Emmanuelle Lerat

From Gene Trees to Organismal Phylogeny in Prokaryotes:The Case of the γ-Proteobacteria

The rapid increase in published genomic sequences for bacteria presents the first opportunity to reconstruct evolutionary events on the scale of entire genomes. However, extensive lateral gene transfer (LGT) may thwart this goal by preventing the establishment of organismal relationships based on individual gene phylogenies. The group for which cases of LGT are most frequently documented and for which the greatest density of complete genome sequences is available is the γ-Proteobacteria, an ecologically diverse and ancient group including free-living species as well as pathogens and intracellular symbionts of plants and animals. We propose an approach to multigene phylogeny using complete genomes and apply it to the case of the γ-Proteobacteria. We first applied stringent criteria to identify a set of likely gene orthologs and then tested the compatibilities of the resulting protein alignments with several phylogenetic hypotheses. Our results demonstrate phylogenetic concordance among virtually all (203 of 205) of the selected gene families, with each of the exceptions consistent with a single LGT event. The concatenated sequences of the concordant families yield a fully resolved phylogeny. This topology also received strong support in analyses aimed at excluding effects of heterogeneity in nucleotide base composition across lineages. Our analysis indicates that single-copy orthologous genes are resistant to horizontal transfer, even in ancient bacterial groups subject to high rates of LGT. This gene set can be identified and used to yield robust hypotheses for organismal phylogenies, thus establishing a foundation for reconstructing the evolutionary transitions, such as gene transfer, that underlie diversity in genome content and organization.

Evolutionary Origins of Genomic Repertoires in Bacteria

Explaining the diversity of gene repertoires has been a major problem in modern evolutionary biology. In eukaryotes, this diversity is believed to result mainly from gene duplication and loss, but in prokaryotes, lateral gene transfer (LGT) can also contribute substantially to genome contents. To determine the histories of gene inventories, we conducted an exhaustive analysis of gene phylogenies for all gene families in a widely sampled group, the γ-Proteobacteria. We show that, although these bacterial genomes display striking differences in gene repertoires, most gene families having representatives in several species have congruent histories. Other than the few vast multigene families, gene duplication has contributed relatively little to the contents of these genomes; instead, LGT, over time, provides most of the diversity in genomic repertoires. Most such acquired genes are lost, but the majority of those that persist in genomes are transmitted strictly vertically. Although our analyses are limited to the γ-Proteobacteria, these results resolve a long-standing paradox—i.e., the ability to make robust phylogenetic inferences in light of substantial LGT.

Identifying repeats and transposable elements in sequenced genomes: how to find your way through the dense forest of programs

The production of genome sequences has led to another important advance in their annotation, which is closely linked to the exact determination of their content in terms of repeats, among which are transposable elements (TEs). The evolutionary implications and the presence of coding regions in some TEs can confuse gene annotation, and also hinder the process of genome assembly, making particularly crucial to be able to annotate and classify them correctly in genome sequences. This review is intended to provide an overview as comprehensive as possible of the automated methods currently used to annotate and classify TEs in sequenced genomes. Different categories of programs exist according to their methodology and the repeat, which they can identify. I describe here the main characteristics of the programs, their main goals and the difficulties they can entail. The drawbacks of the different methods are also highlighted to help biologists who are unfamiliar with algorithmic methods to understand this methodology better. Globally, using several different programs and carrying out a cross comparison of their results has the best chance of finding reliable results as any single program. However, this makes it essential to verify the results provided by each program independently. The ideal solution would be to test all programs against the same data set to obtain a true comparison of their actual performance.

Recognizing the pseudogenes in bacterial genomes

Pseudogenes are now known to be a regular feature of bacterial genomes and are found in particularly high numbers within the genomes of recently emerged bacterial pathogens. As most pseudogenes are recognized by sequence alignments, we use newly available genomic sequences to identify the pseudogenes in 11 genomes from 4 bacterial genera, each of which contains at least 1 human pathogen. The numbers of pseudogenes range from 27 in Staphylococcus aureus MW2 to 337 in Yersinia pestis CO92 (e.g. 1–8% of the annotated genes in the genome). Most pseudogenes are formed by small frameshifting indels, but because stop codons are A + T-rich, the two low-G + C Gram-positive taxa (Streptococcus and Staphylococcus) have relatively high fractions of pseudogenes generated by nonsense mutations when compared with more G + C-rich genomes. Over half of the pseudogenes are produced from genes whose original functions were annotated as ‘hypothetical’ or ‘unknown’; however, several broadly distributed genes involved in nucleotide processing, repair or replication have become pseudogenes in one of the sequenced Vibrio vulnificus genomes. Although many of our comparisons involved closely related strains with broadly overlapping gene inventories, each genome contains a largely unique set of pseudogenes, suggesting that pseudogenes are formed and eliminated relatively rapidly from most bacterial genomes.

Hypervariable and Highly Divergent Intron-Exon Organizations in the Chordate Oikopleura dioica

Oikopleura dioica is a pelagic tunicate with a very small genome and a very short life cycle. In order to investigate the intron–exon organizations in Oikopleura, we have isolated and characterized ribosomal protein EF-1α, Hox, and α-tubulin genes. Their intron positions have been compared with those of the same genes from various invertebrates and vertebrates, including four species with entirely sequenced genomes. Oikopleura genes, like Caenorhabditis genes, have introns at a large number of nonconserved positions, which must originate from late insertions or intron sliding of ancient insertions. Both species exhibit hypervariable intron–exon organization within their α-tubulin gene family. This is due to localization of most nonconserved intron positions in single members of this gene family. The hypervariability and divergence of intron positions in Oikopleura and Caenorhabditis may be related to the predominance of short introns, the processing of which is not very dependent upon the exonic environment compared to large introns. Also, both species have an undermethylated genome, and the control of methylation-induced point mutations imposes a control on exon size, at least in vertebrate genes. That introns placed at such variable positions in Oikopleura or C. elegans may serve a specific purpose is not easy to infer from our current knowledge and hypotheses on intron functions. We propose that new introns are retained in species with very short life cycles, because illegitimate exchanges including gene conversion are repressed. We also speculate that introns placed at gene-specific positions may contribute to suppressing these exchanges and thereby favor their own persistence.

