Functional Requirement for a Highly Conserved Charged Residue at Position 75 in the Gap Junction Protein Connexin 32
Functional Requirement for a Highly Conserved Charged Residue at Position 75 in the Gap Junction Protein Connexin 32
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DOI：
10.1074/jbc.m112.392670
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发表时间：
2013-02-01
影响因子：
4.8
通讯作者：
Freidin, Mona M.
中科院分区：
文献类型：
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作者：
Abrams, Charles K.;Islam, Mahee;Freidin, Mona M.
Charcot玛丽牙病（CMT）是一组以周围神经功能障碍为主要特征的遗传性疾病。CMT 1X是X连锁CMT的最常见形式，由连接蛋白32（Cx 32）突变引起。在这项工作中，我们使用双全细胞膜片钳记录来检查Arg（75）位置突变的功能效应。该残基在连接蛋白家族的成员中高度保守，并且已经在Cx 26、Cx43和Cx46中的该（或相应）位置处鉴定了致病突变。因此，更好地了解Cx 32中该位置突变的影响可能与许多不同人类疾病的发病机制有关。与CMT 1X相关的所有三种突变体（R75 P、R75 Q和R75 W）显示出与单独用载体转染的细胞相似的非常低水平的偶联。与Cx 32 WT的异型配对表明，这些突变体在同型构型中不存在偶联，这可以通过它们的半通道G（j）-V-j关系的变化来解释。在该位置处的另外七个突变体（R75 A、R75 D、R75 E、R75 H、R75 K、R75 L和R75 V）的表达水平和门控特征的检查表明，在Cx 32中的位置75处的正电荷是正常通道功能所需的，但不是间隙连接组装所需的。我们的研究还表明，CMT 1X的疾病治疗策略，纠正Cx 32的贩运异常，可能是无效的突变组也赋予Cx 32通道的门控特性的变化。
Charcot Marie Tooth disease (CMT) is a group of inherited disorders characterized clinically by exclusively or predominantly peripheral nerve dysfunction. CMT1X, the most common form of X-linked CMT is caused by mutations in connexin 32 (Cx32). In this work, we used dual whole cell patch clamp recording to examine the functional effects of mutations at the Arg(75) position. This residue is highly conserved among members of the connexin family, and disease-causing mutations have been identified at this (or the corresponding) position in Cx26, Cx43, and Cx46. Thus, a better understanding of the effects of mutations of this position in Cx32 may have relevance to pathogenesis of a number of different human diseases. All three mutants associated with CMT1X (R75P, R75Q, and R75W) showed very low levels of coupling similar to those of the cells transfected with vector alone. Heterotypic pairing with Cx32 WT showed that the absence of coupling for these mutants in the homotypic configuration could be explained by shifts in their hemichannel G(j)-V-j relations. Examination of the expression levels and gating characteristics of seven additional mutants (R75A, R75D, R75E, R75H, R75K, R75L, and R75V) at this position suggest that the positive charge at position 75 in Cx32 is required for normal channel function but not for gap junction assembly. Our studies also suggest that disease treatment strategies for CMT1X, which correct trafficking abnormalities in Cx32, may be ineffective for the group of mutations also conferring changes in gating properties of Cx32 channels.