Mechanisms and Architecture of Endo-lysosomal Ca2+ Signalling

内溶酶体 Ca2 信号传导的机制和结构

• 批准号: BB/T01640X/1
• 负责人: A Galione
• 金额: $ 103.31万
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2020
• 资助国家: 英国
• 起止时间: 2020 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FT01640X%2F1
• 关键词: Mechanisms; Architecture; Endo; lysosomal; Ca2+

Ca2+ serves an essential signal within every cell. Ca2+ levels inside cells are very low, partly because it is sequestered inside Ca2+-storing compartments. In response to cell stimuli, Ca2+ is released from these stores by opening resident ion channels to activate detector proteins ('decoders') and change cell processes. The increase in intracellular Ca2+ is a universal signal in virtually all cells types e.g. for fertilization, muscle contraction, nerve impulses, gene expression.The largest and best understood Ca2+ store is the endoplasmic reticulum (ER) which contains millimolar Ca2+. ER Ca2+ channels, IP3 receptors, are activated by a second messenger, IP3, which is synthesised in response to cell stimuli (e.g. hormones, neurotransmitters, antibodies, cell contact). IP3 synthesis, IP3R activation and Ca2+ release occur rapidly upon stimulation. However, the ER is not the only Ca2+ store and we discovered that small acidic vesicles (including endosomes and lysosomes) are important Ca2+ stores, but with their own unique second messenger (NAADP) and Ca2+ channels (TPCs), analogous to IP3/IP3Rs. This axis forms our focus.Although they are better known as cellular waste-bins, endo-lysosomes are emerging as dynamic signalling hubs, integrating and delivering signals in response to the environment. A major endo-lysosomal signal is Ca2+. Many stimuli couple to endo-lysosomal Ca2+ release as a transduction pathway: depending on the stimulus, cells synthesise the messenger, nicotinic acid adenine dinucleotide phosphate, NAADP, which opens TPCs (two-pore channels) expressed on endo-lysosomes. These are Ca2+-permeable and elevate cytosolic Ca2+. However, each vesicle is small so the limited amount of Ca2+ that is released generates local Ca2+ 'nanodomains' (a locally high concentration, restricted in space).Why does the cell contain different Ca2+ stores? The answer is that Ca2+ does not increase uniformly in the cytosol but rather is delivered discretely where it is needed. Different stores therefore deliver Ca2+ to different targets and different downstream physiology. We find endo-lysosomes 'pair-up' with their own unique detectors via highly localized and privileged conversations, for which the ER Ca2+ store cannot substitute. Therefore, each cell stimulus selects the appropriate Ca2+ sources for its downstream physiology.However, it is unclear how these essential endo-lysosomal Ca2+ signals are generated and decoded which therefore forms our focus. The NAADP/TPC axis is poorly defined in terms of targets. A further complexity is that endo-lysosomes are small, heterogeneous, motile and exquisitely positioned, interacting physically/functionally with other organelles in specialized junctions (e.g. with ER, mitochondria).Given the multiplicity of inputs/outputs, our aim is to understand how endo-lysosomes establish and regulate these 'private' local Ca2+ conversations with targets (proteins, organelles), thereby solving the Ca2+-specificity conundrum. Clearly, targets must closely associate with endo-lysosomes in order to detect the local 'Ca2+ plume' that emanates from TPCs. We aim to understand how targets are brought to endo-lysosomes or, conversely, how endo-lysosomes are dynamically brought to targets. We will identify how, when and where targets and decoders associate with TPCs. The placement and motility of endo-lysosomes is crucial for the physiology and we also will test whether there is a specialist sub-class of endo-lysosomes for different Ca2+ signalling roles.Potential Benefits & Applications. Standing at the crossroads of multiple processes, defining endo-lysosomal Ca2+ signals will:(a) illuminate basic science (Ca2+ signals, signal compartmentation, endo-lysosomal biology, organelle/membrane dynamics, Ca2+-decoding);(b) provide new tools to the broader cell biology community (reporters, pharmacology);(c) strengthen the case for organelle ion channels as new drug targets.

