Protease modulation of the activity of the epithelial sodium channel expressed in Xenopus oocytes.
Protease modulation of the activity of the epithelial sodium channel expressed in Xenopus oocytes.
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DOI：
10.1085/jgp.111.1.127
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发表时间：
1998-01
影响因子：
3.8
通讯作者：
Horisberger, JD
中科院分区：
文献类型：
--
作者：
Chra誰bi, A;Vallet, V;Firsov, D;Hess, SK;Horisberger, JD
我们研究了胞外蛋白水解酶对表达大鼠α，β和γ三个亚基的非洲爪哇卵母细胞的阿米洛利敏感钠电流(INA)的影响。低浓度的胰酶(2μg/ml)可在几分钟内引起INa的显著升高，这种作用可被大豆胰酶抑制剂完全阻断，但不能被阿米洛利所阻断。胰凝乳酶也有类似的作用，但激肽释放酶则不然。非洲爪哇和大鼠ENaC均可观察到胰酶诱导的INa升高，且在非洲爪哇ENaC的∼和α亚基与ENaC的β和γ亚基共表达所获得的通道中，INA的增加幅度很大(约为20倍)。胰酶对ENaC的作用是选择性的，K+通道ROMK2的表达对电流没有影响。细胞内注射EGTA或用GTP-γS预处理均不能阻断胰酶的作用，提示这一作用不是由G蛋白介导的。通过与FLAG表位结合的抗体对卵母细胞表面通道蛋白表达的测定表明，胰酶的作用并不伴随着通道蛋白密度的增加，这表明蛋白降解改变了存在于卵母细胞表面的通道的活性，而不是细胞表面的表达。在单通道水平，在细胞附着模式下，当吸管中有胰酶存在时，贴片中观察到更活跃的通道，而当在贴片吸管周围的浴液中加入胰酶时，通道活性没有变化。我们得出结论，细胞外蛋白水解酶能够增加上皮性钠通道的开放概率，这种作用不是通过激活G蛋白偶联受体而发生的，而是通过对作为通道本身的组成部分或与通道密切相关的蛋白质的蛋白分解来实现的。
We have investigated the effect of extracellular proteases on the amiloride-sensitive Na+ current (INa) in Xenopus oocytes expressing the three subunits α, β, and γ of the rat or Xenopus epithelial Na+ channel (ENaC). Low concentrations of trypsin (2 μg/ml) induced a large increase of INa within a few minutes, an effect that was fully prevented by soybean trypsin inhibitor, but not by amiloride. A similar effect was observed with chymotrypsin, but not with kallikrein. The trypsin-induced increase of INa was observed with Xenopus and rat ENaC, and was very large (∼20-fold) with the channel obtained by coexpression of the α subunit of Xenopus ENaC with the β and γ subunits of rat ENaC. The effect of trypsin was selective for ENaC, as shown by the absence of effect on the current due to expression of the K+ channel ROMK2. The effect of trypsin was not prevented by intracellular injection of EGTA nor by pretreatment with GTP-γS, suggesting that this effect was not mediated by G proteins. Measurement of the channel protein expression at the oocyte surface by antibody binding to a FLAG epitope showed that the effect of trypsin was not accompanied by an increase in the channel protein density, indicating that proteolysis modified the activity of the channel present at the oocyte surface rather than the cell surface expression. At the single channel level, in the cell-attached mode, more active channels were observed in the patch when trypsin was present in the pipette, while no change in channel activity could be detected when trypsin was added to the bath solution around the patch pipette. We conclude that extracellular proteases are able to increase the open probability of the epithelial sodium channel by an effect that does not occur through activation of a G protein-coupled receptor, but rather through proteolysis of a protein that is either a constitutive part of the channel itself or closely associated with it.
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影响因子：
64.8
作者：
Vallet, V;Chraibi, A;Rossier, BC
通讯作者：
Rossier, BC
DOI：
10.1085/jgp.81.6.785
发表时间：
1983-06
期刊：
The Journal of general physiology
影响因子：
--
作者：
Garty H;Edelman IS
通讯作者：
Edelman IS
DOI：
10.1085/jgp.92.1.121
发表时间：
1988-07
期刊：
The Journal of general physiology
影响因子：
--
作者：
Palmer LG;Frindt G
通讯作者：
Frindt G
影响因子：
2.4
作者：
LEWIS, SA;CLAUSEN, C
通讯作者：
CLAUSEN, C
DOI：
10.1073/pnas.83.14.5345
发表时间：
1986-07-01
影响因子：
11.1
作者：
LEWIS, SA;ALLES, WP
通讯作者：
ALLES, WP