequences of several pairs of closely related prokaryotes (conspecific strains or congeneric species). Surprisingly, even strains of the same species can differ by as much as 20% in gene content. Conceptual and methodological approaches for dealing with such diversity are now being developed, and should transform microbial genomics.

Class I release factors in ciliates with variant genetic codes.

In eukaryotes with the universal genetic code a single class I release factor (eRF1) most probably recognizes all stop codons (UAA, UAG and UGA) and is essential for termination of nascent peptide synthesis. It is well established that stop codons have been reassigned to amino acid codons at least three times among ciliates. The codon specificities of ciliate eRF1s must have been modified to accommodate the variant codes. In this study we have amplified, cloned and sequenced eRF1 genes of two hypotrichous ciliates, Oxytricha trifallax (UAA and UAG for Gln) and Euplotes aediculatus (UGA for Cys). We also sequenced/identified three protist and two archaeal class I RF genes to enlarge the database of eRF1/aRF1s with the universal code. Extensive comparisons between universal code eRF1s and those of Oxytricha, Euplotes, and Tetrahymena which represent three lineages that acquired variant codes independently, provide important clues to identify stop codon-binding regions in eRF1. Domain 1 in the five ciliate eRF1s, particularly the TASNIKS heptapeptide and its adjacent region, differs significantly from domain 1 in universal code eRF1s. This observation suggests that domain 1 contains the codon recognition site, but that the mechanism of eRF1 codon recognition may be more complex than proposed by Nakamura et al. or Knight and Landweber.