title: Localization and enzymatic activity profiles of the proteases responsible for tachykinin-directed oocyte growth in the protochordate, Ciona intestinalis.
abstract: We previously substantiated that Ci-TK, a tachykinin of the protochordate, Ciona intestinalis (Ci), triggered oocyte growth from the vitellogenic stage (stage II) to the post-vitellogenic stage (stage III) via up-regulation of the gene expression and enzymatic activity of the proteases: cathepsin D, carboxypeptidase B1, and chymotrypsin. In the present study, we have elucidated the localization, gene expression and activation profile of these proteases. In situ hybridization showed that the Ci-cathepsin D mRNA was present exclusively in test cells of the stage II oocytes, whereas the Ci-carboxypeptidase B1 and Ci-chymotrypsin mRNAs were detected in follicular cells of the stage II and stage III oocytes. Double-immunostaining demonstrated that the immunoreactivity of Ci-cathepsin D was largely colocalized with that of the receptor of Ci-TK, Ci-TK-R, in test cells of the stage II oocytes. Ci-cathepsin D gene expression was detected at 2h after treatment with Ci-TK, and elevated for up to 5h, and then slightly decreased. Gene expression of Ci-carboxypeptidase B1 and Ci-chymotrypsin was observed at 5h after treatment with Ci-TK, and then decreased. The enzymatic activities of Ci-cathepsin D, Ci-carboxypeptidase B1, and Ci-chymotrypsin showed similar alterations with 1-h lags. These gene expression and protease activity profiles verified that Ci-cathepsin D is initially activated, which is followed by the activation of Ci-carboxypeptidase B1 and Ci-chymotrypsin. Collectively, the present data suggest that Ci-TK directly induces Ci-cahtepsin D in test cells expressing Ci-TK receptor, leading to the secondary activation of Ci-chymotrypsin and Ci-carboxypeptidase B1 in the follicle in the tachykininergic oocyte growth pathway.
authors.affiliation: Suntory Institute for Bioorganic Research, Wakayamadai, Mishima-gun, Osaka, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Aoyama
authors.forename: Tsuyoshi
authors.initials: T
authors.lastname: Kawada
authors.forename: Honoo
authors.initials: H
authors.lastname: Satake
mesh_terms: D000818:Animals; D002938:Ciona intestinalis / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D005260:Female; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D009866:Oogenesis / Q000502:physiology; D010447:Peptide Hydrolases / Q000378:metabolism*; D015320:Tachykinins / Q000378:metabolism*
chemical_list: D015320:Tachykinins; D010447:Peptide Hydrolases