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Phylogenetic relationships within the green algal phylum Chlorophyta have proven difficult to resolve. The core Chlorophyta include Chlorophyceae, Ulvophyceae, Trebouxiophyceae, Pedinophyceae and Chlorodendrophyceae, but the relationships among these classes remain unresolved and the monophyly of Ulvophyceae and Trebouxiophyceae are highly controversial. We analyzed a dataset of 101 green algal species and 73 protein-coding genes sampled from complete and partial chloroplast genomes, including six newly sequenced ulvophyte genomes (Blidingia minima NIES-1837, Ulothrix zonata, Halochlorococcum sp. NIES-1838, Scotinosphaera sp. NIES-154, Caulerpa brownii and Cephaleuros sp. HZ-2017). We applied the Tree Certainty (TC) score to quantify the level of incongruence between phylogenetic trees in chloroplast genomic datasets, and show that the conflicting phylogenetic trees of core Chlorophyta stem from the most GC-heterogeneous sites. With removing the most GC-heterogeneous sites, our chloroplast phylogenomic analyses using heterogeneous models consistently support monophyly of the Chlorophyceae and of the Trebouxiophyceae, but the Ulvophyceae was resolved as polyphyletic. Our analytical framework provides an efficient approach to reconstruct the optimal phylogenetic relationships by minimizing conflicting signals.

QUOKKA is a 40 m pinhole small-angle neutron scattering instrument in routine user operation at the OPAL research reactor at the Australian Nuclear Science and Technology Organisation. Operating with a neutron velocity selector enabling variable wavelength, QUOKKA has an adjustable collimation system providing source–sample distances of up to 20 m. Following the large-area sample position, a two-dimensional 1 m2 position-sensitive detector measures neutrons scattered from the sample over a secondary flight path of up to 20 m. Also offering incident beam polarization and analysis capability as well as lens focusing optics, QUOKKA has been designed as a general purpose SANS instrument to conduct research across a broad range of scientific disciplines, from structural biology to magnetism. As it has recently generated its first 100 publications through serving the needs of the domestic and international user communities, it is timely to detail a description of its as-built design, performance and operation as well as its scientific highlights. Scientific examples presented here reflect the Australian context, as do the industrial applications, many combined with innovative and unique sample environments.

The relatively low chemical stability of cytosine compared with other nucleobases is a key concern in origin-of-life scenarios, but the effect of pressure on the rate of hydrolysis of cytosine to uracil remains unknown. Through in situ NMR spectroscopy measurements, it has been determined that the half-life of cytosine at 373.15 K decreases from (18.0±0.7) days at ambient pressure (0.1 MPa) to (8.64±0.18) days at high pressure (200 MPa). This yields an activation volume for hydrolysis of (-11.8±0.5) cm3 mol-1 ; a decrease that is similar to the molar volume of water (18.0 cm3 mol-1 ) and consistent with a tetrahedral 3,3-hydroxyamine transition-state/intermediate species. Similar behaviour was also observed for cytidine. At both ambient and high pressures, the half-life of cytosine decreases significantly as the pH decreases from 7.0 to 6.0. These results provide scant support for the notion that RNA-based life forms originated in high-temperature, high-pressure, acidic environments.

Green plants comprise two main clades: the Streptophyta, which include charophyte green algae and the embryophytic land plants, and the Chlorophyta including a wide diversity of marine, freshwater, and terrestrial green algae. Establishing a robust phylogeny is important to provide an evolutionary framework for comparative and functional studies. During the last two decades our understanding of the evolution of green algae has profoundly changed, first by phylogenetic analyses of nuclear ribosomal sequence data (mainly 18S), and more recently by analyses of multi-gene and chloroplast genomic data. The phylogenetic relationships among the main streptophytan lineages have been extensively studied and are now relatively well resolved. Although a lot of progress has been made in the last few years, the phylogenetic relationships in the Chlorophyta are still less well established. Here we review how chloroplast genomic data have contributed to address relationships among the main chlorophytan lineages. We highlight recent progress and conflicts among different studies, and discuss future directions in chloroplast phylogenomics of green algae.

