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Identification of CALHM proteins as ion channels

CALHM 蛋白作为离子通道的鉴定

基本信息

批准号：
10044119
负责人：
James Kevin FOSKETT
金额：
$ 16.21万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-08-15 至 2021-11-14
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10044119
关键词：
Action Potentials Binding Biochemical Biological Assay Biological Process Brain Caenorhabditis elegans Caliber Cell Line Cell membrane Cells Chromosome 10 Chromosome 6 Chromosomes Connexins Drosophila genus Electrophysiology (science)Evolution Family G-Protein-Coupled Receptors Gene Family Genes Genetic Goals Grant Homologous Gene Human Ion Channel Ion Channel Gating Ion Channel Protein Knockout Mice Knowledge Lead Mammalian Cell Membrane Membrane Proteins Molecular Mus Nerve Neuraxis Neurotransmitters Oocytes Perception Permeability Physiological Physiology Property Proteins Regulation Research Role Sequence Homology Signal Transduction Structure Taste Bud Cell Taste Buds Taste Perception Testing Therapeutic Tissues Type II Epithelial Receptor Cell Vertebrates Work Xenopus oocyte Yeasts extracellular insight member novel novel therapeutics optical imaging response success voltage

项目摘要

PROJECT SUMMARY The goal of our proposed studies is to determine whether IDG-eligible CALHM4, CALHM5 and CALHM6 are ion channels or ion channel subunits. The CALHM (formerly FAM26) gene family is comprised of six homologs. We previously discovered the molecular mechanisms and physiological roles of CALHM1 and CALHM3 as novel ion channels. CALHM1 encodes a membrane protein expressed throughout the brain and in taste buds that lacks significant homology to other proteins, although five homologs have been identified with 20-44% sequence similarity, and CALHM proteins are conserved across species. We identified CALHM1 as a pore- forming subunit of an ion channel with a large pore-diameter and gating regulation by voltage and extracellular Ca2+ (Ca2+o). We discovered that CALHM1 is essential for perceptions of sweet, bitter and umami tastes by type II taste-bud cells, since CALHM1-knockout mice cannot perceive these tastants. We identified the essential role of CALHM1 by discovering that it is a voltage-gated ATP-permeable ion channel, and that tastant-evoked Na+ action potentials trigger ATP release as a neurotransmitter through CALHM1-associated channels to transduce taste information from the periphery to the central nervous system. We further discovered that whereas CALHM3 expression does not confer a novel ion channel, it is an essential component of the native voltage-gated ATP-release channel, contributing as a pore-forming subunit with CALHM1 to create a hetero-hexameric ATP-release channel in type II cells. Genetic deletion of CALHM3 also eliminates the ability of mice to perceive sweet, bitter and umami substances. These results suggest that other CALHM homologs are also ion channels or ion channel subunits, but there is no information regarding their molecular function. We hypothesize that CALHM4, CALHM5 and CALHM6 are ion channels, and furthermore that the CALHM family represents a family of ATP-release channels the contributes to purinergic signaling throughout the body. We will exploit our insights and technical approaches used to elucidate functions of CALHM1 and CALHM3 to determine whether CALHM4, CALHM5 and CALHM6 are ion channels or ion channel subunits. We will employ electrophysiology, optical imaging, ATP-release assays and biochemical approaches to test this overarching hypothesis. The impact of our research is expected to be the discovery of the functions of CALHM4, CALHM5 and CALHM6. Given the identification of CALHM1 and CALHM3 as ion- channel subunits, our findings are expected to provide information about whether CALHMs are a family of ion channel proteins. Identification of novel ion channels is expected to lead to new understanding of cell and tissue physiology, as our previous work in taste buds and taste perception did with the discoveries of the functions of CALHM1 and CALHM3. Accordingly, identification of the roles of the other CALHM homologs may inform new physiological insights and therapeutic opportunities.

项目概要我们提出的研究的目标是确定符合 IDG 条件的 CALHM4、CALHM5 和 CALHM6 是否离子通道或离子通道亚基。 CALHM（以前称为 FAM26）基因家族由六个同源物组成。我们之前发现了 CALHM1 和 CALHM3 的分子机制和生理作用：新型离子通道。 CALHM1 编码一种在整个大脑和味蕾中表达的膜蛋白与其他蛋白质缺乏显着的同源性，尽管已鉴定出 5 种同源性为 20-44% 序列相似性，并且 CALHM 蛋白在物种间是保守的。我们将 CALHM1 鉴定为孔形成具有大孔径的离子通道亚基并通过电压和细胞外进行门控调节 Ca2+ (Ca2+o)。我们发现 CALHM1 对于甜味、苦味和鲜味的感知至关重要 II 型味蕾细胞，因为 CALHM1 敲除小鼠无法感知这些促味剂。我们确定了 CALHM1 的重要作用是发现它是一个电压门控 ATP 渗透性离子通道，并且促味剂诱发的 Na+ 动作电位通过 CALHM1 相关触发 ATP 作为神经递质的释放将味觉信息从外周神经系统传递到中枢神经系统的通道。我们进一步发现虽然 CALHM3 表达并不赋予新的离子通道，但它是一种重要的离子通道天然电压门控 ATP 释放通道的组成部分，作为孔形成亚基起作用 CALHM1 在 II 型细胞中创建异源六聚体 ATP 释放通道。 CALHM3 的基因删除也消除小鼠感知甜味、苦味和鲜味物质的能力。这些结果表明其他 CALHM 同系物也是离子通道或离子通道亚基，但没有关于它们的信息分子功能。我们假设 CALHM4、CALHM5 和 CALHM6 是离子通道，并且进一步 CALHM 家族代表 ATP 释放通道家族，有助于嘌呤能信号传导遍布全身。我们将利用我们的见解和技术方法来阐明 CALHM1 和 CALHM3 确定 CALHM4、CALHM5 和 CALHM6 是离子通道还是离子通道子单元。我们将采用电生理学、光学成像、ATP 释放测定和生物化学检验这一总体假设的方法。我们研究的影响预计将是发现 CALHM4、CALHM5 和 CALHM6 的功能。鉴于 CALHM1 和 CALHM3 被鉴定为离子通道亚基，我们的研究结果有望提供有关 CALHM 是否是离子家族的信息通道蛋白。新型离子通道的鉴定有望带来对细胞和组织生理学，正如我们之前在味蕾和味觉方面的工作所发现的那样 CALHM1 和 CALHM3 的功能。因此，其他 CALHM 同系物的作用的鉴定可能提供新的生理见解和治疗机会。