Induction of molecular chaperones by hyperosmotic stress in mouse inner medullary collecting duct cells.
Induction of molecular chaperones by hyperosmotic stress in mouse inner medullary collecting duct cells.
复制标题
小鼠内髓集合管细胞高渗应激诱导分子伴侣。
DOI：
10.1152/ajprenal.1997.273.1.f9
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发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Gullans,SR
中科院分区：
文献类型：
--
作者：
Rauchman,MI;Pullman,J;Gullans,SR
存在于肾内髓质的极端高渗条件使尿浓缩机制发挥作用。在这项研究中，我们评估是否应激相关的分子伴侣诱导高渗应激小鼠内髓集合管（MIMCD 3）细胞。暴露的细胞培养基中补充有100 mM NaCl 4或24小时导致热休克蛋白-72（HSP-72）（诱导型）的蛋白质印迹法增加。免疫细胞化学证实了HSP-72的增加，并表明，高渗应激导致HSP-72主要定位于核仁周围的核质和细胞质，亚细胞分布模式与热休克不同。使用变性蛋白质（酪蛋白）亲和柱与ATP洗脱，我们确定了一些假定的分子伴侣（46，60，78，和200 kDa），上调响应4小时的高渗NaCl处理。微测序鉴定这些蛋白质之一是线粒体伴侣蛋白mtHSP-70，HSP-70家族的成员，另一个类似于β-肌动蛋白。我们还发现高水平的HSP-72在长期适应高渗的细胞中，这表明即使在已知非干扰性有机渗透剂积累之后，仍然需要伴侣蛋白来维持某些细胞功能。这些结果表明，分子伴侣在肾脏髓质上皮细胞的适应高渗条件下，存在于体内的内部髓质中的重要作用。
The extreme hyperosmotic conditions that exist in the renal inner medulla enable the urinary concentrating mechanism to function. In this study, we evaluated whether stress-related molecular chaperones are induced in response to hyperosmotic stress in mouse inner medullary collecting duct (mIMCD3) cells. Exposure of cells to medium supplemented with 100 mM NaCl for 4 or 24 h resulted in an increase in heat shock protein-72 (HSP-72) (inducible form) by Western blot. Immunocytochemistry confirmed the increase of HSP-72 and showed that hyperosmotic stress resulted in a localization of HSP-72 predominantly to the nucleoplasm that surrounds the nucleoli and to the cytoplasm, a subcellular distribution pattern different from that seen with heat shock. Using a denatured protein (casein)-affinity column with ATP elution, we identified a number of putative molecular chaperones (46, 60, 78, and 200 kDa) that are upregulated in response to 4 h of hyperosmotic NaCl treatment. Microsequencing identified one of these proteins to be the mitochondrial chaperone mtHSP-70, a member of HSP-70 family, and another to be similar to beta-actin. We also found high levels of HSP-72 in cells chronically adapted to hypertonicity, indicating that chaperones are still required to maintain certain cellular functions even after nonperturbing organic osmolytes are known to accumulate. These results suggest an important role for molecular chaperones in the adaptation of renal medullary epithelial cells to the hyperosmotic conditions that exist in the inner medulla in vivo.