Golgi and plasma membrane pools of PI(4)P contribute to plasma membrane PI(4,5)P2 and maintenance of KCNQ2/3 ion channel current
Golgi and plasma membrane pools of PI(4)P contribute to plasma membrane PI(4,5)P2 and maintenance of KCNQ2/3 ion channel current
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DOI：
10.1073/pnas.1407133111
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发表时间：
2014-06-03
影响因子：
11.1
通讯作者：
Hille, Bertil
中科院分区：
文献类型：
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作者：
Dickson, Eamonn J.;Jensen, Jill B.;Hille, Bertil
质膜（PM）磷脂酰肌醇4，5-二磷酸[PI（4，5）P-2]调节许多离子通道和其他膜相关蛋白的活性。为了确定tsA-201细胞中PM PI（4，5）P-2池的前体来源，我们监测KCNQ 2/3通道电流和PHPLC delta 1结构域的移位作为PM PI（4，5）P-2的实时指标，以及PHOSH 2x2和PHOSH 1结构域的移位分别作为PM和高尔基体磷脂酰肌醇4-磷酸[PI（4）P]的指标。我们选择性地耗尽PI（4）P池在PM，高尔基体，或两者使用雷帕霉素募集脂质4-磷酸酶。用可再募集的4-磷酸酶（Sac 1）耗尽PM处的PI（4）P导致通过电或光学指示剂测量的PI（4，5）P-2的减少。用4-磷酸酶耗尽高尔基体处的PI（4）P或破坏膜转运马达诱导PM PI（4，5）P-2下降。在高尔基体和PM处同时消耗PI（4）P诱导PI（4，5）P-2的较大降低。PI（4，5）P-2的下降后4-磷酸酶的招聘需要1-2分钟。招聘内质网（ER）对高尔基体膜模仿的影响，耗尽PI（4）P在高尔基体，显然是由于内源性ER Sac 1的transactions。因此，PI（4，5）P-2的PM库的维持似乎取决于PM和高尔基体中PI（4）P的前体库。当Sac 1被募集到高尔基体时，PM PI（4，5）P-2的减少表明高尔基体的贡献是持续的，PI（4，5）P-2的产生可能与重要的细胞生物学过程如膜运输或脂质转移活性偶联。
Plasma membrane (PM) phosphatidylinositol 4,5-bisphosphate [PI(4,5)P-2] regulates the activity of many ion channels and other membrane-associated proteins. To determine precursor sources of the PM PI(4,5)P-2 pool in tsA-201 cells, we monitored KCNQ2/3 channel currents and translocation of PHPLC delta 1 domains as real-time indicators of PM PI(4,5)P-2, and translocation of PHOSH2x2, and PHOSH1 domains as indicators of PM and Golgi phosphatidylinositol 4-phosphate [PI(4) P], respectively. We selectively depleted PI(4) P pools at the PM, Golgi, or both using the rapamycin-recruitable lipid 4-phosphatases. Depleting PI(4) P at the PM with a recruitable 4-phosphatase (Sac1) results in a decrease of PI(4,5)P-2 measured by electrical or optical indicators. Depleting PI(4) P at the Golgi with the 4-phosphatase or disrupting membrane-transporting motors induces a decline in PM PI(4,5)P-2. Depleting PI(4) P simultaneously at both the Golgi and the PM induces a larger decrease of PI(4,5)P-2. The decline of PI(4,5)P-2 following 4-phosphatase recruitment takes 1-2 min. Recruiting the endoplasmic reticulum (ER) toward the Golgi membranes mimics the effects of depleting PI(4) P at the Golgi, apparently due to the trans actions of endogenous ER Sac1. Thus, maintenance of the PM pool of PI(4,5)P-2 appears to depend on precursor pools of PI(4) P both in the PM and in the Golgi. The decrease in PM PI(4,5)P-2 when Sac1 is recruited to the Golgi suggests that the Golgi contribution is ongoing and that PI (4,5)P-2 production may be coupled to important cell biological processes such as membrane trafficking or lipid transfer activity.