Two basic questions are considered that approach protein evolution from different directions; the problems arising from using Markov models for the deeper divergences, and then the origin of proteins themselves. The real problem for the first question (going backwards in time) is that at deeper phylogenies the Markov models of sequence evolution must lose information exponentially at deeper divergences, and several testable methods are suggested that should help resolve these deeper divergences. For the second question (coming forwards in time) a problem is that most models for the origin of protein synthesis do not give a role for the very earliest stages of the process. From our knowledge of the importance of replication accuracy in limiting the length of a coding molecule, a testable hypothesis is proposed. The length of the code, the code itself, and tRNAs would all have prior roles in increasing the accuracy of RNA replication; thus proteins would have been formed only after the tRNAs and the length of the triplet code are already formed. Both questions lead to testable predictions.

Passerines are the largest avian order, and the 6,000 species comprise more than half of all extant bird species. This successful radiation probably had its origin in the Australasian region, but dating this origin has been difficult due to a scarce fossil record and poor biogeographic assumptions. Many of New Zealand’s endemic passerines fall within the deeper branches of the passerine radiation, and a well resolved phylogeny for the modern New Zealand element in the deeper branches of the oscine lineage will help us understand both oscine and passerine biogeography. To this end we present complete mitochondrial genomes representing all families of New Zealand passerines in a phylogenetic framework of over 100 passerine species. Dating analyses of this robust phylogeny suggest Passeriformes originated in the early Paleocene, with the major lineages of oscines “escaping” from Australasia about 30 Ma, and radiating throughout the world during the Oligocene. This independently derived conclusion is consistent with the passerine fossil record.

Land plants are a natural group, and Charophyte algae are the closest lineages of land plants and have six morphologically diverged groups. The conjugating green algae (Zygnematales) are now suggested to be the extant sister group to land plants, providing the novel understanding for character evolution and early multicellular innovations in land plants. We review recent molecular phylogenetic work on the origin of land plants and discuss some future directions in phylogenomic analyses.

Aim We used endemic frogs to test the hypothesis that New Zealand was fullysubmerged during the Oligocene marine transgression (c.25–35 Ma, the Oligo-cene drowning) and that all the land biota subsequently arrived by dispersal –including the poorly dispersing frogs.Location New Zealand.Methods The complete mitochondrial genome of the New Zealand nativefrog Leiopelma hochstetteri was sequenced, assembled and analysed, and com-pared with that of L. archeyi.Results Using many phylogenetic analyses we examined the evolutionary rela-tionships of the two species of Leiopelma, both for the deepest divergencewithin New Zealand and relative to other frogs. Our results give estimates ofwell over 65 Ma for the first divergence of leiopelmatid frogs within NewZealand, and over 150 Ma for the divergence of Leiopelma from its closest rela-tive, the North American tailed frog Ascaphus truei.Main conclusions The identification of such a deep divergence among theextant Leiopelma species provides some of the strongest support yet for amixed-age model for the origin of New Zealand’s terrestrial biota, where someelements are of vicariant origin and others have arisen by long-distance dis-persal. This contradicts a full drowning model and supports the view that theremust have been some continuous land through the Oligocene.KeywordsDivergence times, frogs, Leiopelmatidae, long-range dispersal, New Zealand,Oligocene drowning, vicariance.INTRODUCTIONLeiopelma is a genus of frogs native to New Zealand thatconsists of four extant and three recently extinct species, aswell as Miocene fossils. It is an ancient lineage of frogs thatpossesses morphological features not found in most otherfrogs – such as the absence of vocal sacs, lack of toe pads,absence of external ear drums, and the presence of nine ver-tebrae instead of eight (Hay et al., 1995). Two of the species,Leiopelma hochstetteri Fitzinger, 1861 and Leiopelma archeyiTurbott, 1942, are found on the North Island in remnantmontane forests, while Leiopelma pakeka Bell, Daugherty &Hay, 1998 is found on Maud’s Island and Leiopelma hamil-toni McCullough, 1919 on Stephen’s Island, both smallislands in Cook Strait. The latter two species differphenotypically and genetically only marginally from oneanother (Holyoake et al., 2001) and together with L. archeyishow molecular divergences expected between related specieswithin a genus. Physically, L. hamiltoni and L. archeyi haveidentical bone shape, differing only in size. However, L.hochstetteri is reasonably distinct from the other three speciesby being more aquatic, having partially webbed toes (Gibbs,2006, p. 154), lacking parental care, and having some differ-ences in bone shape. All the New Zealand frogs show similarmorphologies and traditionally all have been regarded as arelated group of species within the limits of a single genus.It has been suggested (see Nielson et al., 2006) that theclosest relative of Leiopelma is Ascaphus truei (hereafter Asca-phus), also an ancient lineage of frogs, now found along thewest coast of North America from British Columbia to

