Laboratory of Genetic Skin Diseases Division of Dermatology, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan.
Br J Dermatol. 2013 Mar;168(3):629-33. doi: 10.1111/bjd.12018. Epub 2012 Dec 13.
Hypohidrotic ectodermal dysplasia (HED) is a rare condition characterized by hypotrichosis, hypohidrosis and hypodontia. A de novo heterozygous mutation in the tumour necrosis factor receptor-associated factor 6 gene (TRAF6) was recently identified in a patient with HED, while functional consequences resulting from the mutation remained unknown.
To determine the mechanism by which the TRAF6 mutation results in HED.
We performed coimmunoprecipitation (co-IP) studies to determine whether the mutation would affect the interaction of TRAF6 with transforming growth factor β-activated kinase 1 (TAK1), TAK1-binding protein 2 (TAB 2) and ectodysplasin-A receptor-associated death domain protein (EDARADD). We then performed co-IP and glutathione S-transferase-pulldown assays to determine the TRAF6 binding sequences in EDARADD. In addition, we analysed the effect of the mutant TRAF6 protein on the affinity between wild-type TRAF6 and EDARADD, as well as on EDARADD-mediated nuclear factor (NF)-κB activation.
The mutant TRAF6 protein was capable of forming a complex with TAK1 and TAB 2 in a similar way to wild-type TRAF6. However, the mutant TRAF6 protein completely lost the affinity to EDARADD, while the wild-type TRAF6 bound to the N-terminal domain of EDARADD. Furthermore, the mutant TRAF6 inhibited the interaction between the wild-type TRAF6 and EDARADD, and also potentially reduced the EDARADD-mediated NF-κB activity.
We conclude that the mutant TRAF6 protein shows a dominant negative effect against the wild-type TRAF6 protein, which is predicted to affect the EDARADD-mediated activation of NF-κB during the development of ectoderm-derived organs, and to lead to the HED phenotype.
先天性外胚层发育不良（HED）是一种罕见的疾病，其特征为毛发稀疏、少汗和牙齿缺失。最近，在一名 HED 患者中发现肿瘤坏死因子受体相关因子 6 基因（TRAF6）的新生杂合突变，而突变引起的功能后果尚不清楚。
确定 TRAF6 突变导致 HED 的机制。
我们进行了共免疫沉淀（co-IP）研究，以确定该突变是否会影响 TRAF6 与转化生长因子β激活激酶 1（TAK1）、TAK1 结合蛋白 2（TAB 2）和外胚层发育不良蛋白相关死亡域蛋白（EDARADD）的相互作用。然后，我们进行了 co-IP 和谷胱甘肽 S-转移酶下拉实验，以确定 EDARADD 中 TRAF6 的结合序列。此外，我们分析了突变型 TRAF6 蛋白对野生型 TRAF6 与 EDARADD 之间亲和力的影响，以及对 EDARADD 介导的核因子（NF）-κB 激活的影响。
突变型 TRAF6 蛋白能够以类似于野生型 TRAF6 的方式形成与 TAK1 和 TAB 2 的复合物。然而，突变型 TRAF6 蛋白完全失去与 EDARADD 的亲和力，而野生型 TRAF6 结合 EDARADD 的 N 端结构域。此外，突变型 TRAF6 抑制了野生型 TRAF6 与 EDARADD 之间的相互作用，并可能降低 EDARADD 介导的 NF-κB 活性。
我们的结论是，突变型 TRAF6 蛋白对野生型 TRAF6 蛋白表现出显性负效应，这可能会影响外胚层衍生器官发育过程中 EDARADD 介导的 NF-κB 激活，并导致 HED 表型。
