Sodium-potassium ATPase 1 subunit is a molecular partner of Wolframin, an endoplasmic reticulum protein involved in ER stress.
Sodium-potassium ATPase 1 subunit is a molecular partner of Wolframin, an endoplasmic reticulum protein involved in ER stress.
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DOI：
10.1093/hmg/ddm296
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发表时间：
2008-01-15
影响因子：
3.5
通讯作者：
Barrett TG
中科院分区：
文献类型：
--
作者：
Zatyka M;Ricketts C;da Silva Xavier G;Minton J;Fenton S;Hofmann-Thiel S;Rutter GA;Barrett TG
Wolfram综合征是一种常染色体隐性遗传病，以糖尿病和视神经萎缩为特征，由编码内质网（ER）膜蛋白Wolframin的WFS1基因突变引起。虽然其确切功能尚不清楚，但Wolframin缺乏增加内质网应激，损害细胞周期进程并影响钙稳态。为了进一步了解其功能并鉴定其分子伙伴，我们将wfs1 -羧基末端结构域作为诱饵，在酵母双杂交筛选中与人脑cDNA文库结合。Na+/K+ atp酶β1亚基被鉴定为一个互作克隆。我们将相互作用定位到WFS1的羧基末端和跨膜结构域，而不是氨基末端结构域。我们的图谱数据显示，这种相互作用最有可能发生在急诊室。我们在哺乳动物细胞中通过共免疫沉淀证实了这种相互作用，并在JEG3胎盘细胞和神经母细胞瘤SKNAS细胞中与内源性蛋白相互作用。与野生型相比，人WFS1突变型成纤维细胞中Na+/K+ atp酶β1亚基的表达降低，与野生型相比，WFS1沉默的MIN6胰腺β-细胞中Na+/K+ atp酶β1亚基的表达降低。在野生型细胞中，内质网应激的诱导仅部分解释了Na+/K+ atp酶β1亚基表达的降低。Na+/K+ atp酶β1亚基是Wolframin的第一个蛋白伴侣；这种相互作用最可能发生在内质网，并且可能对Na+/K+ atp酶β1亚基的表达、折叠或成熟很重要。
Wolfram syndrome, an autosomal recessive disorder characterised by diabetes mellitus and optic atrophy, is caused by mutations in the WFS1 gene encoding an endoplasmic reticulum (ER) membrane protein, Wolframin. Although its precise functions are unknown, Wolframin deficiency increases ER stress, impairs cell cycle progression and affects calcium homeostasis. To gain further insight into its function and identify molecular partners, we used the WFS1-carboxy-terminal domain as bait in a yeast two-hybrid screen with a human brain cDNA library. Na+/K+ ATPase β1 subunit was identified as an interacting clone. We mapped the interaction to the WFS1 carboxy-terminal and transmembrane domains, but not the amino-terminal domain. Our mapping data suggest that the interaction most likely occurs in the ER. We confirmed the interaction by co-immunoprecipitation in mammalian cells, and with endogenous proteins in JEG3 placental cells and neuroblastoma SKNAS cells. The expression of Na+/K+ ATPase β1 subunit was reduced in human WFS1 mutant fibroblasts compared with wild type, and in WFS1 silenced MIN6 pancreatic β-cells compared with wild type. Induction of ER stress in wild type cells only partly accounted for the reduced Na+/K+ ATPase β1 subunit expression observed. Na+/K+ ATPase β1 subunit is the first identified protein partner of Wolframin; the interaction is most likely to occur in the ER, and may be important for Na+/K+ ATPase β1 subunit expression, folding or maturation.
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