Ca2+ 在每个细胞内提供重要信号。细胞内的 Ca2+ 水平非常低，部分原因是它被隔离在 Ca2+ 储存室中。为了响应细胞刺激，Ca2+ 通过打开驻留离子通道从这些储存中释放出来，激活检测蛋白（“解码器”）并改变细胞过程。细胞内 Ca2+ 的增加几乎是所有细胞类型（例如细胞）的普遍信号。用于受精、肌肉收缩、神经冲动、基因表达。最大且最了解的 Ca2+ 储存是内质网 (ER)，其中含有毫摩尔 Ca2+。 ER Ca2+ 通道（IP3 受体）由第二信使 IP3 激活，IP3 是根据细胞刺激（例如激素、神经递质、抗体、细胞接触）而合成的。 IP3 合成、IP3R 激活和 Ca2+ 释放在刺激后迅速发生。然而，ER 并不是唯一的 Ca2+ 储存，我们发现小的酸性囊泡（包括内体和溶酶体）是重要的 Ca2+ 储存，但具有自己独特的第二信使 (NAADP) 和 Ca2+ 通道 (TPC)，类似于 IP3/IP3R 。这个轴构成了我们的焦点。虽然内溶酶体更被称为细胞废物箱，但它们正在成为动态信号中心，集成和传递信号以响应环境。主要的内溶酶体信号是 Ca2+。许多刺激与内溶酶体 Ca2+ 释放耦合，作为转导途径：根据刺激，细胞合成信使烟酸腺嘌呤二核苷酸磷酸 (NAADP)，打开内溶酶体上表达的 TPC（双孔通道）。它们具有 Ca2+ 渗透性，可升高胞质 Ca2+。然而，每个囊泡都很小，因此释放的 Ca2+ 数量有限，会产生局部 Ca2+“纳米域”（局部高浓度，受空间限制）。为什么细胞含有不同的 Ca2+ 储备？答案是 Ca2+ 不会在细胞质中均匀增加，而是离散地输送到需要的地方。因此，不同的存储将 Ca2+ 输送到不同的目标和不同的下游生理。我们发现内溶酶体通过高度本地化和特权对话与它们自己独特的检测器“配对”，这是 ER Ca2+ 存储无法替代的。因此，每种细胞刺激都会为其下游生理学选择合适的 Ca2+ 来源。然而，目前尚不清楚这些重要的内溶酶体 Ca2+ 信号是如何产生和解码的，这也是我们关注的焦点。 NAADP/TPC 轴的目标定义不明确。进一步的复杂性是，内溶酶体体积小、异质、活动且位置精巧，与特殊连接处的其他细胞器（例如与内质网、线粒体）进行物理/功能相互作用。鉴于输入/输出的多样性，我们的目标是了解如何内溶酶体建立并调节这些“私人”局部 Ca2+ 与目标（蛋白质、细胞器）的对话，从而解决 Ca2+ 特异性难题。显然，目标必须与内溶酶体密切相关，才能检测 TPC 发出的局部“Ca2+ 羽流”。我们的目标是了解靶标如何被带到内溶酶体，或者相反，内溶酶体如何动态地被带到靶标。我们将确定目标和解码器如何、何时、何地与 TPC 关联。内溶酶体的位置和运动性对于生理学至关重要，我们还将测试是否存在针对不同 Ca2+ 信号传导作用的内溶酶体的专门子类。潜在益处和应用。站在多个过程的十字路口，定义内溶酶体 Ca2+ 信号将：(a) 阐明基础科学（Ca2+ 信号、信号区室、内溶酶体生物学、细胞器/膜动力学、Ca2+ 解码）；(b) 提供新工具更广泛的细胞生物学界（记者、药理学）；(c) 强化细胞器离子通道作为新药物靶点的理由。

项目成果

A cellular protection racket: How lysosomal Ca2+ fluxes prevent kidney injury.

细胞保护球拍：溶酶体 Ca2 通量如何预防肾损伤。

• DOI: 10.1016/j.ceca.2020.102328
• 发表时间: 2021
• 期刊: Cell calcium
• 影响因子: 4
• 作者: Galione A

Choreographing endo-lysosomal Ca2+ throughout the life of a phagosome.

• DOI: 10.1016/j.bbamcr.2021.119040
• 发表时间: 2021-04
• 期刊: Biochimica et biophysica acta. Molecular cell research
• 作者: A. Morgan;Lianne C. Davis;A. Galione

Endolysosomal TPCs regulate social behavior by controlling oxytocin secretion.

• DOI: 10.1073/pnas.2213682120
• 发表时间: 2023-02-14
• 期刊: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• 影响因子: 11.1
• 作者: Martucci, Lora L.;Launay, Jean-Marie;Kawakami, Natsuko;Sicard, Cecile;Desvignes, Nathalie;Dakouane-Giudicelli, Mbarka;Spix, Barbara;Tetu, Maude;Gilmaire, Franck -Olivier;Paulcan, Sloane;Callebert, Jacques;Vaillend, Cyrille;Bracher, Franz;Grimm, Christian;Fossier, Philippe;de la Porte, Sabine;Sakamoto, Hirotaka;Morris, John;Galione, Antony;Granon, Sylvie;Cancela, Jose-Manuel
• 通讯作者: Cancela, Jose-Manuel