Functional architecture of intracellular Ca2+ signals

细胞内 Ca2 信号的功能结构

• 批准号: 9021183
• 负责人: JONATHAN S MARCHANT
• 金额: $ 12.49万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9021183
• 关键词: Affinity Chromatography; Age; Architecture; Attention; Behavior; Binding; Binding Sites; Calcium; Calcium Channel; Calculi; Cell physiology; Cells; Chemicals; Chronic; Complex; Congenital Heart Defects; Coupled; Coupling; Cues; Data; Data Set; Disease; Endoplasmic Reticulum; Event; Family; Functional disorder; Genetic; Goals; Grant; Gray unit of radiation dose; Health; Homeostasis; Inositol; Ion Channel; Kinetics; Knowledge; Left; Life; Ligands; Link; Lysosomes; Malignant Neoplasms; Mass Spectrum Analysis; Membrane; Molecular; NAADP; Nerve Degeneration; Neurodegenerative Disorders; Neurons; Organelles; Outcome; Pattern; Persons; Pharmaceutical Preparations; Physiological; Process; Property; Proteins; Proteomics; Regulation; Reporting; Role; Second Messenger Systems; Signal Pathway; Signal Transduction; Site; Stimulus; Stroke; System; Testing; Work; base; cell type; cellular imaging; design; insight; interest; nervous system disorder; neuron loss; novel; overexpression; rab GTP-Binding Proteins; receptor; response; second messenger; sigma receptors; sigma-2 receptor; spatiotemporal; therapeutic target; trafficking

DESCRIPTION (provided by applicant): Increases in Ca2+ resulting from activation of intracellular Ca2+ channels regulate many physiological events. Dysfunction of these Ca2+ channels is involved in neurodegenerative and neurological disease, as well as exocrine, cardiac abnormalities and cancer. To appreciate how cell functions are controlled by Ca2+ signals, and how pathological cues subvert their function, we must understand how both the distribution and the properties (the 'functional architecture') of intracellular Ca2+ channels controls the patterning of Ca2+ signals. Here, we investigate the structural and functional coupling between two intracellular Ca2+-permeable channels that are activated by distinct second messengers: (i) nicotinic acid adenine dinucleotide phosphate (NAADP) which activates the recently discovered two-pore channels (TPCs) within endolysosomes, and (ii) inositol trisphosphate (IP3), which activates IP3 receptors (IP3Rs) in the endoplasmic reticulum. Despite localization in separate organelles, the activity of these Ca2+ channels is intimately related: a functional coupling garnering increasing attention in neurodegenerative disorders involving lysosomal proliferation. Here, by defining the mammalian TPC interactome we provide insight into two key unknowns: (i) how the functional architecture between TPCs and IP3Rs is established and (ii) the molecular identity of the NAADP receptor (NAADP-R, part of the TPC complex). Both are key pieces of knowledge for designing new drugs to modify this coupling. Our six person team, combining chemical, proteomic, molecular and live cell imaging expertise, will resolve: (1) Whether TPCs are Rab effectors? TPCs associate with a clade of Rab GTPases. We will define how TPC channels act as a node for coupling Ca2+ signaling to endolysosomal trafficking and fusion events. (2) How TPC/IP3R activity is coordinated between discrete organelles. We will use novel molecular insight from the TPC interactome to interrogate the functional architecture of TPCs/IP3Rs at membrane contact sites, and uncover how dysregulation triggers lysosomal proliferation. (3) Identify the NAADP-R. We have designed and optimized a novel bifunctional photoprobe to unmask the NAADP-R within the TPC interactome. This is a key roadblock, hampering knowledge of TPC activation. The broad significance of this work is in understanding principles controlling ion channel dynamics, and thereby the kinetics of Ca2+ signals that control compartmentalized cellular and system-levels outcomes. Such data will aid our understanding of the role of ubiquitous Ca2+ signaling pathways in health and disease.

描述（由申请人提供）：由于细胞内Ca2+通道的激活而导致的Ca2+增加，可调节许多生理事件。这些CA2+通道的功能障碍参与神经退行性和神经系统疾病以及外分泌，心脏异常和癌症。要欣赏细胞功能如何由Ca2+信号控制，以及病理提示如何颠覆其功能，我们必须了解细胞内Ca2+通道的分布和属性（“功能架构”）如何控制Ca2+信号的模式。在这里，我们研究了两种细胞内Ca2+可渗透的通道之间的结构和功能耦合，这些通道被不同的第二使者激活：（i）烟酸腺苷二核苷酸磷酸盐（NAADP）激活最近在最近发现的两孔通道（TPC）内的内溶解体和（II）ii siplosol osipol osipol osipol osipol osipol ossitol trissitol ossipolol ossitol ossiitol ossitol ossitol ospossitol ossitol ossiitol ossiitol trissipoloslosoll ospossitol， （IP3R）在内质网中。尽管在单独的细胞器中定位，但这些Ca2+通道的活性与之密切相关：一种功能性耦合，吸引了涉及溶酶体增殖的神经退行性疾病的越来越注意。在这里，通过定义哺乳动物TPC相互作用组，我们可以洞悉两个关键未知数：（i）如何建立TPCS和IP3RS之间的功能架构，以及（ii）NAADP受体的分子身份（NaADP-R（NaADP-R）的分子身份（NAADP-R，TPC复合物的一部分）。两者都是设计新药来修改这种耦合的关键知识。我们的六人团队结合了化学，蛋白质组学，分子和活细胞成像专业知识，将解决：（1）TPC是否是RAB效应器？ TPC与Rab GTPases的进化枝相连。我们将定义TPC通道如何充当耦合Ca2+信号传导与内侧溶性交通和融合事件的节点。 （2）如何在离散细胞器之间协调TPC/IP3R活动。我们将使用TPC Interactome的新型分子见解来询问TPC/IP3RS在膜接触位点的功能结构，并发现失调症如何触发溶酶体增殖。 （3）识别NAADP-R。我们已经设计并优化了一种新型的双功能光探针，以揭示TPC Interactome中的NAADP-R。这是一个关键的障碍，阻碍了TPC激活的知识。这项工作的广泛意义在于理解控制离子通道动力学的原理，从而控制控制分隔的细胞和系统级别结果的Ca2+信号的动力学。这些数据将有助于我们理解无处不在的CA2+信号通路在健康和疾病中的作用。