Kashlan Ossama B, Mueller Gunhild M, Qamar Mohammad Z, Poland Paul A, Ahner Annette, Rubenstein Ronald C, Hughey Rebecca P, Brodsky Jeffrey L, Kleyman Thomas R
Renal-Electrolyte Division, Department of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
J Biol Chem. 2007 Sep 21;282(38):28149-56. doi: 10.1074/jbc.M703409200. Epub 2007 Jul 30.
Integral membrane proteins are synthesized on the cytoplasmic face of the endoplasmic reticulum (ER). After being translocated or inserted into the ER, they fold and undergo post-translational modifications. Within the ER, proteins are also subjected to quality control checkpoints, during which misfolded proteins may be degraded by proteasomes via a process known as ER-associated degradation. Molecular chaperones, including the small heat shock protein alphaA-crystallin, have recently been shown to play a role in this process. We have now found that alphaA-crystallin is expressed in cultured mouse collecting duct cells, where apical Na(+) transport is mediated by epithelial Na(+) channels (ENaC). ENaC-mediated Na(+) currents in Xenopus oocytes were reduced by co-expression of alphaA-crystallin. This reduction in ENaC activity reflected a decrease in the number of channels expressed at the cell surface. Furthermore, we observed that the rate of ENaC delivery to the cell surface of Xenopus oocytes was significantly reduced by co-expression of alphaA-crystallin, whereas the rate of channel retrieval remained unchanged. We also observed that alphaA-crystallin and ENaC co-immunoprecipitate. These data are consistent with the hypothesis that small heat shock proteins recognize ENaC subunits at ER quality control checkpoints and can target ENaC subunits for ER-associated degradation.
整合膜蛋白在内质网（ER）的细胞质面上合成。在转运或插入到内质网后，它们会折叠并进行翻译后修饰。在内质网中，蛋白质还要经过质量控制检查点，在此期间，错误折叠的蛋白质可能会通过一种称为内质网相关降解的过程被蛋白酶体降解。分子伴侣，包括小热休克蛋白αA-晶状体蛋白，最近已被证明在这个过程中发挥作用。我们现在发现αA-晶状体蛋白在培养的小鼠集合管细胞中表达，在这些细胞中，顶端钠（Na⁺）转运由上皮钠通道（ENaC）介导。在非洲爪蟾卵母细胞中，共表达αA-晶状体蛋白会降低ENaC介导的钠电流。ENaC活性的这种降低反映了细胞表面表达的通道数量减少。此外，我们观察到，共表达αA-晶状体蛋白会显著降低非洲爪蟾卵母细胞ENaC转运到细胞表面的速率，而通道回收速率保持不变。我们还观察到αA-晶状体蛋白和ENaC能进行共免疫沉淀。这些数据与以下假设一致：小热休克蛋白在内质网质量控制检查点识别ENaC亚基，并可将ENaC亚基靶向进行内质网相关降解。
