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Functional analysiss of OASIS, a glia-specific ER stress sensax protein.

OASIS（一种神经胶质细胞特异性 ER 应激感觉蛋白）的功能分析。

基本信息

批准号：
18500298
负责人：
WANAKA Akio
金额：
$ 2.6万
依托单位：
Nara Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18500298/
关键词：
CREB ATF family Transcription factor ER stress Unfolded nrotein resnonse Astrocyte 細胞死 ATF ファミリーノックアウトマウス

项目摘要

We have identified a novel member of CREB/ATF family and named it OASIS. OASIS has a transmembrane domain in addition to a bZIP domain, reminiscent of another family member ATF6. Via this transmembrane domain, OASIS is localized to the endoplasmic reticulum (ER). Our previous study showed that OASIS is specifically expressed by astrocytes in the brain. Since ATF6 has been implicated in the stress responses, we assumed that OASIS is also involved in some kind of stress responses.In the present study, we first tested whether OASIS is liberated from ER membrane by proteolytic cleavage. We employed cultured astrocytes for this analysis, because OASIS is endogenously expressed in the cell. First if all, we applied ER stress to astrocytes by treating them with tunicamicin or thapsigargin. Under this condition, OASIS protein has smaller molecular weight than the control condition. This suggested that OASIS received regulated intramembrene proteolysis. When we knocked down the membrane-associated protease, Si protease, by siRNA, the cleavage of the OASIS protein was inhibited. In the case of 52 protease knockdown, the cleavage was also blocked. These result suggested that Si and S2 proteases are both involved in the liberation of OASIS N-terminal fragment. These two proteases are known to reside in the Golgi apparatus. So we checked subcellular localization of OASIS protein in the ER stress condition. Unlike the normal state, OASIS protein was translocated to the Golgi apparatus. Mutation of intraluminal domain of the OASIS protein did not affect the translocation to the Golgi apparatus, suggesting that Golgi translocation was promoted by the mechanisms other than the ordinary Golgi targeting signals, which ATF6 utilizes.

我们鉴定了CREB/ATF家族中的一个新成员，并将其命名为OASIS。除了bZIP结构域之外，OASIS还有一个跨膜结构域，这让人想起另一个家族成员ATF6。通过这个跨膜区，绿洲定位于内质网(ER)。我们之前的研究表明，OASIS是由大脑中的星形胶质细胞特异性表达的。由于ATF6参与了应激反应，我们推测OASIS也参与了某种应激反应。在本研究中，我们首先测试了OASIS是否通过蛋白水解性切割从ER膜上释放出来。我们使用培养的星形胶质细胞进行分析，因为OASIS在细胞中内源性表达。首先，我们对星形胶质细胞施加内质网应激，用衣康星或thapsigargin处理它们。在此条件下，绿洲蛋白的相对分子质量小于对照条件。这表明绿洲细胞受到了膜内蛋白降解的调控。当我们用siRNA敲除膜相关蛋白Si蛋白酶时，OASIS蛋白的切割被抑制。在52蛋白水解酶被敲除的情况下，切割也被阻断。这些结果表明，S_1和S_2酶都参与了绿洲N-末端片段的释放。这两种蛋白水解酶已知存在于高尔基体中。因此，我们检测了内质网应激条件下绿洲蛋白的亚细胞定位。与正常状态不同，绿洲蛋白被转移到高尔基体。OASIS蛋白腔内结构域的突变不影响高尔基体的易位，这表明高尔基体的易位是由ATF6利用的普通高尔基体靶向信号以外的机制促进的。