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Chaetognath phylogenomics: a protostome with deuterostome-like development

2006; Elsevier BV; Volume: 16; Issue: 15 Linguagem: Inglês

10.1016/j.cub.2006.07.016

1879-0445

Ferdinand Marlétaz, Elise Martin, Yannick Pérez, Daniel Papillon, Xavier Caubit, Christopher J. Lowe, Bob Freeman, Laurent Fasano, Carole Dossat, Patrick Wincker, Jean Weissenbach, Yannick Le Parco,

Parasite Biology and Host Interactions

Traditional textbook phylogeny splits bilaterians into protostomes and deuterostomes according to whether their mouth derives from the blastopore or not. This scheme has been largely confirmed by small subunit ribosomal RNA (SSU) molecular phylogeny. However, some phyla, such as the lophphorate phyla Phoronida and Brachiopoda as well as the Chaetognatha exhibit classical deuterostome embryological features such as formation of the mesoderm from the gut (enterocoely) and secondary opening of the mouth. However, their SSU sequences do not classify them as deuterostomes [1Adoutte A. Balavoine G. Lartillot N. Lespinet O. Prud'homme B. de Rosa R. The new animal phylogeny: reliability and implications.Proc. Natl. Acad. Sci. USA. 2000; 97: 4453-4456Crossref PubMed Scopus (424) Google Scholar]. For example, using SSU based phylogeny the lophophorates are now grouped with protostome trochozoans (Annelida and Mollusca) to form lophotrochozoans [1Adoutte A. Balavoine G. Lartillot N. Lespinet O. Prud'homme B. de Rosa R. The new animal phylogeny: reliability and implications.Proc. Natl. Acad. Sci. USA. 2000; 97: 4453-4456Crossref PubMed Scopus (424) Google Scholar, 2Halanych K.M. Bacheller J.D. Aguinaldo A.M. Liva S.M. Hillis D.M. Lake J.A. Evidence from 18S ribosomal DNA that the lophophorates are protostome animals.Science. 1995; 267: 1641-1643Crossref PubMed Scopus (451) Google Scholar]. But in the case of chaetognaths, a phylum of small marine predators, phylogenetic placement has been more problematic. Indeed, although they exhibit some protostome features, such as ventral nerve cords and circum-oesophageal fibers, SSU phylogeny first led to them being positioned wrongly as stem bilaterians because their fast evolving SSU resulted in a a long branch attraction artefact [3Telford M.J. Holland P.W. The phylogenetic affinities of the chaetognaths: a molecular analysis.Mol. Biol. Evol. 1993; 10: 660-676PubMed Google Scholar]. More recently, analysis of several mitochondrial markers showed affinities of chaetognaths to protostomes [4Papillon D. Perez Y. Caubit X. Le Parco Y. Identification of chaetognaths as protostomes is supported by the analysis of their mitochondrial genome.Mol. Biol. Evol. 2004; 21: 2122-2129Crossref PubMed Scopus (73) Google Scholar]. In order to address the discrepancy of morphological characters and molecular phylogeny, we attempted to resolve the phylogenetic position of chaetognaths using genomic data. Here, we clearly position chaetognaths among protostomes, likely as a sister-group of all other protostome phyla using a ribosomal protein dataset including hemichordate genomic data. We sequenced 11,526 expressed sequence tags (ESTs) from a juvenile cDNA library of the benthic chaetognath Spadella cephaloptera (Busch, 1851) (Supplemental data). BLAST comparisons of transcriptome similarities between chaetognaths and various species (specific databases: Drosophila, Lumbricus, Homo) or clades (composite databases: Deuterostomes, Lophotrochozoans, Ecdysozoans) showed conservation of a large set of genes within bilaterians (Table S1) as previously observed by Kortschak et al.[5Kortschak R.D. Samuel G. Saint R. Miller D.J. EST analysis of the cnidarian Acropora millepora reveals extensive gene loss and rapid sequence divergence in the model invertebrates.Curr. Biol. 2003; 13: 2190-2195Abstract Full Text Full Text PDF PubMed Scopus (287) Google Scholar]. Among these genes, ribosomal proteins were retrieved as valuable markers for phylogenomic analysis because of their conservation among eukaryotes and relative abundance in the EST collections of different phyla we used. We determined that BLAST searches provide significantly higher similarity between input and matched sequences when performed against a composite database that gathers all the available sequences for the various species belonging to a clade, rather than against a one-species database. As current problems in phylogenetic reconstruction are often related to long branch attraction [6Philippe H. Lartillot N. Brinkmann H. Multigene analyses of bilaterian animals corroborate the monophyly of Ecdysozoa, Lophotrochozoa, and Protostomia.Mol. Biol. Evol. 2005; 22: 1246-1253Crossref PubMed Scopus (436) Google Scholar], this composite database strategy was employed to shorten branches. For each validated monophyletic group (i.e. phylum), we retrieved the sequences of phylogenetic markers from a composite database (Supplemental data) and selected the less derived sequences (the sequences with the best blast score). This strategy allowed a reduction of branch lengths for fast evolving phyla, such as nematodes (Supplemental data). However, our method and data did not succeed in reducing the branch length of platyhelminthes, thus they were excluded from the main analysis. We built a dataset of 79 ribosomal proteins concatenated for 17 taxa, representing 14,558 raw positions (11,667 after Gblock filter). This dataset was validated for both composition homogeneity and saturation (Supplemental data), which are the strongest possible biases in phylogenomic analyses [7Jeffroy O. Brinkmann H. Delsuc F. Philippe H. Phylogenomics: the beginning of incongruence?.Trends Genet. 