Magnesium flux compendium: Discover ligands, channels, and metabolic signals

镁通量概要：发现配体、通道和代谢信号

• 批准号: 10791996
• 负责人: MADESH MUNISWAMY
• 金额: $ 25万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-01 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10791996
• 关键词: Acinar Cell; Binding; Biochemical Reaction; Bioenergetics; Biological; Biophysics; Buffers; CRISPR/Cas technology; Cations; Cell membrane; Cell model; Cell physiology; Cells; Cellular biology; Complex; Cytosol; Deficiency Diseases; Dissociation; Endosomes; Enzymes; Equilibrium; Event; Functional disorder; Funding; Future; Homeostasis; Hormonal; Ion Channel; Laboratories; Ligands; Link; Lysosomes; Magnesium; Membrane; Metabolic; Metabolism; MgATP; Mitochondria; Mitochondrial Matrix; Molecular; Nucleic Acids; Nucleotides; Organelles; Phenotype; Physiological; Proteins; RNA interference screen; Rest; Role; Route; Shapes; Signal Pathway; Signal Transduction; Stimulus; Testing; Work; cofactor; ionization; mouse model; programs; uptake

ABSTRACT/SUMMARY Free ionized intracellular Mg2+ (iMg2+) is estimated to be in the range of 0.5–1.2 mM. In general, it is accepted that under resting conditions, the concentration of ionized cytosolic Mg2+ is `muffled' by phosphometabolites, nucleic acids and proteins. For example, ATP binds with a Kd value of 50 M-70 μM and therefore Mg2+ in the cytosol and the mitochondrial matrix is primarily complexed with ATP (Mg-ATP2-). Because of its abundance (~5 mM), ATP is considered to be the largest iMg2+ `store'. Fluctuations in free cytosolic (cMg2+) following hormonal stimuli have been touted as passive adjustments of Mg2+ dissociating from the exuberant Mg-ATP contingent and other `buffered' pools of Mg2+. Apart from iMg2+ `buffering' mechanism, Mg2+ ion channels and transporters controlling Mg2+ entry as well as efflux across the plasma membrane are thought to maintain the equilibrium of free cMg2+. Currently, several candidates are correlated to Mg2+ entry machinery (TRPM6, TRPM7, MagT1), but are still awaiting convincing biophysical and physiological evidence for such roles. The Mg2+/Na+ exchanger SLC41A1 was proposed to contribute Mg2+ efflux from the cell, whereas Mrs2 was proposed as a mitochondrial Mg2+ transporter. Very little is known about the molecular details of Mg2+ transport into/from cellular organelles like the ER, mitochondria, endosomes and lysosomes. A few studies have speculated that free [Mg2+] in the ER and mitochondria are likely to be similar to [cMg2+]. However, the temporal and spatial dynamics, let alone the biological relevance of iMg2+ mobilization, remain a mystery in cell biology. Nevertheless, Mg2+ is an essential cation controlling many biochemical reactions. Our recent work has shown that L-lactate acts as an activator that triggers a dynamic transfer of Mg2+ between the ER and mitochondria to shape bioenergetics and cellular metabolism (Cell 2020). Mechanistically, L-lactate stimulates Mg2+ release from the ER followed by Mg2+ uptake by mitochondria. The mitochondrial localized Mrs2 transporter was found to be responsible for the accumulation of Mg2+ in mitochondria. However, the L-lactate-induced ER release molecular machinery remains unidentified. I propose to identify ER Mg2+ release component, plasma membrane entry machinery and the resultant molecular signaling pathways. I will take advantage of unbiased RNAi screen and targeted CRISPR/Cas9 editing approaches to answer these mysteries in the Mg2+ signaling field. Identification of these molecular machineries would aid in our understanding of iMg2+ dynamics and the cause-effect relationships that exist between iMg2+ flux and cellular processes. Additionally, I will test and define the Mg2+-dependent signaling events based on the cellular and mouse model phenotypes. It is thrilling to define the molecular link between cellular Mg2+ homeostasis and physiological function. Our identification and characterization of the Mg2+ flux components will further investigate how, and if, these signaling routes impinge on the pathophysiology of a growing number of Mg2+ deficiency diseases in humankind. Overall, the R35/MIRA funding will support the testing of this unconventional hypothesis and my laboratory will address these major mysteries in the near future.

