Hsp90 Co-localizes with Rab-GDI-1 and regulates agonist-induced amylase release in AR42J cells.
Hsp90 Co-localizes with Rab-GDI-1 and regulates agonist-induced amylase release in AR42J cells.
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Hsp90 与 Rab-GDI-1 共定位并调节 AR42J 细胞中激动剂诱导的淀粉酶释放。
DOI：
10.1159/000257429
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发表时间：
2009
期刊：
影响因子：
--
通讯作者：
Rafiq,Sarwish
中科院分区：
文献类型：
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作者：
Raffaniello,Robert;Fedorova,Daria;Ip,Dawn;Rafiq,Sarwish
Rab蛋白是通过分泌和内吞途径运输囊泡所需的小GTP酶。Rab GDP解离抑制剂（rab-GDI）通过维持Rab蛋白处于GDP结合构象来调节Rab蛋白的功能和定位。rab-GDI的两种同种型存在于大多数哺乳动物细胞中：GDI-1和GDI-2。最近已经证明，热休克蛋白90（Hsp 90）伴侣复合物调节Rab蛋白和Rab-GDI-1之间的相互作用。AR 42 J细胞系来源于大鼠胰腺外分泌肿瘤细胞，当用地塞米松（Dex）处理时，其形成腺泡样表型。本研究的目的是研究在存在或不存在Dex的情况下，rab-GDI亚型和Hsp 90在AR 42 J细胞中的表达。Rab-GDI：Hsp 90相互作用也被检查。通过免疫印迹在AR 42 J细胞中检测到两种rab-GDI同种型。在Dex处理的细胞中，定量免疫印迹显示rab-GDI-1表达增加了28%，尽管这种变化没有统计学意义。Rab-GDI-2水平未被Dex处理改变。大约21%的rab-GDI-1是膜相关的，而rab-GDI-2是专门的胞质。地塞米松处理不影响rab-GDI亚型的亚细胞分布。热休克蛋白90存在于AR 42 J细胞的胞浆和膜组分中，并与胞浆rab-GDI-1共免疫沉淀。此外，AR 42 J细胞膜的密度梯度离心显示，热休克蛋白90和rab-GDI-1共定位在低密度和高密度的膜组分，包括含淀粉酶的分泌颗粒。Hsp 90抑制剂格尔德霉素以剂量依赖性方式抑制CCK-8诱导的这些细胞的淀粉酶释放。我们的研究结果表明，作为AR 42 J细胞分化为腺泡样细胞，rab-GDI异构体的表达和定位没有显着改变。此外，我们的研究结果表明，热休克蛋白90调节激动剂诱导的外分泌细胞与rab-GDI-1相互作用。
Rab proteins are small GTPases required for vesicle trafficking through the secretory and endocytic pathways. Rab GDP-dissociation inhibitor (rab-GDI) regulates Rab protein function and localization by maintaining Rab proteins in the GDP-bound conformation. Two isoforms of rab-GDI are present in most mammalian cells: GDI-1 and GDI-2. It has recently been demonstrated that a Heat shock protein 90 (Hsp90) chaperone complex regulates the interactions between Rab proteins and Rab-GDI-1. The AR42J cell line is derived from rat pancreatic exocrine tumor cells and develops an acinar-like phenotype when treated with dexamethasone (Dex). The aim of the present study was to examine the expression of rab-GDI isoforms and Hsp90 in AR42J cells in the presence or absence of Dex. Rab-GDI:Hsp90 interactions were also examined. Both rab-GDI isoforms were detected in AR42J cells by immunoblotting. In Dex-treated cells, quantitative immunoblotting revealed that rab-GDI-1 expression increased by 28%, although this change was not statistically significant. Rab-GDI-2 levels were unaltered by Dex treatment. Approximately 21% rab-GDI-1 was membrane associated, whereas rab-GDI-2 was exclusively cytosolic. Dex treatment did not affect the subcellular distribution of rab-GDI isoforms. Hsp90 was present in the cytosolic and membrane fractions of AR42J cells and co-immunoprecipitated with cytosolic rab-GDI-1. Moreover, density gradient centrifugation of AR42J cell membranes revealed that Hsp90 and rab-GDI-1 co-localize on low- and high-density membrane fractions, including amylase-containing secretory granules. The Hsp90 inhibitor, geldanamycin, inhibited CCK-8-induced amylase release from these cells in a dose-dependent manner. Our results indicate that as AR42J cells differentiate into acinar-like cells, rab-GDI isoform expression and localization is not significantly altered. Moreover, our findings suggest that Hsp90 regulates agonist-induced secretion in exocrine cells by interacting with rab-GDI-1.