An increasing amount of Expressed Sequence Tag (EST) and genomic data predominantly for the cnidarians Acropora, Hydra and Nematostella, reveals that despite being one of the morphologically simplest multicellular animals, cnidarians possess a high genomic complexity. In order to contribute towards a broader coverage of this phylum, an EST project was performed to analyze the transcriptome of Hydractinia echinata. Moreover, transcriptional profiling experiments were carried out to characterize the i-cell population and the immune system of the hydroid. In this work a cDNA-library containing about 20,000 clones was constructed, which covers the entire life cycle of the organism and also represents some stress-induced conditions. After randomly sequencing almost 9,000 clones, EST characterization revealed a broad diversity of genes, with higher sequence similarity to vertebrates than to ecdysozoan invertebrates. Furthermore, a significant number of sequences hitherto unknown in metazoans were detected. The identification of unique Hydractinia sequences is consistent with the suggested high diversity and complexity of genes within the phylum. To store all the acquired information a database aimed at making the data widely available was created, which is accessible at www.mchips.org/hydractinia_echinata.html. To further characterize Hydractinia genes, a cDNA-microarray was constructed including the already sequenced ESTs as well as PCR-products from almost 5,000 un-sequenced cDNAs. Genes associated with the i-cell lineage were identified by the analysis of the gene expression profile of colonies depleted from their i-cells using mitomycin-C and colonies after the recovery from the treatment. Microarray normalized data ended up with 162 significant differentially expressed genes. Several growth and transcription factors as well as genes associated with undifferentiated cells were identified including; BMPs, Bzip/Mafl and CnPL10. In addition, i-cell depleted organisms exhibited an activation of genes involved in detoxification and wound healing activities. These genes are good candidates to define the i-cell population of Hydractinia. Genes associated with the immune system of Hydractinia were identified by the analysis of the expression profile of organisms having a LPS mimicked Gram-negative bacterial infection as well as an allogeneic reaction. 245 candidate genes with a significantly different expression level were identified. Genes associated with an LPS response encode for e.g. HSP70, lipocalin-like proteins, serine protease inhibitors, proteins with TSR domains and lectins. In the case of allorecognition, a probable whole genome response with up-and down regulation of hundreds of genes was observed; demonstrating a complex process. Some of the identified genes encode for e.g. minicollagens, transcriptional and growth factors, proteins with a protective function against oxygen metabolites or with potent inflammatory and neurotoxicity effects. Gene expression pattern analysis provided insights towards the function of many genes which are still unknown. In the case of genes with a known functional annotation, the microarray experiments either corroborated their characterization or defined an alternative one for Hydractinia. This project is the first high-throughput effort aimed to identify and characterize the transcriptome of the colonial marine hydroid Hydractinia echinata. The combination of the EST dataset, database and the microarray, provides a solid platform to promote and facilitate molecular research not only in Hydractinia but also in other cnidarians