Comparative analysis of transposable elements in the melanogaster subgroup sequenced genomes.

Transposable elements (TEs) are indwelling components of genomes, and their dynamics have been a driving force in genome evolution. Although we now have more information concerning their amounts and characteristics in various organisms, we still have little data from overall comparisons of their sequences in very closely-related species. While the Drosophila melanogaster genome has been extensively studied, we have only limited knowledge regarding the precise TE sequences in the genomes of the related species Drosophila simulans, Drosophila sechellia and Drosophila yakuba. In this study we analyzed the number and structure of TE copies in the sequenced genomes of these four species. Our findings show that, unexpectedly, the number of TE insertions in D. simulans is greater than that in D. melanogaster, but that most of the copies in D. simulans are degraded and in small fragments, as in D. sechellia and D. yakuba. This suggests that all three species were invaded by numerous TEs a long time ago, but have since regulated their activity, as the present TE copies are degraded, with very few full-length elements. In contrast, in D. melanogaster, a recent activation of TEs has resulted in a large number of almost-identical TE copies. We have detected variants of some TEs in D. simulans and D. sechellia, that are almost identical to the reference TE sequences in D. melanogaster, suggesting that D. melanogaster has recently been invaded by active TE variants from the other species. Our results indicate that the three species D. simulans, D. sechellia, and D. yakuba seem to be at a different stage of their TE life cycle when compared to D. melanogaster. Moreover, we show that D. melanogaster has been invaded by active TE variants for several TE families likely to come from D. simulans or the ancestor of D. simulans and D. sechellia. The numerous horizontal transfer events implied to explain these results could indicate introgression events between these species.

A call for benchmarking transposable element annotation methods.

DNA derived from transposable elements (TEs) constitutes large parts of the genomes of complex eukaryotes, with major impacts not only on genomic research but also on how organisms evolve and function. Although a variety of methods and tools have been developed to detect and annotate TEs, there are as yet no standard benchmarks—that is, no standard way to measure or compare their accuracy. This lack of accuracy assessment calls into question conclusions from a wide range of research that depends explicitly or implicitly on TE annotation. In the absence of standard benchmarks, toolmakers are impeded in improving their tools, annotators cannot properly assess which tools might best suit their needs, and downstream researchers cannot judge how accuracy limitations might impact their studies. We therefore propose that the TE research community create and adopt standard TE annotation benchmarks, and we call for other researchers to join the authors in making this long-overdue effort a success.

Codon usage by transposable elements and their host genes in five species.

Abstract. We compared the codon usage of sequences of transposable elements (TEs) with that of host genes from the species Drosophila melanogaster, Arabidopsis thaliana, Caenorhabditis elegans, Saccharomyces cerevisiae, and Homo sapiens. Factorial correspondence analysis showed that, regardless of the base composition of the genome, the TEs differed from the genes of their host species by their AT-richness. In all species, the percentage of A + T on the third codon position of the TEs was higher than that on the first codon position and lower than that in the noncoding DNA of the genomes. This indicates that the codon choice is not simply the outcome of mutational bias but is also subject to selection constraints. A tendency toward higher A + T on the third position than on the first position was also found in the host genes of A. thaliana, C. elegans, and S. cerevisiae but not in those of D. melanogaster and H. sapiens. This strongly suggests that the AT choice is a host-independent characteristic common to all TEs. The codon usage of TEs generally appeared to be different from the mean of the host genes. In the AT-rich genomes of Arabidopsis thaliana, Caenorhabditis elegans, and Saccharomyces cerevisiae, the codon usage bias of TEs was similar to that of weakly expressed genes. In the GC-rich genome of D. melanogaster, however, the bias in codon usage of the TEs clearly differed from that of weakly expressed genes. These findings suggest that selection acts on TEs and that TEs may display specific behavior within the host genomes.

Is the evolution of transposable elements modular

The evolution of transposable element structures can be analyzed in populations and species and by comparing the functional domains in the main classes of elements. We begin with a synthesis of what we know about the evolution of the mariner elements in the Drosophilidae family in terms of populations and species. We suggest that internal deletion does not occur at random, but appears to frequently occur between short internal repeats. We compared the functional domains of the DNA and/or amino acid sequences to detect similarities between the main classes of elements. This included the gag, reverse transcriptase, and envelope genes of retrotransposons and retroviruses, and the integrases of retrotransposons and retroviruses, and transposases of class II elements. We find that each domain can have its own evolutionary history. Thus, the evolution of transposable elements can be seen to be modular.

Reversing Gene Erosion – Reconstructing Ancestral Bacterial Genomes from Gene-Content and Order Data

In the last few years, it has become routine to use gene-order data to reconstruct phylogenies, both in terms of edge distances (parsimonious sequences of operations that transform one end point of the edge into the other) and in terms of genomes at internal nodes, on small, duplication-free genomes. Current gene-order methods break down, though, when the genomes contain more than a few hundred genes, possess high copy numbers of duplicated genes, or create edge lengths in the tree of over one hundred operations. We have constructed a series of heuristics that allow us to overcome these obstacles and reconstruct edges distances and genomes at internal nodes for groups of larger, more complex genomes. We present results from the analysis of a group of thirteen modern γ-proteobacteria, as well as from simulated datasets.