Ovarian expression and localization of a vitellogenin receptor with eight ligand binding repeats in the cutthroat trout (Oncorhynchus clarki)
Ovarian expression and localization of a vitellogenin receptor with eight ligand binding repeats in the cutthroat trout (Oncorhynchus clarki)
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DOI：
10.1016/j.cbpb.2013.07.005
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发表时间：
2013-09-01
影响因子：
2.2
通讯作者：
Hiramatsu, Naoshi
中科院分区：
文献类型：
--
作者：
Mizuta, Hiroko;Luo, Wenshu;Hiramatsu, Naoshi
从切喉鳟鱼 Oncorhynchus clarki 的卵巢中克隆了编码具有 8 个配体结合重复序列 (vtgr) 的卵黄蛋白原受体的 cDNA。原位杂交和定量PCR分析表明vtgr mRNA的主要表达位点是卵母细胞。在核周期卵母细胞中强烈检测到表达，随着卵母细胞生长逐渐减少，并且在卵黄发生卵母细胞中几乎检测不到。针对重组 Vtgr 蛋白产生兔抗体 (a-Vtgr)，以便免疫学检测和定位卵泡内的 Vtgr。使用 a-Vtgr 进行蛋白质印迹检测，在卵巢制剂中检测到一条粗体条带，其表观质量类似于 95-105 kDa，配体印迹中也与库页岛哲罗鱼、哲罗鱼、卵黄原蛋白结合。使用a-Vtgr的免疫组织化学显示，Vtgr均匀分布在核周期卵母细胞的整个卵质中，随后易位到脂滴期卵母细胞的外围，并在卵黄发生期间定位于卵膜。我们首次对割喉鳟鱼的转录和翻译水平上的 Vtgr 进行了表征，我们的结果表明该受体参与了该物种卵母细胞对 Vtg 的摄取。 (C) 2013 Elsevier Inc. 保留所有权利。
A cDNA encoding a vitellogenin receptor with 8 ligand binding repeats (vtgr) was cloned from ovaries of the cutthroat trout, Oncorhynchus clarki. In situ hybridization and quantitative PCR analyses revealed that the main site of vtgr mRNA expression was the oocytes. Expression was strongly detected in perinucleous stage oocytes, gradually decreased as oocytes grew, and became hardly detectable in vitellogenic oocytes. A rabbit antibody (a-Vtgr) was raised against a recombinant Vtgr protein in order to immunologically detect and localize Vtgr within the ovarian follicles. Western blotting using a-Vtgr detected a bold band with an apparent mass of similar to 95-105 kDa in an ovarian preparation that also bound Sakhalin taimen, Hucho perryi, vitellogenin in ligand blots. Immunohistochemistry using a-Vtgr revealed that the Vtgr was uniformly distributed throughout the ooplasm of perinucleolus stage oocytes, subsequently translocated to the periphery of lipid droplet stage oocytes, and became localized to the oolemma during vitellogenesis. We provide the first characterization of Vtgr at both the transcriptional and the translational levels in the cutthroat trout, and our results suggest that this receptor is involved in uptake of Vtg by oocytes of this species. (C) 2013 Elsevier Inc. All rights reserved.