Polycystin-1 Interacts with Inositol 1,4,5-Trisphosphate Receptor to Modulate Intracellular Ca2+ Signaling with Implications for Polycystic Kidney Disease
Polycystin-1 Interacts with Inositol 1,4,5-Trisphosphate Receptor to Modulate Intracellular Ca2+ Signaling with Implications for Polycystic Kidney Disease
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DOI：
10.1074/jbc.m109.068916
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发表时间：
2009-12-25
影响因子：
4.8
通讯作者：
Guggino, William B.
中科院分区：
文献类型：
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作者：
Li, Yun;Santoso, Netty G.;Guggino, William B.
PKD 1或PKD 2基因编码与多囊肾病相关的多囊蛋白（PC）1和2。以前，我们证明，PC 2与肌醇1，4，5-三磷酸受体（IP 3R）相互作用，以调节Ca 2+信号。在这里，我们调查PC 1是否也调节IP 3R。我们产生了一个片段编码的最后六个跨膜（TM）结构域的PC 1和C-末端尾（QIF 38），与PC 2的同源性最高的部分。使用非洲爪蟾卵母细胞Ca 2+成像系统，我们观察到QIF 38的表达显着降低IP 3诱导的Ca 2+瞬变的初始幅度，而缺乏C-末端尾部的突变没有。因此，C末端对于QIF 38功能是必需的。免疫共沉淀试验表明，QIF 38通过其C末端与IP 3R的IP 3结合结构域结合。一个较短的PC 1片段跨越只有最后的TM和C-末端的尾巴也减少IP 3诱导的Ca 2+释放，而另一个C-末端片段缺乏任何TM域没有。因此，只有内质网定位的PC 1可以调节IP 3R。最后，我们表明，在极化的Madin-Darby犬肾细胞，异源表达的全长PC 1导致在一个较小的IP 3诱导的Ca 2+反应。过表达的IP 3结合结构域的IP 3R逆转了PC 1的抑制作用，这表明全长PC 1（或其裂解形式）与内源性IP 3R在Madin-Darby犬肾细胞中的相互作用。这些结果表明，在哺乳动物细胞中的全长PC 1的行为是一致的PC 1的C-末端片段在卵母细胞系统。这些数据表明，PC 1抑制Ca 2+释放，可能与PC 2的作用相反，后者通过IP 3R促进Ca 2+释放。
The PKD1 or PKD2 genes encode polycystins (PC) 1 and 2, which are associated with polycystic kidney disease. Previously we demonstrated that PC2 interacts with the inositol 1,4,5-trisphosphate receptor (IP3R) to modulate Ca2+ signaling. Here, we investigate whether PC1 also regulates IP3R. We generated a fragment encoding the last six transmembrane (TM) domains of PC1 and the C-terminal tail (QIF38), a section with the highest homology to PC2. Using a Xenopus oocyte Ca2+ imaging system, we observed that expression of QIF38 significantly reduced the initial amplitude of IP3-induced Ca2+ transients, whereas a mutation lacking the C-terminal tail did not. Thus, the C terminus is essential to QIF38 function. Co-immunoprecipitation assays demonstrated that through its C terminus, QIF38 associates with the IP3-binding domain of IP3R. A shorter PC1 fragment spanning only the last TM and the C-terminal tail also reduced IP3-induced Ca2+ release, whereas another C-terminal fragment lacking any TM domain did not. Thus, only endoplasmic reticulum-localized PC1 can modulate IP3R. Finally, we show that in the polarized Madin-Darby canine kidney cells, heterologous expression of full-length PC1 resulted in a smaller IP3-induced Ca2+ response. Overexpression of the IP3-binding domain of IP3R reversed the inhibitory effect of PC1, suggesting interaction of full-length PC1 (or its cleavage forms) with endogenous IP3R in Madin-Darby canine kidney cells. These results indicate that the behavior of full-length PC1 in mammalian cells is congruent with that of PC1 C-terminal fragments in the oocyte system. These data demonstrate that PC1 inhibits Ca2+ release, perhaps opposing the effect of PC2, which facilitates Ca2+ release through the IP3R.