FUNCTION OF MST4-EZRIN-ACAP4 SIGNALING IN GASTRIC PARIETAL CELL SECRETION AND HOMEOSTASIS

MST4-EZRIN-ACAP4 信号传导在胃壁细胞分泌和稳态中的功能

• 批准号: 9753750
• 负责人: XUEBIAO YAO
• 金额: $ 32.23万
• 依托单位: MOREHOUSE SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-19 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9753750
• 关键词: Achlorhydria; Acids; Actin-Binding Protein; Address; Affect; Animals; Apical; Atrophic; Atrophic Gastritis; Binding; Biochemical; Biological Assay; Calpain; Cell division; Cell membrane; Cell physiology; Cell secretion; Chemicals; Chronic; Clinical; Coupled; Couples; Coupling; Cyclic AMP-Dependent Protein Kinases; Data; Development; Digestive Physiology; Emerging Technologies; Epithelial; Epithelial Cells; Epitopes; Exhibits; Foxes; Gastric Acid; Gastric Glands; Gastric Metaplasia; Gastric Parietal Cells; Gastric mucosa; Gastritis; Goals; Guanosine Triphosphate Phosphohydrolases; Helicobacter; Helicobacter Infections; Histamine; Homeostasis; Human; Hyperplasia; Image Analysis; In Vitro; Infection; Inflammatory; Inflammatory Response; Investigation; K ATPase; Light; Link; Malignant Neoplasms; Mediating; Membrane; Metaplasia; Microscope; Molecular; Organoids; Outcome; Parietal; Peptides; Pharmacology; Phosphoproteins; Phosphorylation; Physiological; Physiology; Point Mutation; Prevention; Prevention strategy; Protein Kinase; Proteins; Proteolysis; Proton Pump; Publications; Regulation; Research; Resolution; Role; Secretory Vesicles; Signal Pathway; Signal Transduction; Stomach; Structure; Testing; Toxin; apical membrane; basolateral membrane; cytokine; design; experience; ezrin; human pathogen; improved; inhibitor/antagonist; innovation; knock-down; knockout animal; mutant; novel; optical imaging; prevent; protein activation; real-time images; recruit; response; spatiotemporal; success; trafficking

The digestive function of the stomach depends on acidification of the gastric lumen. Acid secretion into the lumen is triggered by activation of a cAMP-dependent protein kinase (PKA) cascade, which ultimately results in the insertion of gastric H,K-ATPases into the apical plasma membranes of parietal cells. A coupling protein is ezrin, an 80 kDa phosphoprotein, whose phosphorylation at Ser66 by PKA is required for parietal cell activation. Our early study demonstrated that H. pylori VacA perturbs parietal cell secretion and induces parietal cell loss via calpain-elicited ezrin proteolysis. Recent studies demonstrate that gastric metaplasia and hyperplasia are results of ezrin loss in the parietal cells, suggesting the importance of ezrin signaling in parietal cell homeostasis and renewal. However, little is known regarding the molecular mechanism(s) by which the ezrin signaling pathway operates in gastric acid secretion and responds to H. pylori infection. The long-term goal of our research is to delineate how ezrin functions in gastric parietal cell secretion and homeostasis in response to the physiological stimulation and to H. pylori infection. To address this question, three Specific Aims are proposed: first, we will evaluate how MST4 interacts with ACAP4 and ezrin using epitope-tagging and chemical footprinting approaches. These studies will involve a detailed analysis of the structural determinants that mediate a direct MST4-ACAP4 contact. Binding domain data will be used to design peptides that potently and specifically perturb MST4-ACAP4 interactions in in vitro binding assays. The function of this interaction will then be evaluated by the effects of the peptides on acid secretion using permeabilized gastric glands and by introducing MST4-interaction deficient ACAP4 mutants into cultured parietal cells from ACAP4 knock-out animals. Second, we will define the role of MST4 in regulating spatiotemporal dynamics of ARF6 GTPase during parietal cell activation. The importance of such an interaction in acid secretion in response to physiological stimulation of H. pylori infection will then be evaluated by functional assay and super-resolution imaging analysis. Third, we plan to illustrate the molecular mechanisms by which MST4-ezrin-ACAP4 signaling axis orchestrates parietal cell homeostasis in human gastric organoids in the presence of H. pylori infection. These studies will be facilitated by biochemical and functional characterization coupled with optical imaging of secretory vesicle trafficking in live parietal cells with novel ACAP4 inhibitor MSM0601. Studying the molecular mechanisms underlying parietal cell secretion and homeostasis is of great significance in understanding the cellular physiology of regulated epithelial secretion in the gut, and is also expected to be of great benefit in leading to development of pharmacological strategies for correction and/or prevention of H. pylori infection-elicited gastric atrophy.

