title: cDNA cloning, localization, and candidate binding partners of acid-extractable vitelline-coat protein Ci-v-Themis-like in the ascidian Ciona intestinalis.
abstract: Ascidians are hermaphrodites, although several ascidian species show self-sterility because of the occurrence of a self/nonself-recognition system called the self-incompatibility system. We previously reported that two pairs of sperm polycystin 1-like receptors, s-Themis-A and s-Themis-B, and egg fibrinogen-like ligands, v-Themis-A and v-Themis-B, are responsible for self-incompatibility in the ascidian Ciona intestinalis. Our previous results showed that v-Themis-A and v-Themis-B were hardly extracted from the vitelline coat (VC) by acid treatment, which is not in accordance with a report that an acid-extractable VC factor has the ability to distinguish self- from nonself-sperm. These results led us to explore a novel factor from acid-extractable VC proteins that could be involved in self-incompatibility. Here, we report cDNA cloning, expression, and localization of Ci-v-Themis-like, a major acid-extractable VC protein. This protein has a fibrinogen-like domain, as do v-Themis-A and v-Themis-B, but it showed no polymorphisms. Phylogenic analysis suggested that Ci-v-Themis-like is an ancestral protein of v-Themis-A and v-Themis-B. Whole mount in situ hybridization revealed that Ci-v-Themis-like mRNA is expressed in the ovary and testis. Western blotting and immunocytochemistry showed the occurrence of Ci-v-Themis-like in developing oocytes and on the VC of mature eggs. Yeast two-hybrid screenings using testis and ovary libraries revealed candidate interacting proteins; among these candidates, we succeeded in identifying several testis-specific proteins, including sperm proteases and coiled-coil-domain-containing proteins. The results suggest that Ci-v-Themis-like and its binding partners are involved in sperm binding to the VC prior to the allorecognition process during C. intestinalis fertilization.
authors.affiliation: Sugashima Marine Biological Laboratory, Graduate School of Science, Nagoya University, Sugashima, Toba, Japan.
authors.forename: Kei
authors.initials: K
authors.lastname: Otsuka
authors.forename: Lixy
authors.initials: L
authors.lastname: Yamada
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Sawada
mesh_terms: D000818:Animals; D002938:Ciona intestinalis / Q000235:genetics* / Q000378:metabolism; D004527:Egg Proteins / Q000096:biosynthesis / Q000378:metabolism*; D005260:Female; D007246:Infertility / Q000235:genetics / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism; D008297:Male; D010053:Ovary / Q000378:metabolism; D010802:Phylogeny; D017434:Protein Structure, Tertiary / Q000235:genetics; D057945:Self-Fertilization / Q000235:genetics / Q000502:physiology*; D013737:Testis / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D014561:Urochordata / Q000235:genetics* / Q000378:metabolism; D014817:Vitelline Membrane / Q000378:metabolism; D047048:Vitellins / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism
chemical_list: D004527:Egg Proteins; D047908:Intracellular Signaling Peptides and Proteins; D047048:Vitellins