Cyclic nucleotide-gated channel subunit glycosylation regulates matrix metalloproteinase-dependent changes in channel gating.
Cyclic nucleotide-gated channel subunit glycosylation regulates matrix metalloproteinase-dependent changes in channel gating.
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DOI：
10.1021/bi400824x
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发表时间：
2013-11-19
期刊：
影响因子：
2.9
通讯作者：
Varnum MD
中科院分区：
文献类型：
--
作者：
Meighan SE;Meighan PC;Rich ED;Brown RL;Varnum MD
环核苷酸门控（CNG）通道是视网膜光感受器内的光传导所必需的。我们之前已经证明基质金属蛋白酶-2和-9的酶活性增强了杆状（CNGA1 + CNGB1）和锥形CNGA3 + CNGB3) CNG通道的配体敏感性。此外，我们还观察到最大CNG通道电流（IMAX）的下降，这种下降在mmp定向门控变化的后期开始。在这里，我们证明了CNG通道在长时间暴露于MMP后变得不导电。与导电通道的损失同时发生的是表现出非修饰门控特性的通道的相对贡献增加，这表明存在一个亚群的通道免受mmp诱导的门控效应的保护。已知CNGA亚基具有一个细胞外核心糖基化位点，位于靠近孔形成区域的转塔环内的两个可能位置之一。我们的研究结果表明，CNGA糖基化可以阻碍mmp依赖性的CNG通道修饰。此外，孔转塔内糖基化位点的相对位置影响mmp依赖性蛋白水解的程度。发现在CNGA3亚基位点的糖基化具有保护作用，而在牛CNGA1位点的糖基化则没有。将CNGA1中的糖基化位点重新定位到CNGA3中发现的位置，再现了对mmp依赖性加工的CNGA3样保护。综上所述，这些数据表明CNGA糖基化可能保护CNG通道免受MMP依赖性蛋白水解的影响，这与MMP对通道功能的修饰需要物理进入通道的细胞外表面是一致的。
Cyclic-nucleotide gated (CNG) channels are essential for phototransduction within retinal photoreceptors. We have demonstrated previously that enzymatic activity of matrix metalloproteinase-2 and -9, members of the MMP family of extracellular, Ca+2- and Zn+2-dependent proteases, enhances the ligand sensitivity of both rod (CNGA1 + CNGB1) and cone CNGA3 + CNGB3) CNG channels. Additionally, we have observed a decrease in maximal CNG channel current (IMAX) that begins late during MMP-directed gating changes. Here we demonstrate that CNG channels become non-conductive after prolonged MMP exposure. Concurrent with the loss of conductive channels is the increased relative contribution of channels exhibiting non-modified gating properties, suggesting the presence of a subpopulation of channels that are protected from MMP-induced gating effects. CNGA subunits are known to possess one extracellular core glycosylation site, located at one of two possible positions within the turret loop near the pore-forming region. Our results indicate that CNGA glycosylation can impede MMP-dependent modification of CNG channels. Furthermore, the relative position of the glycosylation site within the pore turret influences the extent of MMP-dependent proteolysis. Glycosylation at the site found in CNGA3 subunits was found to be protective, while glycosylation at the bovine CNGA1 site was not. Relocating the glycosylation site in CNGA1 to the position found in CNGA3 recapitulated CNGA3-like protection from MMP-dependent processing. Taken together, these data indicate that CNGA glycosylation may protect CNG channels from MMP-dependent proteolysis, consistent with MMP modification of channel function having a requirement for physical access to the extracellular face of the channel.