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To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2007.09.005 Byline: Yuval Rinkevich (a), Jacob Douek (b), Omer Haber (a), Baruch Rinkevich (b), Ram Reshef (a) Keywords: Innate immunity; Ascidians; Botrylloides; Development; Serine protease; Regeneration; EST library; Urochordates Abstract: The phenomenon of whole body regeneration (WBR) from minute soma fragments is a rare event in chordates, confined to the subfamily of botryllid ascidians and is poorly understood on the cellular and molecular levels. We assembled a list of 1326 ESTs from subtracted mRNA, at early stages of Botrylloides leachi WBR, and classified them into functional categories. Sixty-seven (15%) ESTs with roles in innate immunity signaling were classified into a broad functional group, a result supported by domain search and RT-PCR reactions. Gene ontology analysis for human homologous to the immune gene category, identified 22 significant entries, of which 'peptidase activity' and 'protease inhibitor activity', stood out as functioning during WBR. Analyzing expressions of serine protease Bl-TrSP, a representative candidate gene from the 'peptidase activity' subgroup, revealed low transcript levels in naA[macron]ve vasculature with upregulated expression during WBR. This was confirmed by in situ hybridization that further elucidated staining restricted to a circulating population of macrophage cells. Furthermore, Bl-TrSP was localized in regeneration niches within vasculature, in regenerating buds, and in buds, during blastogenesis. Functional inhibition of serine protease activity disrupts early remodeling processes of the vasculature microenvironment and hinders WBR. Comparison of genome-wide transcription of WBR with five other developmental processes in ascidians (including metamorphosis, budding and blastogenesis), revealed a broad conservation of immune signaling expressions, suggesting a ubiquitous route of harnessing immune-related genes within a broader range of tunicate developmental context. This, in turn, may have enabled the high diversity of life history traits represented by urochordate ascidians. Author Affiliation: (a) Faculty of Biology, Technion-Israel Institute of Technology, Haifa, Israel (b) Israel Oceanography and Limnological Research, National Institute of Oceanography, Tel Shikmona, Haifa 31080, Israel Article History: Received 9 April 2007; Revised 28 August 2007; Accepted 6 September 2007