Identification and functional characterization of an N-terminal oligomerization domain for polycystin-2.
Identification and functional characterization of an N-terminal oligomerization domain for polycystin-2.
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DOI：
10.1074/jbc.m803834200
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发表时间：
2008-10-17
期刊：
影响因子：
--
通讯作者：
Ong AC
中科院分区：
文献类型：
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作者：
Feng S;Okenka GM;Bai CX;Streets AJ;Newby LJ;DeChant BT;Tsiokas L;Obara T;Ong AC
常染色体显性遗传性多囊肾病（ADPKD）是肾衰竭最常见的遗传性原因，由PKD 1（85%）或PKD 2（15%）突变引起。PKD 2蛋白，多囊蛋白-2（PC 2或TRPP 2），是瞬时受体电位（TRP）超家族的成员，并作为非选择性钙通道发挥作用。已发现PC 2在天然组织中形成寡聚体，这表明它可能与其他亚基形成功能性同四聚体或异四聚体，类似于其他TRP通道。我们的实验出乎意料地发现，缺乏已知的C-末端二聚化结构域的PC 2突变蛋白仍然能够形成寡聚体和共免疫沉淀全长PC 2，这意味着可能存在的近端二聚化结构域。利用酵母双杂交和生物化学测定，我们已经映射了一个替代的二聚化结构域的N端的PC 2（NT 2 -1-223，L224 X）。该结构域的功能表征表明，它足以诱导斑马鱼胚胎中的囊肿形成，并可能通过显性负性机制抑制mIMCD 3细胞中的PC 2表面电流。总之，我们提出了一个模型PC 2组件作为一个功能性的四聚体，这取决于C-和N-末端二聚化结构域。这些结果对我们理解PC 2功能和ADPKD发病机制具有重要意义，并为研究PC 2功能提供了新的策略。
Autosomal dominant polycystic kidney disease (ADPKD), the most common inherited cause of kidney failure, is caused by mutations in either PKD1 (85%) or PKD2 (15%). The PKD2 protein, polycystin-2 (PC2 or TRPP2), is a member of the transient receptor potential (TRP) superfamily and functions as a non-selective calcium channel. PC2 has been found to form oligomers in native tissues suggesting that it may form functional homo- or heterotetramers with other subunits, similar to other TRP channels. Our experiments unexpectedly revealed that PC2 mutant proteins lacking the known C-terminal dimerization domain were still able to form oligomers and co-immunoprecipitate full-length PC2, implying the possible existence of a proximal dimerization domain. Using yeast two-hybrid and biochemical assays, we have mapped an alternative dimerization domain to the N terminus of PC2 (NT2-1-223, L224X). Functional characterization of this domain demonstrated that it was sufficient to induce cyst formation in zebrafish embryos and inhibit PC2 surface currents in mIMCD3 cells probably by a dominant-negative mechanism. In summary, we propose a model for PC2 assembly as a functional tetramer which depends on both C- and N-terminal dimerization domains. These results have significant implications for our understanding of PC2 function and disease pathogenesis in ADPKD and provide a new strategy for studying PC2 function.