Derlin-1在上皮细胞钠通道(ENaC)泛素化降解中的调控作用

肾脏远端肾单位上皮细胞钠离子通道 (ENaC) 的主要功能是介导钠离子的跨膜转运，调节水钠平衡并稳定血压。ENaC功能失调会引起人体血压调节障碍。ENaC表达转运的调控有赖于ENaC蛋白合成和蛋白降解的动态平衡，但是目前人们对ENaC降解的分子机制还知之甚少。Derlin-1及其相关蛋白在泛素化蛋白酶体降解途径中起着至关重要的作用。本课题的研究目的在于探讨derlin-1在ENaC泛素化蛋白酶体降解中的调控机制。我们将利用肾脏集合管上皮细胞derlin-1转基因及基因敲除小鼠模型，在体观察derlin-1对ENaC降解的调控作用；也将利用小鼠肾脏集合管上皮细胞进行体外实验，从四个方面 (详见研究内容) 来深入阐明derlin-1在介导ENaC泛素化降解过程中的分子机制。本课题的完成将为研究机体对钠离子重吸收开拓新的思路，为发现高血压新的治疗靶点奠定新的理论基础。

The regulated activity of the distal nephron epithelial sodium channel (ENaC) is an important determinant of sodium balance, extracellular fluid volume, and blood pressure. The physiological significance of ENaC is illustrated most clearly by human genetic diseases in which channel mutations produce clinical defects in renal salt and water transport.The regulation of ENaC function really dependent on the balance of its Channel synthesis and channel degradation. However, our understanding of ENaC degradation in is quite incomplete. Recent studies show that derlin and its associated proteins play a critical role in ubiquitin mediated protein degradation. Our preliminary data suggest that the derlin complex effectively degrades ENaC in mouse cortical collecting duct (mCCD) cells. Our hypothesis is that the derlin complex is a key regulator of ubiquitin mediated ENaC degradation. The goal for this proposal is to identify components of the derlin complex that recognize and extract ENaC from the ER, and to identify the basis for their selective degradation of ENaC. The studies (in vivo) will be performed to examine whether derlin-1 complex participate in the regulation of ubiquitin-mediated epithelial sodium channel (ENaC) degradation using derlin-1 transgenic/knockout mice. We will test other four specific aims in mCCD cells (in vitro): (1) to define the role of derlin complex in ENaC degradation; (2) to examine the role of lysine-linkage of polyubiquitin chains in ENaC degradation; (3) to identify the processes mediating ENaC retro-translocation from the ER to the cytosol; and (4) to evaluate the ubiquitin E3 ligases that evoke ubiquitin-mediated ENaC degradation. Successful completion of this proposal will advance our understanding of molecular mechanism of ENaC regulation and provide a roadmap for identifying novel potential therapeutic targets to salt-sensitive hypertension.