Congenital chloride-losing diarrhea causing mutations in the STAS domain result in misfolding and mistrafficking of SLC26A3
Congenital chloride-losing diarrhea causing mutations in the STAS domain result in misfolding and mistrafficking of SLC26A3
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DOI：
10.1074/jbc.m704328200
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发表时间：
2008-03-28
影响因子：
4.8
通讯作者：
Thomas, Philip J.
中科院分区：
文献类型：
--
作者：
Dorwart, Michael R.;Shcheynikov, Nikolay;Thomas, Philip J.
先天性失氯性腹泻（CLD）是一种导致水样便和脱水的遗传性疾病。溶质载体26家族成员3（SLC26A3）作为一种偶联的Cl⁻/HCO₃⁻交换体，其发生突变会导致CLD。SLC26A3是一种膜蛋白，预计包含12个跨膜α-螺旋以及一个与细菌抗σ因子拮抗剂同源的C末端STAS（硫酸盐转运体和抗σ因子）结构域。STAS结构域是SLC26A3的Cl⁻/HCO₃⁻交换功能以及SLC26A3激活囊性纤维化跨膜传导调节因子所必需的。在此我们研究了STAS结构域中的四种导致CLD的突变（ΔY526/7、I544N、I675/6ins和G702Tins）引发疾病的分子机制。在异源哺乳动物表达系统中，生化、免疫组织化学和离子转运实验表明，这四种CLD突变导致SLC26A3转运体错误折叠和/或错误运输。对分离的STAS结构域进行的表达研究表明，I675/6ins和G702Tins突变直接破坏STAS结构域，而有限的蛋白水解实验表明，ΔY526/7和I544N突变影响折叠和/或运输途径中的后续步骤。数据表明，这些导致CLD的突变通过至少两种不同的分子机制引发疾病，最终都导致质膜上功能性蛋白质的缺失。
Congenital chloride-losing diarrhea (CLD) is a genetic disorder causing watery stool and dehydration. Mutations in SLC26A3 ( solute carrier 26 family member 3), which functions as a coupled Cl-/HCO3- exchanger, cause CLD. SLC26A3 is a membrane protein predicted to contain 12 transmembrane-spanning alpha-helices and a C-terminal STAS (sulfate transporters and anti-sigma-factor) domain homologous to the bacterial anti-sigma-factor antagonists. The STAS domain is required for SLC26A3 Cl-/HCO3- exchange function and for the activation of cystic fibrosis transmembrane conductance regulator by SLC26A3. Here we investigate the molecular mechanism(s) by which four CLD-causing mutations (Delta Y526/ 7, I544N, I675/6ins, and G702Tins) in the STAS domain lead to disease. In a heterologous mammalian expression system biochemical, immunohistochemical, and ion transport experiments suggest that the four CLD mutations cause SLC26A3 transporter misfolding and/or mistrafficking. Expression studies with the isolated STAS domain suggest that the I675/6ins and G702Tins mutations disrupt the STAS domain directly, whereas limited proteolysis experiments suggest that the Delta Y526/ 7 and I544N mutations affect a later step in the folding and/ or trafficking pathway. The data suggest that these CLD-causing mutations cause disease by at least two distinct molecular mechanisms, both ultimately leading to loss of functional protein at the plasma membrane.