Role of cytoskeletal interactions with mechanogated ion channel Piezo2 in the mechanism of mechanotransduction by enteroendocrine cells

细胞骨架与机械离子通道 Piezo2 的相互作用在肠内分泌细胞机械转导机制中的作用

• 批准号: 10375392
• 负责人: Arnaldo Mercado-Perez
• 金额: $ 4.7万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10375392
• 关键词: Actins; Adult; Affect; Biology; Calcium; Cell Adhesion Molecules; Cells; Clinical; Co-Immunoprecipitations; Collaborations; Cytoskeleton; Data; Disease; Doctor of Philosophy; Electrophysiology (science); Elements; Enteroendocrine Cell; Epithelial; Esthesia; Exhibits; Face; Fiber; Functional disorder; Future; Gastrointestinal Diseases; Goals; Hormone secretion; Hormones; Image; Imaging Techniques; Impairment; Individual; Intestines; Ion Channel; Ion Channel Gating; Ions; Irritable Bowel Syndrome; Knowledge; Lateral; Light; Link; Measures; Mechanical Stress; Mechanics; Mediating; Membrane; Mentors; Microscopy; Molecular; Mucous Membrane; Muscle Contraction; Neurons; Organoids; Patients; Pharmacology; Physicians; Physiological; Physiology; Piezo 2 ion channel; Play; Population; Process; Proteins; Reflex action; Research Activity; Resolution; Role; Scientist; Sensory; Serotonin; Signal Transduction; Signaling Molecule; Skin; Small Interfering RNA; Specific qualifier value; Stretching; Surgeon; Testing; Tight Junctions; Tissue Model; Tissues; Touch sensation; Training; Work; arm; base; cell motility; claudin 4; clinically relevant; experience; experimental study; force sensor; knock-down; mechanotransduction; novel; optogenetics; patch clamp; receptor; response; selective expression; skills; supportive environment; targeted treatment; tool; transmission process

Project Summary/Abstract Around 15% of the US population suffers from a motility or functional GI disorder (FGID), like irritable bowel syndrome (IBS). The pathophysiology of FGIDs remains poorly understood, leading to poorly targeted treatments. Enteroendocrine cells (EECs) in the GI epithelium release signaling molecules that control many processes affected in FGID patients, like motility and secretion. A large proportion of FGID patients have abnormalities in mechanosensation. A population of mechanosensitive EECs in the GI epithelium exhibits structural and functional similarities with specialized sensory epithelia, such as light touch receptors in the skin. The mechanosensitive EECs sense physical forces and convert them into hormone release. Thus, mechanosensitive EECs are primary mechanotransducers in the GI epithelium, and they may be targets for the treatment of FGIDs. They are characterized by expression of Piezo2, an ion channel that opens in response to force. Piezo2 generates a receptor current that initiates EEC mechanotransduction. Piezo2 is not the only mechanosensitive protein in these cells: actin fibers and tight junctions are also critical in epithelial force transmission. Understanding Piezo2 localization and its functional interactions with other mechanosensitive proteins is important for understanding sensory epithelial mechanotransduction. The overall goal of this proposal is to uncover mechanisms by which mechanosensitive proteins work together in EECs to make them efficient force sensors. The hypothesis is that the actin cytoskeleton plays a critical role in Piezo2+ EEC mechanotransduction by linking the channel to other mechanosensors and directly changing channel currents. Aim 1 investigates the assembly of mechanosensory proteins in the EEC by superresolution imaging and co- immunoprecipitation studies. These experiments explore direct and indirect interactions between the mechanosensors Piezo2, actin fibers and claudin-4. Aim 2 investigates how these mechanosensors affect overall EEC mechanotransduction by tracking force-induced calcium transients and electrophysiological studies of Piezo2. These experiments explore how force is transmitted in epithelial sheets to initiate the receptor current. The results of this work are poised to bridge knowledge gaps in Piezo2+ EEC mechanotransduction, as well as inform broader mechanosensing mechanisms in sensory epithelia. The proposed work will be carried out in a supportive environment that provides cutting edge tools and expert knowledge towards achieving the specified goal, including collaborations with experts in cytoskeletal biology,and an imaging core with vast experience in the proposed imaging techniques. The proposal includes a comprehensive training plan with physician-scientist mentors, by which the PI will gain valuable skills in the study of molecular mechanotransduction on clinically relevant questions. Along with research activities, the plan also includes clinical training and shadowing activities to prepare the PI for his transition to the next stage of training as a future surgeon-scientist.

项目摘要/摘要 大约15％的美国人口患有运动或功能性GI障碍（FGID），例如肠易激 综合征（IBS）。 FGID的病理生理学仍然很少了解，导致靶向较差 治疗。 GI上皮释放信号分子中的肠内分泌细胞（EEC）控制了许多 在FGID患者中受影响的过程，例如运动和分泌。大部分FGID患者具有 机械敏化异常。胃肠道上皮中的机械敏感EEC群体 具有专门感觉上皮的结构和功能相似性，例如皮肤中的轻触摸受体。 机械敏感的EEC感知物理力并将其转化为激素释放。因此， 机械敏感的EEC是GI上皮中的主要机械转换器，它们可能是目标的目标 fGID的处理。它们的特征是表达压电2，这是一个对响应的离子通道的表达 力量。压电2产生了启动EEC机械转导的受体电流。压电不是唯一的 这些细胞中的机械敏感蛋白：肌动蛋白纤维和紧密连接在上皮力也很重要 传播。了解Piezo2定位及其与其他机械敏感的功能相互作用 蛋白质对于理解感觉上皮机械转移很重要。总体目标 建议是揭示机制在EEC中共同起作用的机制 有效的力传感器。假设是肌动蛋白细胞骨架在压电2+ EEC中起关键作用 通过将通道连接到其他机械传感器并直接更改通道电流来传递机械传输。 AIM 1通过超级分辨率成像和共同进行研究，研究了EEC中机械感觉蛋白的组装 免疫沉淀研究。这些实验探索了直接和间接相互作用 机械传感器压电2，肌动蛋白纤维和Claudin-4。 AIM 2研究了这些机械传感器如何影响 通过跟踪力诱导的钙瞬变和电生理学的总体EEC机械转传 Piezo2的研究。这些实验探讨了如何在上皮板上传输力以启动 受体电流。这项工作的结果有望弥合Piezo2+ EEC中的知识差距 机械转导，并为感觉上皮中的更广泛的机械传感机理提供信息。这 建议的工作将在提供尖端工具和专家的支持环境中进行 实现指定目标的知识，包括与细胞骨架专家的合作 生物学，以及在提出的成像技术中具有丰富经验的成像核心。该提案包括一个 与医师科学家导师的全面培训计划，PI将获得宝贵的技能 研究分子机械转导在临床相关问题上。与研究活动一起 计划还包括临床培训和阴影活动，以准备PI，以使其过渡到下一阶段 作为未来外科医生科学家的培训。