Phospho-regulated ACAP4-Ezrin Interaction Is Essential for Histamine-stimulated Parietal Cell Secretion
Phospho-regulated ACAP4-Ezrin Interaction Is Essential for Histamine-stimulated Parietal Cell Secretion
复制标题
磷酸调节的 ACAP4-Ezrin 相互作用对于组胺刺激的壁细胞分泌至关重要
DOI：
10.1074/jbc.m110.129007
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发表时间：
2010-06-11
影响因子：
4.8
通讯作者：
Yao, Xuebiao
中科院分区：
文献类型：
--
作者：
Ding, Xia;Deng, Hui;Yao, Xuebiao
ezrin-radixin-moesin蛋白在膜蛋白和皮质细胞骨架之间提供调节的连接，并且还参与信号转导途径。Ezrin定位于壁细胞的顶膜，并将蛋白激酶A激活级联与受调节的HCl分泌偶联。我们最近的蛋白质组学研究揭示了对挥发性膜重塑必需的ezrin-ACAP 4-ARF 6的蛋白质复合物（Fang，Z.，苗，Y.，丁某，Deng，H.，Liu，S.，王福，周，R.，沃森，C.，傅，C.，胡昆，利拉德，J.W.，小的，鲍威尔，M.，陈玉，Forte，J.G.，和Yao，X.（2006）Mol. Cell Proteomics 5，1437-1449）。然而，关于ACAP 4是否与ezrin物理相互作用以及它们的相互作用如何整合到膜细胞骨架重塑中的知识仍然难以捉摸。在这里，我们提供了第一个证据表明，ezrin与ACAP 4相互作用的蛋白激酶A介导的磷酸化依赖的方式通过N-末端400氨基酸的ACAP 4。ACAP 4位于静息壁细胞的细胞质膜中，但在组胺刺激时移位至顶端质膜。ACAP 4与埃兹蛋白从分泌而非静息壁细胞裂解物中沉淀，表明磷酸调节的相互作用。事实上，这种相互作用被磷酸酶处理消除，并通过使用磷酸模拟埃兹蛋白（S66 D）的体外重建测定法进行验证。重要的是，埃兹蛋白指定分泌壁细胞中的ACAP 4的顶端分布，因为埃兹蛋白的抑制或非磷酸化埃兹蛋白的过表达阻止了ACAP 4的顶端定位。此外，过表达GTP酶激活蛋白缺陷型ACAP 4导致顶端膜细胞骨架重塑和胃酸分泌的抑制。总之，这些结果定义了一种新的分子机制连接ACAP 4-ezrin相互作用极化上皮分泌。
The ezrin-radixin-moesin proteins provide a regulated linkage between membrane proteins and the cortical cytoskeleton and also participate in signal transduction pathways. Ezrin is localized to the apical membrane of parietal cells and couples the protein kinase A activation cascade to the regulated HCl secretion. Our recent proteomic study revealed a protein complex of ezrin-ACAP4-ARF6 essential for volatile membrane remodeling (Fang, Z., Miao, Y., Ding, X., Deng, H., Liu, S., Wang, F., Zhou, R., Watson, C., Fu, C., Hu, Q., Lillard, J. W., Jr., Powell, M., Chen, Y., Forte, J. G., and Yao, X. (2006) Mol. Cell Proteomics 5, 1437-1449). However, knowledge of whether ACAP4 physically interacts with ezrin and how their interaction is integrated into membrane-cytoskeletal remodeling has remained elusive. Here we provide the first evidence that ezrin interacts with ACAP4 in a protein kinase A-mediated phosphorylation-dependent manner through the N-terminal 400 amino acids of ACAP4. ACAP4 locates in the cytoplasmic membrane in resting parietal cells but translocates to the apical plasma membrane upon histamine stimulation. ACAP4 was precipitated with ezrin from secreting but not resting parietal cell lysates, suggesting a phospho-regulated interaction. Indeed, this interaction is abolished by phosphatase treatment and validated by an in vitro reconstitution assay using phospho-mimicking ezrin(S66D). Importantly, ezrin specifies the apical distribution of ACAP4 in secreting parietal cells because either suppression of ezrin or overexpression of non-phosphorylatable ezrin prevents the apical localization of ACAP4. In addition, overexpressing GTPase-activating protein-deficient ACAP4 results in an inhibition of apical membrane-cytoskeletal remodeling and gastric acid secretion. Taken together, these results define a novel molecular mechanism linking ACAP4-ezrin interaction to polarized epithelial secretion.