We need to estimate protein tertiary structure, as well as using primary sequences, in order to further our understanding of protein evolution and evolutionary processes in general.

A hypothesis is presented that the lipids of Archaea are an adaptation to a high temperature origin, possibly as an adaptation to higher sea temperatures during an early bombardment? This is separate from the issue that Eukaryotes arose before Prokaryotes (Akaryotes on our terminology). Archaeal lipids are basically isoprenoid compounds. All three domains of life have isoprenoids, but only Archaea have them as lipids are generally isoprenoid derivates, and are quite variable, and are ether liked (not ester linked). They may also be continuous, or split in the middle. Several methods of studying the temperatures at which membranes can be formed are discussed, the situation is ripe for experimental study. The ideas put forward here are all testable. A few authors make two clades of Archaea, but these are based on Markov models that are known to mislead at deeper divergences.

Early evolutionary thinkers proposed relatively simple models to describe processes of evolution, and these are the basis of evolutionary models still used today. Recent research has since shown that evolutionary relationships among plants can be complex and difficult to reconstruct even from molecular data. In plants there is a continuum of processes, ranging from reticulate relationships within a sexually reproducing population, incomplete lineage sorting and hybridization between recently diverged species, allopolyploidy between more distantly related species, to symbioses and endosymbiosis. These aspects of plant biology can create practical problems for interpreting bifurcating gene trees and identifying species. The promise of “omics” is that it will provide data and analyses to improve our understanding of the nature of species and their phylogenetic relationships. We highlight the importance of distinguishing evolutionary processes and evolutionary models, and stress that improving the un...

The Last Eukaryote Common Ancestor (LECA) appears to have the genetics required for meiosis, mitosis, nucleus and nuclear substructures, an exon/intron gene structure, spliceosomes, many centres of DNA replication, etc. (and including mitochondria). Most of these features are not generally explained by models for the origin of the Eukaryotic cell based on the fusion of an Archeon and a Bacterium. We find that the term 'prokaryote' is ambiguous and the non-phylogenetic term akaryote should be used in its place because we do not yet know the direction of evolution between eukaryotes and akaryotes. We use the term 'protoeukaryote' for the hypothetical stem group ancestral eukaryote that took up a bacterium as an endosymbiont that formed the mitochondrion. It is easier to make detailed models with a eukaryote to an akaryote transition, rather than vice versa. So we really are at a phylogenetic impasse in not being confident about the direction of change between eukaryotes and akaryotes.

It is not really helpful to consider modern environmental epigenetics as neo-Lamarckian; and there is no evidence that Lamarck considered the idea original to himself. We must all keep learning about inheritance, but attributing modern ideas to early researchers is not helpful, and can be misleading.

