PARENT SESSION
GENE REGULATION AND FUNCTION IN THE REPRODUCTIVE AXIS
Kent
7:30 AM-10:00 AM

(369) ANALYSIS OF AN OOCYTE-SPECIFIC NOVEL GENE, oogenesin, IN THE MOUSE.

Minami, Naojiro¹, Miyamoto, Mazakazu¹, Ohashi, Akihiro¹, Aizawa, Akira², Kigami, Daisuke¹, Imai, Hiroshi¹, ¹ Laboratory of Reproductive Biology, Kyoto, Japan² Maebashi Institute of Animal Science, Maebashi, Japan

ABSTRACT- In the present study, we report the protein expression of a novel gene (named oogenesin) in mouse oocytes and embryos. The gene was first isolated based on its elevated expression in 2-cell embryos developed without oviductal tissue, in which embryos cannot develop beyond the 2-cell stage. The full length of the cDNA was constructed from a set of homologous clones obtained from the EST (Expressed Sequence Tag) database and a 5' RACE analysis. Molecular cloning of a near full-length cDNA revealed that the novel gene encodes a protein composed of 398 amino acids corresponding to a predicted molecular mass of 46 kDa with no significant homology to the sequences previously reported. A remarkable characteristic of the gene is that it contains a leucine zipper at positions 203-224. Northern blot analysis of RNA from multiple mouse tissues demonstrated that the gene expressed only in ovary as a single transcript of approx. 1.8 kbp. In situ hybridization using an RNA probe in sections of the adult ovary resulted in distinct signals in the oocyte and surrounding cumulus cells. However, an RT-PCR analysis of cDNA obtained from cumulus cells of ovulated oocytes demonstrated that the gene is not expressed in the cumulus cells. Expression analysis of a novel cDNA isolated from various stages of embryos revealed that c-1 expression abruptly decreased after the 2-cell stage and disappeared after the 4-cell stage. Immunohistochemical analysis of ovary sections using an antibody raised against a specific peptide sequence revealed that the protein also expressed specifically in oocytes. Western blot analysis using the same antibody also demonstrated that the gene expressed from oocytes to the 4-cell stage embryos at the predicted position of 46 kDa. An RT-PCR experiment using various stages of fetal ovary revealed that de novo expression of the novel gene starts at 15.5 dpc ovary. This time coincides with the start of meiotic cell cycle and of the expression of apoptosis related proteins Bax, suggesting that the novel gene may be involved in regulating the meiotic cell cycle and/or apoptosis in the oocytes.

KEY WORDS: oocyte, novel gene, mouse, 15.5 embryo