Screening of a ligand for hypoxia-induced receptor by negative selection method

负选择法筛选缺氧诱导受体配体

• 批准号: 16601002
• 负责人: MIZUKAMI Yoichi
• 金额: $ 2.5万
• 依托单位: Yamaguchi University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2004
• 资助国家: 日本
• 起止时间: 2004 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16601002/
• 关键词: Estrogen; GPR30; membrane; がん; プロテオーム; リガンド探索

Recently, GPR30 was reported to be a novel estrogen receptor ; however, its intracellular localization has remained controversial. To investigate the intracellular localization of GPR30 in vivo, we produced four kinds of polyclonal antibodies for distinct epitopes on GPR30. Immunocytochemical observations using anti-GPR30 antibody and anti-FLAG antibody show that FLAG-GPR30 localizes to the plasma membrane 24 hr after transfection.Treatment with estrogen (17β-estradiol or E2) causes an elevation in the intracellular Ca^<2+> concentration ([Ca^<2+>]_i) within 10 sec in HeLa cells expressing FLAG-GPR30. In addition, E2 induces the translocation of GPR30 from the plasma membrane to the cytoplasm by 1 hr after stimulation. Immunohistochemical analysis shows that GPR30 exists on the cell surface of CA2 pyramidal neuronal cells. The images on transmission electron microscopy show that GPR30 is localized to a particular region on the associated with the plasma membranes of the pyramidal cells. These data indicate that GPR30,a transmembrane receptor for estrogen, is localized to the plasma membrane of CA2 pyramidal neuronal cells of the hippocampus in rat brain.

最近，据报道GPR30是一种新型的雌激素受体。但是，其细胞内定位仍然存在争议。为了研究GPR30在体内的细胞内定位，我们生产了四种在GPR30上不同表位的多克隆抗体。使用抗GPR30抗体和抗FLAG抗体的免疫细胞化学观察表明，转染后24小时的FLAG-GPR30定位于质膜24小时。用雌激素（17β-雌二醇或E2）对细胞内Ca^<2+>浓度的升高导致升高（表达式）hla hema hepration（ca^<2+> <2^<2^ca^<2+>]。此外，E2在刺激后诱导GPR30从质膜从质膜转换为细胞质。免疫组织化学分析表明，GPR30存在于Ca2锥体神经元细胞的细胞表面上。透射电子显微镜上的图像表明，GPR30位于与金字塔细胞的质膜相关的特定区域。这些数据表明，雌激素的跨膜受体GPR30位于大鼠脑中海马的Ca2锥体神经元细胞的质膜。

项目成果

Growth inhibitory effect of D-allose on human ovarian carcinoma cells in vitro

D-阿洛糖对人卵巢癌细胞的体外生长抑制作用

• 发表时间: 2005
• 期刊: Anticancer Res. 25(4)
• 作者: Hara;K.;Sui.L.
• 通讯作者: Sui.L.

Induction of cytochrome CYP4F3A in all-trans-retinoic acid-treated HL6O cells

全反式视黄酸处理的 HL6O 细胞中细胞色素 CYP4F3A 的诱导

• 发表时间: 2004
• 期刊: Biochem. Biophys. Res. Commun. 314
• 作者: Mizukami;Y
• 通讯作者: Y

Mechanism of the generation of autonomous activity of Ca2+/calmodulin-dependent protein kinase IV

• DOI: 10.1074/jbc.m406534200
• 发表时间: 2004-09-24
• 期刊: JOURNAL OF BIOLOGICAL CHEMISTRY
• 影响因子: 4.8
• 作者: Tokumitsu, H;Hatano, N;Kobayashi, R
• 通讯作者: Kobayashi, R

Regulation of Cardiomyocyte functions by distinct phosphoinositide 3-kinase, "Molecular Mechanisms of Heart diseases"(Edits Mizukami, Y., Ohkusa, T.)

独特的磷酸肌醇 3-激酶对心肌细胞功能的调节，“心脏病的分子机制”（Mizukami, Y.、Ohkusa, T. 编辑）

• 发表时间: 2005
• 作者: Aki;T.;Mizukami;Y.
• 通讯作者: Y.

Identification of active metabolite of ticlopidine from rat in vitro metabolites

大鼠体外代谢物中噻氯匹定活性代谢物的鉴定

• 发表时间: 2004
• 期刊: Brit. J. Pharmacol. 142(3)
• 作者: Yoneda;K
• 通讯作者: K