2006; 22: 225-231Abstract Full Text Full Text PDF PubMed Scopus (481) Google Scholar]. The topology obtained after analysis of this dataset supports the branching of chaetognaths with protostomes and the 'new view' of animal phylogeny (Figure 1) [1Adoutte A. Balavoine G. Lartillot N. Lespinet O. Prud'homme B. de Rosa R. The new animal phylogeny: reliability and implications.Proc. Natl. Acad. Sci. USA. 2000; 97: 4453-4456Crossref PubMed Scopus (424) Google Scholar]. Phylogeny inference was performed by maximum likelihood and Bayesian inference approaches — methods less sensitive to long branch attraction [6Philippe H. Lartillot N. Brinkmann H. Multigene analyses of bilaterian animals corroborate the monophyly of Ecdysozoa, Lophotrochozoa, and Protostomia.Mol. Biol. Evol. 2005; 22: 1246-1253Crossref PubMed Scopus (436) Google Scholar]. We recovered the three main animal clades — deuterostomes, ecdysozoans and lophotrochozoans — with significant bootstrap and posterior probabilities (Figure 1). The chaetognaths clustered strongly with protostomes (bootstrap 98% and posterior probabilty 1.0). Their position as a sistergroup to all other protostomes is supported by weak bootstrap (51%), but by strongest posterior probability (1.0). Likelihood-based tests show that this topology is the most likely one, but not significantly better than others (Supplemental data). Furthermore, we have discovered the Guanidinoacetate N-methyltransferase gene in the chaetognath EST collection. This gene is present in cnidarians and deuterostomes, but lost in protostomes, suggesting that chaetognaths are a stem protostome group (Supplemental data). Further genomic data such as ESTs or whole genome sequences in other protostomes (e.g. Lophophorates) would give further insights into protostome relationships. This should help to resolve the issue of whether chaetognaths should be grouped into an existing protostome clade, as proposed by Telford and colleagues [8Matus D.Q. Copley R.R. Dunn C.W. Hejnol A. Eccleston H. Halanych K.M. Martindale M.Q. Telford M. Broad taxon sampling and Gene Sampling Indicate that Chaetognaths are Sister to Lophotrochozoans.Curr. Biol. 2006; 16: R575-R576Abstract Full Text Full Text PDF PubMed Scopus (79) Google Scholar], or remain positioned as a sister group to all other protostomes as we propose here. In our analyses, we also found the surprising grouping of nematodes with tardigrades (bootstrap: 100%; posterior probability: 1.0) and of vertebrates with urochordates (bootstrap: 89%; posterior probability: 1.0), already observed in a previous phylogenomic study [9Delsuc F. Brinkmann H. Chourrout D. Philippe H. Tunicates and not cephalochordates are the closest living relatives of vertebrates.Nature. 2006; 439: 965-968Crossref PubMed Scopus (1215) Google Scholar]. Interestingly, the introduction of hemichordates into the analysis recovered their grouping with echinoderms in the Ambulacraria clade (bootstrap: 100%; posterior probability: 1) and strongly decreased the support (bootstrap: 57%; posterior probability: 0.71) for a cephalochordates plus echinoderms clade, as proposed in [9Delsuc F. Brinkmann H. Chourrout D. Philippe H. Tunicates and not cephalochordates are the closest living relatives of vertebrates.Nature. 2006; 439: 965-968Crossref PubMed Scopus (1215) Google Scholar]. So, the exact position of cephalochordates remains quite uncertain. Our results challenge some previous phylogenomic studies that argued either for a revival of the coelomata hypothesis [10Wolf Y.I. Rogozin I.B. Koonin E.V. Coelomata and not Ecdysozoa: evidence from genome-wide phylogenetic analysis.Genome Res. 2004; 14: 29-36Crossref PubMed Scopus (174) Google Scholar] or that there is insufficient phylogenetic signal for the resolution of bilaterian phylogeny [11Rokas A. Kruger D. Carroll S.B. Animal evolution and the molecular signature of radiations compressed in time.Science. 2005; 310: 1933-1938Crossref PubMed Scopus (261) Google Scholar]. Furthermore, the peculiar position of chaetognaths among protostomes validates previous mitochondrial analyses [4Papillon D. Perez Y. Caubit X. Le Parco Y. Identification of chaetognaths as protostomes is supported by the analysis of their mitochondrial genome.Mol. Biol. Evol. 2004; 21: 2122-2129Crossref PubMed Scopus (73) Google Scholar]. It confirms, on a genomic basis, that deuterostomy in an embryological sense is not a decisive character for the classification of animals. Some animals, like the chaetognaths, can be protostomes and yet show features of a deuterostome-like embryology (Figure 1) [12Valentine J.W. Cleavage patterns and the topology of the metazoan tree of life.Proc. Natl. Acad. Sci. USA. 1997; 94: 8001-8005Crossref PubMed Scopus (82) Google Scholar]. Nevertheless, the position of chaetognaths as a sistergroup of protostomes prompts us to propose that their development could be reminiscent of the bilaterian ancestor and testify that chaetognaths are a landmark phylum for addressing hypotheses about the origins of bilaterians. We thank Mathilde Paris, Bastien Boussau and François Bonneton from ENS Lyon for their helpful comments and suggestions, Gabriel Nève from Université de Provence for taking part in data management during the EST annotation process, Chris Gruber from Express genomics, and Eric Lander from the Broad Institute MIT for providing Saccoglossus ribosomal proteins. Download .pdf (.16 MB) Help with pdf files Document S1. Supplemental Experimental Procedures