The origin of land plants is a fundamental topic in plant evolutionary biology. Despite the crucial importance for knowing the closest lineages of land plants, this question has not been fully answered yet. Using recently available nuclear sequences from streptophyte algae, the multispecies coalescent model produces a congruent phylogeny that is robust to different data sets, in contrast to the conflicting phylogenies produced by the concatenation method. Using phylogenomic data and the coalescent model, in this opinion article we postulate that the Zygnematales are the closest lineages of land plants. We suggest that the coalescent model can accommodate gene tree heterogeneity in deep-level phylogenies and can be potentially used to resolve other deep species phylogenies.

Vaults are very large oligomeric ribonucleoproteins conserved among a variety of species. The rat vault 3D structure shows an ovoid oligomeric particle, consisting of 78 major vault protein monomers, each of approximately 861 amino acids. Vaults are probably the largest ribonucleoprotein structures in eukaryote cells, being approximately 70 nm in length with a diameter of 40 nm--the size of three ribosomes and with a lumen capacity of 50 million Å(3). We use both protein sequences and inferred ancestral sequences for in silico virtual resurrection of tertiary and quaternary structures to search for vaults in a wide variety of eukaryotes. We find that the vault's phylogenetic distribution is widespread in eukaryotes, but is apparently absent in some notable model organisms. Our conclusion from the distribution of vaults is that they were present in the last eukaryote common ancestor but they have apparently been lost from a number of groups including fungi, insects, and probably plants. Our approach of inferring ancestral 3D and quaternary structures is expected to be useful generally.

We examine the advantages of going beyond sequence similarity and use both protein three-dimensional (3D) structure prediction and then quaternary structure (docking) of inferred 3D structures to help evaluate whether comparable sequences can fold into homologous structures with sufficient lateral associations for quaternary structure formation. Our test case is the major vault protein (MVP) that oligomerizes in multiple copies to form barrel-like vault particles and is relatively widespread among eukaryotes. We used the iterative threading assembly refinement server (I-TASSER) to predict whether putative MVP sequences identified by BLASTp and PSI Basic Local Alignment Search Tool are structurally similar to the experimentally determined rodent MVP tertiary structures. Then two identical predicted quaternary structures from I-TASSER are analyzed by RosettaDock to test whether a pair-wise association occurs, and hence whether the oligomeric vault complex is likely to form for a given MVP sequence. Positive controls for the method are the experimentally determined rat (Rattus norvegicus) vault X-ray crystal structure and the purple sea urchin (Strongylocentrotus purpuratus) MVP sequence that forms experimentally observed vaults. These and two kinetoplast MVP structural homologs were predicted with high confidence value, and RosettaDock predicted that these MVP sequences would dock laterally and therefore could form oligomeric vaults. As the negative control, I-TASSER did not predict an MVP-like structure from a randomized rat MVP sequence, even when constrained to the rat MVP crystal structure (PDB:2ZUO), thus further validating the method. The protocol identified six putative homologous MVP sequences in the heterobolosean Naegleria gruberi within the excavate kingdom. Two of these sequences are predicted to be structurally similar to rat MVP, despite being in excess of 300 residues shorter. The method can be used generally to help test predictions of homology via structural analysis.

In a recent article published in these pages, Bowman and colleagues propose that the ribosome represents a challenge to the RNA world model, a long-standing framework to explain the origin of DNA and genetically encoded proteins from a hypothetical RNA-based system. Specifically, they outline a scenario for the emergence and subsequent coevolution of the peptidyl transferase centre (PTC) of the ribosome with non-templated peptide products of this RNA through chemical evolution. They also propose that the PTC would have predated the emergence of enzymatic RNA replication, and that this in turn indicates that the RNA world never existed. We and others have previously incorporated non-templated peptide production as an early stage in the evolution of protein synthesis, which we would count as a chemical process, in agreement with Bowman and colleagues' model. However, their model raises an important question: to what extent could early protein synthesis and its products have evolved in the absence of Darwinian processes? We argue that evolution of the early ribosome requires Darwinian evolution, and that, while chemical evolution could give rise to peptidyl transferase activity, it is insufficient for subsequent improvement of a proto-PTC, or for ongoing coevolution of the proto-PTC with its early non-templated peptide products. We conclude that it is difficult to preclude the involvement of replicative processes, themselves subject to Darwinian evolution, from the evolution of the PTC. Finally, Bowman et al. call into question current models for the RNA to protein transition. We show that the difficulty that Bowman et al. have with this scenario is down to a misreading of our previous work.

