Carbachol-mediated endocytosis of NHE3 involves a clathrin-independent mechanism requiring lipid rafts and Cdc42.
Carbachol-mediated endocytosis of NHE3 involves a clathrin-independent mechanism requiring lipid rafts and Cdc42.
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DOI：
10.1159/000358659
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发表时间：
2014
期刊：
影响因子：
--
通讯作者：
Kovbasnjuk O
中科院分区：
文献类型：
--
作者：
Zachos NC;Alamelumangpuram B;Lee LJ;Wang P;Kovbasnjuk O
在肠上皮细胞中，刷状边界Na+/H+交换剂NHE3的急性调节通常通过内吞作用和/或胞外作用的改变发生。组成型NHE3胞吞作用涉及网格蛋白。碳水化合物（CCH），提高细胞内Ca2+ ([Ca2+]i)，降低NHE3活性，刺激内吞作用；然而，钙介导的NHE3内吞作用的机制尚不清楚。脂筏中存在大量的NHE3，这有助于基础的NHE3运输，而不是camp介导的NHE3运输，这表明NHE3内吞作用存在另一种机制。Cdc42被证明在一些胆固醇敏感的、不依赖网格蛋白的内吞作用中起着不可或缺的作用。因此，本研究旨在验证以下假设：(1)网格蛋白介导的内吞作用（CME）参与了NHE3的组成性内吞作用，而不是cch介导的内吞作用；(2)cch介导的NHE3的内吞作用是通过脂质筏激活的cdc42依赖途径发生的，而不涉及网格蛋白。在极化Caco-2/BBe细胞中，采用药理学和shRNA敲低方法研究了Cdc42和脂筏对NHE3活性和内吞作用的影响。基础NHE3活性在CME阻滞剂（氯丙嗪；K+消耗）的存在下增加，这支持了先前的报道，即构成性NHE3内吞作用依赖于网格蛋白。相比之下，cch对NHE3活性的抑制作用在MβCD处理的Caco-2/BBe细胞和Cdc42敲低细胞中被消除，但CME阻滞剂不受影响。cch介导的NHE3活性抑制不依赖于网格蛋白，涉及脂质筏，需要Cdc42。
In intestinal epithelial cells, acute regulation of the brush border Na+/H+ exchanger, NHE3, usually occurs by changes in endocytosis and/or exocytosis. Constitutive NHE3 endocytosis involves clathrin. Carbachol (CCH), which elevates intracellular Ca2+ ([Ca2+]i), decreases NHE3 activity and stimulates endocytosis; however, the mechanism involved in calcium-mediated endocytosis of NHE3 is unclear. A pool of NHE3 resides in lipid rafts, which contributes to basal, but not cAMP-mediated, NHE3 trafficking, suggesting that an alternative mechanism exists for NHE3 endocytosis. Cdc42 was demonstrated to play an integral role in some cases of cholesterol-sensitive, clathrin-independent endocytosis. Therefore, the current study was designed to test the hypotheses that (1) clathrin-mediated endocytosis (CME) is involved in constitutive, but not CCH-mediated, endocytosis of NHE3, and (2) CCH-mediated endocytosis of NHE3 occurs through a lipid raft, activated Cdc42-dependent pathway that does not involve clathrin. The role of Cdc42 and lipid rafts on NHE3 activity and endocytosis were investigated in polarized Caco-2/BBe cells using pharmacological and shRNA knockdown approaches. Basal NHE3 activity was increased in the presence of CME blockers (chlorpromazine; K+ depletion) supporting previous reports that constitutive NHE3 endocytosis is clathrin dependent. In contrast, CCH-inhibition of NHE3 activity was abolished in Caco-2/BBe cells treated with MβCD (to disrupt lipid rafts) as well as in Cdc42 knockdown cells but was unaffected by CME blockers. CCH-mediated inhibition of NHE3 activity is not dependent on clathrin and involves lipid rafts and requires Cdc42.
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影响因子：
3.3
作者：
Eyster CA;Cole NB;Petersen S;Viswanathan K;Früh K;Donaldson JG
通讯作者：
Donaldson JG
影响因子：
15.9
作者：
DONOWITZ, M;COHEN, ME;SHARP, GWG
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影响因子：
5.5
作者：
Donowitz, M;Cha, BY;Li, XH
通讯作者：
Li, XH
DOI：
10.1083/jcb.83.1.82
发表时间：
1979-10
期刊：
The Journal of cell biology
影响因子：
--
作者：
Haigler HT;McKanna JA;Cohen S
通讯作者：
Cohen S
影响因子：
4.8
作者：
Janecki, AJ;Montrose, MH;Donowitz, M
通讯作者：
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