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Cloning and expression pattern of a Smad3 homolog from the pearl oyster, Pinctada fucata.

Several transforming growth factor beta (TGFbeta) superfamily members have been identified from mollusks. The significant effects of TGFbeta signaling pathways on bone formation in vertebrates give clues to the signal transduction mechanism and how the shell of oysters is formed. However, what kinds of mediators are involved in the molluscan TGFbeta signaling pathways, and how they play their functions in mollusks has not been well explained due to a lack of genomic information and the failure to establish oyster cell lines. That is, if we knew the genome sequence we could search the TGFbeta superfamily members using the blast program (http://www.ncbi.nlm.nih.gov/), and if we established the molluscan cell line, we could transfect the gene of interest to the cell and detect its influence on expression levels of other genes, such as the matrix proteins. Therefore, to investigate whether there are similar TGFbeta pathways in mollusks, many important mediators should be identified. In this paper, we report a cDNA encoding a Smad3 homolog (designated Pf-Smad3) that was isolated from the pearl oyster, Pinctada fucata. Sequence alignment showed that Pf-Smad3 contains a DNA-binding MH1 domain and a Runx2/Cbfa1-binding MH2 domain, and shares an extremely high similarity with Smad3 proteins in vertebrates. However, Smad proteins in Drosophila and Caenorhabditis elegans are very different from other Smad3 proteins. Reverse transcription-polymerase chain reaction results indicated that Pf-Smad3 mRNA was expressed ubiquitously in adult Pin. fucata and was expressed at different levels at different developmental stages. In situ hybridization results showed that Pf-Smad3 mRNA was expressed mainly at the outer epithelial cells of the middle fold and the inner epithelial cells of the outer fold, especially around the gutter. These results suggested that Pf-Smad3 might take part in many physiological processes, including biomineralization, in oysters.