The green algal phylum Chlorophyta has six diverse classes, but the phylogenetic relationship of the classes within Chlorophyta remains uncertain. In order to better understand the ancient Chlorophyta evolution, we have applied a site pattern sorting method to study compositional heterogeneity and the model fit in the green algal chloroplast genomic data. We show that the fastest-evolving sites are significantly correlated with among-site compositional heterogeneity and these sites have a much poorer fit to the evolutionary model. Our phylogenomic analyses suggest that the class Chlorophyceae is a monophyletic group and the classes Ulvophyceae, Trebouxiophyceae and Prasinophyceae are non-monophyletic groups. Our proposed phylogenetic tree of Chlorophyta will offer new insights to investigate ancient green algae evolution and our analytical framework will provide a useful approach for evaluating and mitigating the potential errors of phylogenomic inferences.

At macroevolutionary time scales, and for a constant mutation rate, there is an expected linear relationship between time and the number of inferred neutral mutations (the "molecular clock"). However, at shorter time scales a number of recent studies have observed an apparent acceleration in the rate of molecular evolution. We study this apparent acceleration under a Jukes-Cantor model applied to a randomly mating population, and show that, under the model, it arises as a consequence of ignoring short term effects due to existing diversity within the population. The acceleration can be accounted for by adding the correction term h_0e^{-4mu*t/3} to the usual Jukes-Cantor formula p(t)=(3/4)(1-e^{-4mu*t/3}), where h_0 is the expected heterozygosity in the population at time t=0. The true mutation rate mu may then be recovered, even if h_0 is not known, by estimating mu and h_0 simultaneously using least squares. Rate estimates made without the correction term (that is, incorrectly assuming the population to be homogeneous) will result in a divergent rate curve of the form mu_{div}=mu+C/t, so that the mutation rate appears to approach infinity as the time scale approaches zero. While our quantitative results apply only to the Jukes-Cantor model, it is reasonable to suppose that the qualitative picture that emerges also applies to more complex models. Our study therefore demonstrates the importance of properly accounting for any ancestral diversity, as it may otherwise play a dominant role in rate overestimation., LaTeX; 11 pages, 3 graphs. v3: Typoes corrected, including some in formulae. v2: Abstract, introduction and conclusions significantly rewritten to better reflect and explain the significance of the results

Resolving the closest relatives of Gnetales has been an enigmatic problem in seed plant phylogeny. The problem is known to be difficult because of the extent of divergence between this diverse group of gymnosperms and their closest phylogenetic relatives. Here, we investigate the evolutionary properties of conifer chloroplast DNA sequences. To improve taxon sampling of Cupressophyta (non-Pinaceae conifers), we report sequences from three new chloroplast (cp) genomes of Southern Hemisphere conifers. We have applied a site pattern sorting criterion to study compositional heterogeneity, heterotachy, and the fit of conifer chloroplast genome sequences to a general time reversible + G substitution model. We show that non-time reversible properties of aligned sequence positions in the chloroplast genomes of Gnetales mislead phylogenetic reconstruction of these seed plants. When 2,250 of the most varied sites in our concatenated alignment are excluded, phylogenetic analyses favor a close evolutionary relationship between the Gnetales and Pinaceae—the Gnepine hypothesis. Our analytical protocol provides a useful approach for evaluating the robustness of phylogenomic inferences. Our findings highlight the importance of goodness of fit between substitution model and data for understanding seed plant phylogeny.

