Functional analysis of oocyte-specific gene, c-1, which express during meiosis

在本研究中，我们报道了一种名为Oogenesin的新基因在小鼠卵母细胞和胚胎中的蛋白表达和定位。该基因首次在没有输卵管组织的2细胞胚胎中被分离出来，在2细胞胚胎中，胚胎不能发育超过2细胞阶段。对一个接近全长cDNA的分子克隆表明，该新基因编码一个由398个氨基酸组成的蛋白质，预测分子质量为46 kDa。该基因的一个显著特征是在203-224位含有亮氨酸拉链，在137-326位含有富含亮氨酸的结构域。在成年卵巢切片中使用RNA探针进行原位杂交，在卵泡的所有阶段的卵母细胞中产生不同的信号。从胚胎着床前不同阶段分离的一种新的cDNA表达分析表明，Oogenesin的表达在2细胞阶段后突然下降，在4细胞阶段后消失。利用兔多克隆抗体对合成肽进行亲和纯化的western blot分析显示，OOGENESIN蛋白的分子大小约为46 kDa，与基于OOGENESIN cDNA预测的大小一致。OOGENESIN主要在卵母细胞和1- 4细胞期胚胎中表达，而在桑葚胚/囊胚期胚胎中表达较弱。卵母细胞至1细胞期表达升高，至4细胞期表达降低。免疫组织化学检测到次级和窦卵泡卵母细胞信号强，原始和初级卵泡卵母细胞信号弱。所有信号均在卵泡卵母细胞细胞质中检测到。然而，对排卵卵母细胞和着床前胚胎的免疫组织化学分析显示，该蛋白定位于1细胞晚期和2细胞早期的细胞核中。这些结果以及该蛋白独特的结构表明，卵原蛋白作为一种新的母体效应基因，在小鼠胚胎的转录激活，特别是合子基因激活中具有重要的作用。少

In the present study, we report the protein expression and localization 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. 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. A remarkable characteristic of the gene is that it contains a leucine zipper at positions 203-224 and a leucine-rich domain at positions 137-326. In situ hybridization using an RNA probe in sections of the adult ovary resulted in distinct signals in the oocyte of all stages of follicles. Expression analysis of a novel cDNA isolated from various stages of preimplantation embryos revealed that Oogenesin expression abruptly decreased after the 2-cell stage and disappeared after the 4-cell stage.A western blot analy … More sis using an affinity-purified rabbit polyclonal antibody raised against a synthetic peptide revealed that the molecular size of OOGENESIN protein is approximately 46 kDa, which is consistent with the size predicted on the basis of the Oogenesin cDNA. OOGENESIN is expressed predominantly in oocytes and 1-to 4-cell stage embryos, whereas it is expressed only weakly in morula/blastocyst stage embryos. Increasing expression was observed from oocytes to the 1-cell stage and the expression decreased toward the 4-cell stage. Immunohistochemistry detected strong signals in oocytes of secondary and antral follicles, and weak signals in oocytes of primordial and primary follicles. All signals were detected in the cytoplasm of follicular oocytes. However, immunohistochemical analysis of ovulated oocytes and preimplantation embryos revealed that the protein localized in the nuclei at late 1-cell and early 2-cell stage. These results together with the unique structure of the protein suggest that Oogenesin has important roles in the transcriptional activation, especially in zygotic gene activation of mouse embryos as a novel maternal effect gene. Less