Post-sequencing functional studies on the pufferfish (Takifugu rubripes) genome

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1.Sequencing of 24,398 ESTs generated from 15 different adult and juvenile torafugu tissues resulted in 10,116 gene tags when compared with the genome data. The EST assemblies showed strong conservation with those of zebrafish, although significant differences remained between the two species.2.Muscle specific genes, myosin heavy chain (MYH) and tropomyosin (TPM), and genes encoding myogenic regulatory factors (MyoD family) such as MyoD and myogenin were analyzed in silico and in vivo. Comparison with mammalian counterparts revealed that although the number of the MyoD family genes was almost the same, torafugu had MYHs and TPMs about twice as many as mammalian counterparts, which suggests genome duplication in teleost.3.A total of 28 MYHs comprising 13 fast skeletal, 5 cardiac, 2 slow skeletal, 1 superfast, 2 smooth and 5 nonmuscle, were found in torafugu.4.Torafugu fast skeletal MYHs were distributed across five genomic regions, three of which contained MYH clusters. While human 6 fast skeletal MYHs formed one cluster, its syntenic region in torafugu was duplicated, although each locus contained a single MYH. These results suggest that fast skeletal MYHs evolved independently in teleosts and tetrapods after they had diverged from a common ancestor.5.Bacterial isolates from the torafugu skin were agglutinated by pufflectin, but could not adhere to the torafugu skin.6.pIgR was isolated as an transporter of the skin mucus IgM and, unlike mammals, torafugu pIgR was expressed not only in the intestine but also in the skin. A fragment of pIgR could bind to tetrameric IgM in the skin mucus.7.Marker genes of antigen presenting cells, CD11c, CD40 and MHC class IIα, and genes encoding costimulatory B7 molecules, B7-H1, B7-H3 and B7-H4, isolated from torafugu represent the first of these kinds from teleosts.