Phylogenetics explores the continuum of shallower to deeper genetic divergences between taxa. Along this continuum increasing lengths of DNA sequence can be used to answer deeper and deeper questions about biological relationships. We use shorter, and then longer mitochondrial DNA sequences to address two aspects of pigeon evolution. Firstly, we examine the phylogenetic relationships of the eight genera within the South Pacific Ducula-Ptilinopus radiation, and examine how this radiation fits into pigeons generally. Within Ducula, taxa are closely related, whereas Ptilinopus is very diverse, and paraphyletic. One third of all pigeon species are within the Ducula-Ptilinopus radiation, however all are very similar ecologically. Secondly, we study the deeper phylogenetic question regarding the relationship of pigeons to other birds. To this end, we report the complete mitochondrial genome of Hemiphaganovaeseelandiae, a member of the Ducula-Ptilinopus radiation. We use this mitochondrial genome, along with additional sandgrouse (Pterocles namaqua) mitochondrial genes to assess various candidates for the closest relative of pigeons. Of parrots, shorebirds, and sandgrouse, we find highest support for the sandgrouse-pigeon grouping. Furthermore in these analyses the pigeon and sandgrouse group closer to the falcons than any other included taxon. The finding that pigeons and sandgrouse may be more closely related to falcons than to previous candidates such as shorebirds or parrots invites further investigation.

Second-generation sequencing technology has allowed a very large increase in sequencing throughput. In order to make use of this high throughput, we have developed a pipeline for sequencing and de novo assembly of multiple mitochondrial genomes without the costs of indexing. Simulation studies on a mixture of diverse animal mitochondrial genomes showed that mitochondrial genomes could be reassembled from a high coverage of short (35 nt) reads, such as those generated by a second-generation Illumina Genome Analyzer. We then assessed this experimentally with long-range polymerase chain reaction products from mitochondria of a human, a rat, a bird, a frog, an insect, and a mollusc. Comparison with reference genomes was used for deconvolution of the assembled contigs rather than for mapping of sequence reads. As proof of concept, we report the complete mollusc mitochondrial genome of an olive shell (Amalda northlandica). It has a very unusual putative control region, which contains a structure that would probably only be detectable by next-generation sequencing. The general approach has considerable potential, especially when combined with indexed sequencing of different groups of genomes.

Commonly used phylogenetic models assume a homogeneous process through time in all parts of the tree. However, it is known that these models can be too simplistic as they do not account for nonhomogeneous lineage-specific properties. In particular, it is now widely recognized that as constraints on sequences evolve, the proportion and positions of variable sites can vary between lineages causing heterotachy. The extent to which this model misspecification affects tree reconstruction is still unknown. Here, we evaluate the effect of changes in the proportions and positions of variable sites on model fit and tree estimation. We consider 5 current models of nucleotide sequence evolution in a Bayesian Markov chain Monte Carlo framework as well as maximum parsimony (MP). We show that for a tree with 4 lineages where 2 nonsister taxa undergo a change in the proportion of variable sites tree reconstruction under the best-fitting model, which is chosen using a relative test, often results in the wrong tree. In this case, we found that an absolute test of model fit is a better predictor of tree estimation accuracy. We also found further evidence that MP is not immune to heterotachy. In addition, we show that increased sampling of taxa that have undergone a change in proportion and positions of variable sites is critical for accurate tree reconstruction.

Eukaryote gene expression is mediated by a cascade of RNA functions that regulate, process, store, transport, and translate RNA transcripts. The RNA network that promotes this cascade depends on a large cohort of proteins that partner RNAs; thus, the modern RNA world of eukaryotes is really a ribonucleoprotein (RNP) world. Features of this "RNP infrastructure" can be related to the high cytosolic density of macromolecules and the large size of eukaryote cells. Because of the densely packed cytosol or nucleoplasm (with its severe restriction on diffusion of macromolecules), partitioning of the eukaryote cell into functionally specialized compartments is essential for efficiency. This necessitates the association of RNA and protein into large RNP complexes including ribosomes and spliceosomes. This is well illustrated by the ubiquitous spliceosome for which most components are conserved throughout eukaryotes and which interacts with other RNP-based machineries. The complexes involved in gene processing in modern eukaryotes have broad phylogenetic distributions suggesting that the common ancestor of extant eukaryotes had a fully evolved RNP network. Thus, the eukaryote genome may be uniquely informative about the transition from an earlier RNA genome world to the modern DNA genome world.

