Single Cl- channels activated by Ca2+ in Drosophila S2 cells are mediated by bestrophins.
Single Cl- channels activated by Ca2+ in Drosophila S2 cells are mediated by bestrophins.
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DOI：
10.1085/jgp.200609581
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发表时间：
2006-09
期刊：
影响因子：
--
通讯作者：
Hartzell HC
中科院分区：
文献类型：
--
作者：
Chien LT;Zhang ZR;Hartzell HC
人类Bestrophin-1(VMD2)的突变在基因上与几种形式的视网膜变性有关，但其潜在机制尚不清楚。Bestrophin-1(HBest1)是一种氯−通道，参与视网膜色素上皮的离子和液体运输。然而，到目前为止，Bestrophin电流只在过表达系统中被描述，而在任何天然细胞中都没有描述。为了检测Bestrophins在生理上是否作为钙激活的Cl-−通道，我们在果蝇S2细胞系中使用了干扰RNA。S2细胞表达四种bestrophin(dest1-4)，并具有内源性CAC电流。CAC电流被几个RNAi结构取消为dest1和dest2，但不是dest3或dest4。通过在HEK细胞中表达dest1来模拟内源性CAC电流，并用半胱氨酸取代F81来改变该电流的整流性和相对通透性。S2Bestrophin电流的单通道分析显示∼2-pS单通道具有快速的门控动力学和线性的电流-电压关系。在转导Dest1的CHO细胞中观察到了类似的通道，而在对Dest1的RNAi处理的S2细胞中没有观察到这种通道。这为Bestrophins是质膜上天然CAC通道的组成部分提供了确凿的证据。
Mutations in human bestrophin-1 (VMD2) are genetically linked to several forms of retinal degeneration but the underlying mechanisms are unknown. Bestrophin-1 (hBest1) has been proposed to be a Cl− channel involved in ion and fluid transport by the retinal pigment epithelium (RPE). To date, however, bestrophin currents have only been described in overexpression systems and not in any native cells. To test whether bestrophins function as Ca2+-activated Cl− (CaC) channels physiologically, we used interfering RNA (RNAi) in the Drosophila S2 cell line. S2 cells express four bestrophins (dbest1–4) and have an endogenous CaC current. The CaC current is abolished by several RNAi constructs to dbest1 and dbest2, but not dbest3 or dbest4. The endogenous CaC current was mimicked by expression of dbest1 in HEK cells, and the rectification and relative permeability of the current were altered by replacing F81 with cysteine. Single channel analysis of the S2 bestrophin currents revealed an ∼2-pS single channel with fast gating kinetics and linear current–voltage relationship. A similar channel was observed in CHO cells transfected with dbest1, but no such channel was seen in S2 cells treated with RNAi to dbest1. This provides definitive evidence that bestrophins are components of native CaC channels at the plasma membrane.
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影响因子：
3.8
作者：
Marmorstein, Lihua Y;Wu, Jiang;McLaughlin, Precious;Yocom, John;Karl, Mike O;Neussert, Rudgar;Wimmers, Soenke;Stanton, J Brett;Gregg, Ronald G;Strauss, Olaf;Peachey, Neal S;Marmorstein, Alan D
通讯作者：
Marmorstein, Alan D
影响因子：
4.8
作者：
Ho, MWY;Kaetzel, MA;Shears, SB
通讯作者：
Shears, SB
影响因子：
4.5
作者：
KLOCKNER, U
通讯作者：
KLOCKNER, U
影响因子：
8.4
作者：
Hartzell, C;Qu, ZQ;Cui, YY
通讯作者：
Cui, YY
影响因子：
4.4
作者：
Bakall, B;Marmorstein, LY;Marmorstein, AD
通讯作者：
Marmorstein, AD