A single amino acid residue constitutes the third dimerization domain essential for the assembly and function of the tetrameric polycystin-2 (TRPP2) channel.
A single amino acid residue constitutes the third dimerization domain essential for the assembly and function of the tetrameric polycystin-2 (TRPP2) channel.
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DOI：
10.1074/jbc.m110.192286
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发表时间：
2011-05-27
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;Rodat-Despoix L;Delmas P;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，这表明与其他TRP通道类似，它可以与其他TRP亚基形成功能性同源或异源四聚体。我们最近已经证明，PC 2的同源二聚化是由N-末端和C-末端结构域介导的，并且已知PC 2可以与PC 1、TRPC 1和TRPV 4异源二聚化。在本文中，我们报告了一个单一的半胱氨酸残基，Cys 632，在一个已知的PKD 2家系突变，构成第三个二聚化结构域的PC 2。缺失N和C末端的PC 2截短突变体在非还原条件下仍能二聚化。Cys 632单独突变废除在这些突变体中的二聚化，表明它是介导PC 2单体之间二硫键形成的关键残基。C632 A PC 2突变体与野生型PC 2通道的共表达减少了HEK 293细胞中ATP敏感性内质网Ca 2+的释放。C632 A和破坏C-末端卷曲螺旋结构域的突变（Val 846、Ile 853、Ile 860、Leu 867或4 M）的组合几乎消除了二聚体形成和ATP依赖性Ca 2+释放。然而，与4 M PC 2突变体不同，C632 A突变体仍然可以与多囊蛋白-1（PC 1）异源二聚体化。我们的研究结果表明，PC 2同源二聚体是由三个不同的域，这些事件调节四聚体PC 2通道的形成。
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 nonselective calcium channel. PC2 has been found to form oligomers in native tissues, suggesting that similar to other TRP channels, it may form functional homo- or heterotetramers with other TRP subunits. We have recently demonstrated that the homodimerization of PC2 is mediated by both N-terminal and C-terminal domains, and it is known that PC2 can heterodimerize with PC1, TRPC1, and TRPV4. In this paper, we report that a single cysteine residue, Cys632, mutated in a known PKD2 pedigree, constitutes the third dimerization domain for PC2. PC2 truncation mutants lacking both N and C termini could still dimerize under nonreducing conditions. Mutation of Cys632 alone abolished dimerization in these mutants, indicating that it was the critical residue mediating disulfide bond formation between PC2 monomers. Co-expression of C632A PC2 mutants with wild-type PC2 channels reduced ATP-sensitive endoplasmic reticulum Ca2+ release in HEK293 cells. The combination of C632A and mutations disrupting the C-terminal coiled-coil domain (Val846, Ile853, Ile860, Leu867 or 4M) nearly abolished dimer formation and ATP-dependent Ca2+ release. However, unlike the 4M PC2 mutant, a C632A mutant could still heterodimerize with polycystin-1 (PC1). Our results indicate that PC2 homodimerization is regulated by three distinct domains and that these events regulate formation of the tetrameric PC2 channel.