Eukaryotes express many functional non-protein-coding RNAs (ncRNAs) that participate in the processing and regulation of other RNA molecules. By focusing on connections between RNA-based processes, common patterns emerge that form a network-like RNA infrastructure. Owing to the intracellular movement of RNA during its processing (both between nuclear compartments and between the nucleus and cytoplasm), the RNA infrastructure contains both spatial and temporal connections. As research moves away from being protein-centric and focuses more on genomics, it is timely to explore these often 'hidden' aspects of the eukaryotic cell. The general and ancestral nature of most basic RNA-processing steps places a new focus on the generality of the spatial and temporal steps in RNA processing.

We report three developments toward resolving the challenge of the apparent basal polytomy of neoavian birds. First, we describe improved conditional down-weighting techniques to reduce noise relative to signal for deeper divergences and find increased agreement between data sets. Second, we present formulae for calculating the probabilities of finding predefined groupings in the optimal tree. Finally, we report a significant increase in data: nine new mitochondrial (mt) genomes (the dollarbird, New Zealand kingfisher, great potoo, Australian owlet-nightjar, white-tailed trogon, barn owl, a roadrunner [a ground cuckoo], New Zealand long-tailed cuckoo, and the peach-faced lovebird) and together they provide data for each of the six main groups of Neoaves proposed by Cracraft J (2001). We use his six main groups of modern birds as priors for evaluation of results. These include passerines, cuckoos, parrots, and three other groups termed "WoodKing" (woodpeckers/rollers/kingfishers), "SCA" (owls/potoos/owlet-nightjars/hummingbirds/swifts), and "Conglomerati." In general, the support is highly significant with just two exceptions, the owls move from the "SCA" group to the raptors, particularly accipitrids (buzzards/eagles) and the osprey, and the shorebirds may be an independent group from the rest of the "Conglomerati". Molecular dating mt genomes support a major diversification of at least 12 neoavian lineages in the Late Cretaceous. Our results form a basis for further testing with both nuclear-coding sequences and rare genomic changes.

Alternative splicing (AS) contributes to increased transcriptome and proteome diversity in various eukaryotic lineages. Previous studies showed low levels of conservation of alternatively spliced (cassette) exons within mammals and within dipterans. We report a strikingly different pattern in Caenorhabditis nematodes-more than 92% of cassette exons from Caenorhabditis elegans are conserved in Caenorhabditis briggsae and/or Caenorhabditis remanei. High levels of conservation extend to minor-form exons (present in a minority of transcripts) and are particularly pronounced for exons showing complex patterns of splicing. The functionality of the vast majority of cassette exons is underscored by various other features. We suggest that differences in conservation between lineages reflect differences in levels of functionality and further suggest that these differences are due to differences in intron length and the strength of consensus boundaries across lineages. Finally, we demonstrate an inverse relationship between AS and gene duplication, suggesting that the latter may be primarily responsible for the emergence of new functional transcripts in nematodes.

Accurate gene prediction in eukaryotes is a difficult and subtle problem. Here we point out a useful feature of expected distributions of spliceosomal intron lengths. Since introns are removed from transcripts prior to translation, intron lengths are not expected to respect coding frame, thus the number of genomic introns that are a multiple of three bases (‘3n introns’) should be similar to the number that are a multiple of three plus one bases (or plus two bases). Skewed predicted intron length distributions thus suggest systematic errors in intron prediction. For instance, a genome-wide excess of 3n introns suggests that many internal exonic sequences have been incorrectly called introns, whereas a deficit of 3n introns suggests that many 3n introns that lack stop codons have been mistaken for exonic sequence. A survey of genomic annotations for 29 diverse eukaryotic species showed that skew in intron length distributions is a common problem. We discuss several examples of skews in genome-wide intron length distributions that indicate systematic problems with gene prediction. We suggest that evaluation of length distributions of predicted introns is a fast and simple method for detecting a variety of possible systematic biases in gene prediction or even problems with genome assemblies, and discuss ways in which these insights could be incorporated into genome annotation protocols.