摘要/总结 游离离子化细胞内 Mg2+ (iMg2+) 估计在 0.5–1.2 mM 范围内。一般来说，这是可以接受的。 在静息条件下，离子化胞质 Mg2+ 的浓度被磷酸代谢物“抑制”， 例如，ATP 的结合 Kd 值为 50 μM-70 μM，因此 Mg2+ 会与核酸和蛋白质结合。 由于其丰度（~5），细胞质和线粒体基质主要与 ATP (Mg-ATP2-) 复合。 mM)，ATP 被认为是激素后游离胞质 (cMg2+) 中最大的 iMg2+“储存”。 刺激被认为是 Mg2+ 从旺盛的 Mg-ATP 队伍中解离出来的被动调节 和其他 Mg2+“缓冲”池 除了 iMg2+“缓冲”机制之外，Mg2+ 离子通道和转运蛋白。 控制 Mg2+ 进入以及穿过质膜的流出被认为可以维持平衡 目前，有几个候选者与 Mg2+ 进入机制相关（TRPM6、TRPM7、MagT1），但是 仍在等待 Mg2+/Na+ 交换剂的这种作用的生物物理学和令人信服的生理学证据。 SLC41A1 被认为有助于从细胞中流出 Mg2+，而 Mrs2 被认为是线粒体 关于 Mg2+ 转运入/转运出细胞器的分子细节知之甚少。 一些研究推测 ER 中存在游离 [Mg2+]。 和线粒体很可能与[cMg2+]相似，但是时间和空间动力学更不用说了。 iMg2+ 动员的生物学相关性在细胞生物学中仍然是一个谜，然而，Mg2+ 是必不可少的。 我们最近的工作表明，L-乳酸作为一种激活剂，可以控制许多生化反应。 触发内质网和线粒体之间 Mg2+ 的动态转移，从而塑造生物能学和细胞学 新陈代谢 (Cell 2020) 从机制上讲，L-乳酸刺激内质网释放 Mg2+，然后吸收 Mg2+。 线粒体定位的 Mrs2 转运蛋白被发现负责积累。 然而，L-乳酸诱导的 ER 释放分子机制仍不清楚。 我建议确定 ER Mg2+ 释放成分、质膜进入机制以及由此产生的结果 我将利用无偏见的 RNAi 筛选和靶向 CRISPR/Cas9 编辑。 解开 Mg2+ 信号传导领域的这些谜团的方法 识别这些分子机器。 有助于我们理解 iMg2+ 动力学以及 iMg2+ 通量之间存在的因果关系 此外，我将根据以下内容测试和定义 Mg2+ 依赖性信号传导事件。 细胞和小鼠模型表型的确定令人兴奋。 我们对 Mg2+ 通量成分的鉴定和表征将 进一步研究这些信号通路如何以及是否影响越来越多的病理生理学 总体而言，R35/MIRA 资金将支持这一测试。 非常规假设和我的实验室将在不久的将来解决这些重大谜团。

项目成果

Negative modulation of mitochondrial calcium uniporter complex protects neurons against ferroptosis.

• DOI: 10.1038/s41419-023-06290-1
• 发表时间: 2023-11-25
• 期刊: CELL DEATH & DISEASE
• 影响因子: 9
• 作者: Marmolejo-Garza, Alejandro;Krabbendam, Inge E.;Luu, Minh Danh Anh;Brouwer, Famke;Trombetta-Lima, Marina;Unal, Osman;O'Connor, Shane J.;Majernikova, Nad'a;Elzinga, Carolina R. S.;Mammucari, Cristina;Schmidt, Martina;Madesh, Muniswamy;Boddeke, Erik;Dolga, Amalia M.
• 通讯作者: Dolga, Amalia M.