胃的消化功能取决于胃腔的酸化。酸分泌进入 该腔由cAMP依赖性蛋白激酶（PKA）级联的激活触发， 最终导致胃H，K-ATP酶插入壁细胞的顶端质膜， 细胞偶联蛋白是ezrin，一种80 kDa的磷蛋白，其通过PKA在Ser 66处磷酸化， 壁细胞激活所必需的。我们的早期研究表明H.幽门VacA干扰顶叶 细胞分泌并通过钙蛋白酶引起的埃兹蛋白水解诱导壁细胞损失。最近的研究 证明胃化生和增生是壁细胞中Ezrin丢失的结果， 提示Ezrin信号在壁细胞稳态和更新中的重要性。然而， 关于ezrin信号通路在胃粘膜中起作用的分子机制已知， 酸分泌，并对H.幽门感染我们研究的长期目标是 埃兹蛋白在胃壁细胞分泌和稳态中的作用 生理刺激和H.幽门感染为了解决这个问题，三个具体目标 首先，我们将使用表位标记来评估MST 4如何与ACAP 4和ezrin相互作用 和化学足迹法。这些研究将涉及详细的结构分析 介导MST 4-ACAP 4直接接触的决定簇。绑定域数据将用于设计 在体外结合测定中有效且特异性地干扰MST 4-ACAP 4相互作用的肽。的 然后通过肽对酸分泌的影响来评价这种相互作用的功能 通过将MST 4相互作用缺陷型ACAP 4突变体引入到胃粘膜中， 来自ACAP 4敲除动物的培养的壁细胞。第二，我们将确定MST 4在以下方面的作用： 在壁细胞激活期间调节ARF 6 GT3的时空动力学。的重要性 这种相互作用在酸分泌响应生理刺激的H。幽门螺杆菌感染将 然后通过功能测定和超分辨成像分析进行评价。第三，我们计划说明 MST 4-ezrin-ACAP 4信号轴协调壁细胞的分子机制 在H.幽门感染这些研究报告将 通过生物化学和功能表征以及分泌的光学成像来促进 囊泡运输活壁细胞与新的ACAP 4抑制剂MSM 0601。 研究壁细胞分泌和稳态的分子机制是非常重要的 在理解调节肠道上皮分泌的细胞生理学方面具有重要意义， 也有望在导致开发药理学策略方面有很大的好处， 纠正和/或预防H.幽门感染引起胃萎缩。

项目成果

Mitotic motor CENP-E cooperates with PRC1 in temporal control of central spindle assembly.

有丝分裂马达CENP-E与PRC1合作对中央纺锤体组件进行时间控制

• DOI: 10.1093/jmcb/mjz051
• 发表时间: 2020-08-01
• 期刊: Journal of molecular cell biology
• 影响因子: 5.5
• 作者: Liu X;Xu L;Li J;Yao PY;Wang W;Ismail H;Wang H;Liao B;Yang Z;Ward T;Ruan K;Zhang J;Wu Q;He P;Ding X;Wang D;Fu C;Dou Z;Yan F;Wang W;Liu X;Yao X
• 通讯作者: Yao X