We consider the problem of finding the maximum likelihood rooted tree of three species under a molecular clock symmetric model of substitution of 2-state characters. For identically distributed rates per site this is probably the simplest phylogenetic estimation problem, and it is readily solved numerically. Analytic solutions, on the other hand, were obtained only recently by Yang [Complexity of the simplest phylogenetic estimation problem, Proc. Roy Soc. London Ser. B 267 (2000) 109–119].In this work we provide analytic solutions for any distribution of rates across sites, provided the moment generating function of the distribution is strictly increasing over the negative real numbers. This class of distributions includes, among others, identical rates across sites, as well as the Gamma, the uniform, and the inverse Gaussian distributions. Our work therefore generalizes Yang's solution and our derivation of the analytic solution is substantially simpler. We use the Hadamard conjugation to prove a general statement about the edge lengths of any neighboring pair of leaves in any phylogenetic tree (on three or more taxa). We then employ this relation, in conjunction with the convexity of an entropy-like function, to derive the analytic solution.

Although spliceosomal introns are present in all characterized eukaryotes, intron numbers vary dramatically, from only a handful in the entire genomes of some species to nearly 10 introns per gene on average in vertebrates. For all previously studied intron-rich species, significant fractions of intron positions are shared with other widely diverged eukaryotes, indicating that 1) large numbers of the introns date to much earlier stages of eukaryotic evolution and 2) these lineages have not passed through a very intron-poor stage since early eukaryotic evolution. By the same token, among species that have lost nearly all of their ancestral introns, no species is known to harbor large numbers of more recently gained introns. These observations are consistent with the notion that intron-dense genomes have arisen only once over the course of eukaryotic evolution. Here, we report an exception to this pattern, in the intron-rich diatom Thalassiosira pseudonana. Only 8.1% of studied T. pseudonana intron positions are conserved with any of a variety of divergent eukaryotic species. This implies that T. pseudonana has both 1) lost nearly all of the numerous introns present in the diatom-apicomplexan ancestor and 2) gained a large number of new introns since that time. In addition, that so few apparently inserted T. pseudonana introns match the positions of introns in other species implies that insertion of multiple introns into homologous genic sites in eukaryotic evolution is less common than previously estimated. These results suggest the possibility that intron-rich genomes may have arisen multiple times in evolution. These results also provide evidence that multiple intron insertion into the same site is rare, further supporting the notion that early eukaryotic ancestors were very intron rich.

International audience; Incomplete taxon sampling has been a major problem in resolving the early divergences in birds. Five new mitochondrial genomes are reported here (brush-turkey, lyrebird, suboscine flycatcher, turkey vulture, and a gull) and three break up long branches that tended to attract the distant reptilian outgroup. These long branches were to galliforms, and to oscine and suboscine passeriformes. Breaking these long branches leaves the root, as inferred by maximum likelihood and Bayesian phylogenetic analyses, between paleognaths and neognaths. This means that morphological, nuclear, and mitochondrial data are now in agreement on the position of the root of the avian tree and we can, move on to other questions. An overview is then given of the deepest divisions in the mitogenomic tree inferred from complete mitochondrial genomes. The strict monophyly of both the galloanseres and the passerines is strongly supported, leaving the deep six-way split within Neoaves as the next major question for which resolution is still lacking. Incomplete taxon sampling was also a problem for Neoaves, and although some resolution is now available there are still problems because current phylogenetic methods still fail to account for real features of DNA sequence evolution.