Characterization of an ascidian DEAD-box gene, Ci-DEAD1: specific expression in the germ cells and its mRNA localization in the posterior-most blastomeres in early embryos

We isolated DEAD-box genes from three ascidian species (Ciona intestinalis, Ciona savignyi, and Halocynthia roretzi) by polymerase chain reaction meth ods. We obtained two types from each of C. infestinalis and C. savignyi, an d four types from H. roretzi. The first type (DEAD1) belonged to the vasa s ubfamily, the second type (DEAD2) to the PL10 subfamily, the third type (DE AD3) to the p68 subfamily, and the forth type (DEAD4) did not belong to any of the subfamilies. We further analyzed in detail the expression pattern o f C. intestinalis vasa-type gene (Ci-DEAD1) by in situ hybridization. In se ctions of the ovary and testis, the Ci-DEAD1-specific probe reacted intense ly to small germ cells, oogonium, and/or oocyte and spermatogonium and/or s permatocyte, respectively. In whole-mount specimens of juveniles this probe specifically reacted to the primordial germ cells in the gonad rudiment. T hese gonad-specific expressions were confirmed by reverse transcriptase pol ymerase chain reaction of RNA from various tissues. The transcript was pres ent in unfertilized eggs and in the central cytoplasm of blastomeres until the two-cell stage. During the second cleavage a part of the transcripts mo ved to the posterior region of embryos and, during early embryogenesis, was localized in the posterior-most blastomeres. In the tailbud, one or two hy bridization signals were detected in the caudal endodermal strand. Based on these observations, we propose precursors of primordial germ cells in asci dians.