Function and regulation of TRPM7 Mg2+-inhibited cation channels

TRPM7 Mg2 抑制的阳离子通道的功能和调节

• 批准号: 7981134
• 负责人: Juliusz Ashot Kozak
• 金额: $ 25.07万
• 依托单位: WRIGHT STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2014-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7981134
• 关键词: 1-Phosphatidylinositol 3-Kinase; Alkalinization; Antigens; Attention; Autoimmune Process; Autoimmunity; Biochemical; Calcium; Calcium ion; Cations; Cell Line; Cell membrane; Cells; Characteristics; Chemosensitization; Clonal Expansion; Dendritic Cells; Dependence; Detection; Drosophila genus; Drosophila melanogaster; Drug Delivery Systems; Effector Cell; Electrophysiology (science); Enzymes; Fura-2; Gene Expression; Gene Targeting; Generations; Genetic; Genetic Engineering; Genetic Transcription; Homologous Gene; Hour; Human; Hydrolysis; Image; Immune; Immune response; Immune system; Immunologic Deficiency Syndromes; Immunosuppression; Ion Channel; Ions; Lipids; Lymphocyte Activation; Mammalian Cell; Measures; Mediating; Membrane Potentials; Metabolism; Molecular; Mus; Organ; Organ Transplantation; Orthologous Gene; Pathway interactions; Phosphatidylinositols; Phospholipase C; Phospholipids; Process; Proliferating; Property; Publishing; RNA Interference; Receptor Signaling; Recombinants; Regulation; Research; Reverse Transcriptase Polymerase Chain Reaction; Role; Signal Transduction; Specificity; Surface; System; T-Cell Activation; T-Cell Receptor; T-Lymphocyte; Testing; base; cell type; immunoregulation; macrophage; new therapeutic target; novel; patch clamp; prevent; public health relevance

DESCRIPTION (provided by applicant): T-lymphocyte activation is accompanied by calcium ion (Ca2+) mobilization, alkalinization, blast transformation (enlargement) and membrane potential changes. Ca2+ in the cell is elevated when antigen recognition initiates a biochemical cascade, leading to the release of this ion from intracellular stores and ensuing influx of Ca2+ through ion channels in the plasma membrane. The calcium entry process is critical for efficient T- cell gene expression and clonal expansion, necessary steps for the adaptive immune response. Even though store-operated Ca2+ entry mechanism in T lymphocytes has received most attention, other Ca2+ -permeable ion channels are present in the T cell, which can provide sustained, hour-long Ca2+ elevations required for transcription. One such channel is TRPM7. It is highly expressed in T cells and is activated by alkalinization of the cell interior and also inhibited by Mg2+. The central hypothesis of the proposed research is that TRPM7 channels function as a Ca2+ entry pathway during T-cell activation and proliferation. The specific aims to test this hypothesis are: 1) to characterize regulation of native T cell and recombinant TRPM7 by plasma membrane PI(3,4,5) phospholipid 2) to characterize regulation of native T cell and recombinant TRPM7 by intracellular pH. 3) to determine if TRPM7 can provide a pathway for a sustained Ca2+ influx during T cell activation. 4) to establish a novel system for studying mammalian TRPM7 channels in isolation that employs Drosophila melanogaster cell line, which lack native TRPM7 homologs. Single-cell Ca2+ and pH imaging and electrophysiology will be employed for the detection of specific ion channel activity. Recombinant DNA technology will be used for heterologous expression of recombinant TRPM7 channels in non-immune and Drosophila cells for detailed study of their characteristics and regulation. The significance of the proposed research is that detailed understanding of the Ca2+ entry components in T cell activation process will likely identify novel drug targets for immunomodulation. PUBLIC HEALTH RELEVANCE: Cells comprising the human adaptive immune system require prolonged elevations of intracellular calcium ion concentration. Genetic disturbances in calcium handling result in immunodeficiency. Conversely, immunosuppression is widely used in organ transplantation and autoimmune conditions. The current proposal aims to elucidate the molecular mechanisms of calcium handling in immune cells, potentially providing new therapeutic targets for immunosuppression and immunomodulation.

描述（由申请人提供）：t淋巴细胞活化伴随着钙离子（Ca2+）动员、碱化、胚转化（扩大）和膜电位变化。当抗原识别启动生化级联时，细胞中的Ca2+升高，导致细胞内储存的离子释放，并随后通过质膜中的离子通道流入Ca2+。钙离子进入过程是T细胞基因高效表达和克隆扩增的关键，是适应性免疫应答的必要步骤。尽管存储操作的Ca2+进入机制在T淋巴细胞中受到了最多的关注，其他Ca2+渗透离子通道存在于T细胞中，可以提供持续的，转录所需的长达一小时的Ca2+升高。其中一个通道是TRPM7。它在T细胞中高度表达，被细胞内部碱化激活，也被Mg2+抑制。该研究的中心假设是TRPM7通道在t细胞活化和增殖过程中作为Ca2+进入途径发挥作用。验证这一假设的具体目的是：1)表征质膜PI（3,4,5）磷脂对天然T细胞和重组TRPM7的调节作用；2)表征细胞内ph对天然T细胞和重组TRPM7的调节作用；3)确定TRPM7是否可以为T细胞活化过程中持续的Ca2+内流提供途径。4)利用缺乏天然TRPM7同源物的果蝇（Drosophila melanogaster）细胞系，建立新的哺乳动物TRPM7通道分离研究体系。单细胞Ca2+和pH成像和电生理学将用于检测特定的离子通道活性。利用重组DNA技术在非免疫细胞和果蝇细胞中异种表达重组TRPM7通道，详细研究其特性和调控。这项研究的意义在于，对T细胞活化过程中Ca2+进入成分的详细了解将有可能确定免疫调节的新药物靶点。