PHYLOBAS · Towards an improved animal phylogeny: Congruence between morphology and molecules in basal metazoans

Towards an improved animal phylogeny: Congruence between morphology and molecules in basal metazoans

Project context Over time, the architecture of the basal metazoan phylogenetic tree has been in a state of flux. From traditional phylogenies based on morphology to phylogenetic analyses of different genes the topology of the basal part of the animal tree has experienced drastic rearrangements. Nowadays, with phylogenomic approaches, interesting and robust relationships appear in the most basal steps of the animal tree. This project emerges as a necessity to increase the taxon sampling on the most basal nodes of the metazoan tree, especially the phylum Porifera, which in most phylogenomic studies counts no more than four representatives. Project objectives Our main objective is to generate transcriptomic data for five species of poriferans and to improve the robustness of the metazoan tree using phylogenomics. But the transcriptomic data generated by the project represents a huge amount of data that can and should be used for many more studies. Therefore, our specific objectives during the present project are as follows: 1. Transcriptome sequencing: our objective is to sequence the transcriptomes of five different sponge species (Sycon coactum, Petrosia ficiformis, Aphrocallistes vastus, Corticium candelabrum, and Spongilla lacustris) using NGS. Initially, it was intended to sequence with 454 platforms (Roche), but we finally used an Illumina HiSeq platform instead, which renders much more data (90 million reads (ca. 100 base pairs)) and also provides data to analyse the expression level of selected genes. To assemble the transcriptomes, we chose the software CLCbio workbench 4.0. Project results As a result of this new approach, we were able to establish new collaborations with different groups both in the United States and Europe. These techniques have never been applied before in sponges, and many different research groups expressed interest in establishing collaborations after the fellow presented the preliminary results during the VIII International Sponge Conference (September 2010, Girona, Spain). We contacted several groups to start collaborative projects. These collaborations have proved fruitful and provide new insights and pathways for this project. The specific sections in which these collaborators are currently working are provided below. 2. Comparative transcriptomics: using the assembled transcriptomes of the nine different species selected for the project, we set three different objectives to achieve: -2.1. Phylogenomic analyses (in collaboration with Dr Iñaki Ruiz-Trillo, IBE-CSIC): the main objective is to create a new database with molecular data to resolve the basal nodes of the animal tree of life. The transcriptomic data for our nine sponge species were uploaded in the database created by Drs Giribet and Dunn and implemented in an extensive holozoan data matrix provided by Dr Ruiz-Trillo. To complete this objective, we also used the available data on expressed sequence tags (ESTs) and whole genomes of poriferans, cnidarians, ctenophores and placozoans. We used several bilaterian outgroups, too. The preliminary results using the holozoan datamatrix suggests Porifera (with only our transcriptomic data analysed so far for Porifera) is a monophyletic group. -2.2. Developmental analyses (in collaboration with Dr Sally Leys, University of Alberta): we selected many gene targets of essential developmental processes (germline determination, cell adhesion, body structuring, signalling pathways, etc.) and the objective is to identify them in the transcriptomes of the selected species. Using the sequence information of the selected genes, we wanted to analyse the expression patterns of important genes in the germ lineage determination of sponges by using in-situ hybridisation techniques. -2.3. Differential expression analyses: genes involved in developmental processes were selected to analyse their differential expression during different life stages of the sponge individuals. The expression level of all the markers will be assessed by an experiment of RNA-seq. analysis performed using the contigs and raw sequences derived from the Illumina sequencing and the CLC bioinformatics workbench 4.0 program. Such an experiment will render the expression value RPKM (number of reads which map per kilobase of exon model per million mapped reads), which will be used to compare the different expression levels of the markers in the different stages of the sponge life cycle. 3. Phylogenetic analysis of the phylum Porifera with genes regularly used for phylogenetic analyses (the nuclear gene 18S, and the mitochondrial gene cytochrome oxidase I) to resolve the ancestral reproductive condition for sponges (gonochoric/hermaphroditic, oviparous/viviparous). We also established a new collaboration with Drs Malcolm Hill, Robert Thacker and Joseph Lopez in the framework of the project THE PORIFERA TREE OF LIFE - http://www.portol.org - supplying sequences of the housekeeping genes from the transcriptomes we sequenced in the lab, in order to resolve the phylogeny of the Porifera.

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