Structural motifs for subtype-specific pH-sensitive gating of vertebrate otopetrin proton channels.
Structural motifs for subtype-specific pH-sensitive gating of vertebrate otopetrin proton channels.
复制标题
脊椎动物otopetrin质子通道亚型特异性ph敏感门控的结构基序。
DOI：
10.7554/elife.77946
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发表时间：
2022-08-03
期刊：
影响因子：
7.7
通讯作者：
Liman, Emily R.
中科院分区：
文献类型：
--
作者：
Teng, Bochuan;Kaplan, Joshua P.;Liang, Ziyu;Krieger, Zachary;Tu, Yu-Hsiang;Burendei, Batuujin;Ward, Andrew B.;Liman, Emily R.
关键词：
耳石蛋白（OTOP）通道是在脊椎动物和无脊椎动物中保守的质子选择性离子通道，与其他离子通道没有结构相似性。脊椎动物中有三种耳石蛋白通道（OTOP1、OTOP2和OTOP3），其中一种（OTOP1）作为酸味受体发挥作用。除了能透过细胞外质子外，细胞外质子是否能调控耳石蛋白通道尚不清楚。在此，我们利用膜片钳记录和胞质pH微量荧光测定法比较了三种小鼠耳石蛋白通道的功能特性。我们发现OTOP1和OTOP3都能被细胞外质子强烈激活，其细胞外pH阈值分别为<6.0和5.5，且动力学过程依赖于pH。相比之下，OTOP2通道在较大的pH范围内（pH 5 - pH 10）广泛具有活性，并在细胞外碱化（>pH 9.0）时产生外向电流。令人惊讶的是，我们可以通过交换连接跨膜结构域的细胞外连接子来改变OTOP2和OTOP3通道对pH敏感的门控特性。对包含跨膜结构域1 - 6的N结构域中的细胞外连接子进行交换，往往会改变嵌合通道在碱性pH下的相对电导，而对包含跨膜结构域7 - 12的C结构域内的连接子进行交换，则往往会改变OTOP3电流激活的速率。我们得出结论，耳石蛋白通道家族成员是质子门控（酸敏感）的质子通道，并且门控装置分布在通道的N和C结构域内的多个细胞外区域。除了味觉系统，耳石蛋白通道还在脊椎动物的前庭和消化系统中表达。我们所描述的独特门控特性可能使它们在这些以及其他生物系统中发挥不同的细胞类型特异性功能。
Otopetrin (OTOP) channels are proton-selective ion channels conserved among vertebrates and invertebrates, with no structural similarity to other ion channels. There are three vertebrate OTOP channels (OTOP1, OTOP2, and OTOP3), of which one (OTOP1) functions as a sour taste receptor. Whether extracellular protons gate OTOP channels, in addition to permeating them, was not known. Here, we compare the functional properties of the three murine OTOP channels using patch-clamp recording and cytosolic pH microfluorimetry. We find that OTOP1 and OTOP3 are both steeply activated by extracellular protons, with thresholds of pHo <6.0 and 5.5, respectively, and kinetics that are pH-dependent. In contrast, OTOP2 channels are broadly active over a large pH range (pH 5 pH 10) and carry outward currents in response to extracellular alkalinization (>pH 9.0). Strikingly, we could change the pH-sensitive gating of OTOP2 and OTOP3 channels by swapping extracellular linkers that connect transmembrane domains. Swaps of extracellular linkers in the N domain, comprising transmembrane domains 1–6, tended to change the relative conductance at alkaline pH of chimeric channels, while swaps within the C domain, containing transmembrane domains 7–12, tended to change the rates of OTOP3 current activation. We conclude that members of the OTOP channel family are proton-gated (acid-sensitive) proton channels and that the gating apparatus is distributed across multiple extracellular regions within both the N and C domains of the channels. In addition to the taste system, OTOP channels are expressed in the vertebrate vestibular and digestive systems. The distinct gating properties we describe may allow them to subserve varying cell-type specific functions in these and other biological systems.