Development of Multiplex Multiomic Mass Spectrometry Methods for Probing the Response to Copper Toxicity in the Blue Crab, Callinectes sapidus

开发多重多组学质谱方法来探测蓝蟹（Callinectes sapidus）对铜毒性的反应

• 批准号: 10404908
• 负责人: Christopher Sauer
• 金额: $ 2万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2022-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10404908
• 关键词: Acute; Address; Agriculture; Amines; Anemia; Animal Model; Biochemical; Biological; Blue Crab; Brain; Chelating Agents; Chronic; Copper; Crabs; Crustacea; Custom; Defense Mechanisms; Development; Disease; Ensure; Event; Excretory function; Exposure to; Gills; Gland; Glutathione; Goals; Harvest; Health; Heavy Metals; Hemolymph; Hepatopancreas; Homeostasis; Hypoxia; Industrialization; Injections; Ions; Knowledge; Label; Leucine; Link; Liquid substance; Liver; Liver diseases; Location; Lysine; Masks; Mass Spectrum Analysis; Measures; Mediating; Mental disorders; Metabolism; Metallothionein; Methods; Nervous system structure; Neurodegenerative Disorders; Neuropeptides; Neurosecretory Systems; Nutrient; Organ; Organism; Oxidation-Reduction; Oxidative Stress; Peptides; Pericardial body location; Physiological; Play; Proteins; Reaction; Reporter; Research; Resolution; Role; Running; Saline; Sampling; Scanning; Signal Pathway; Signaling Molecule; Sinus; Stress; Structure; Techniques; Time; Tissue Extracts; Tissues; Toxic effect; Variant; Vertebrates; biological adaptation to stress; design; environmental stressor; enzyme activity; experimental study; improved; in vivo; insight; instrumentation; liver injury; mass analyzer; metal poisoning; multiple omics; nervous system disorder; protein structure; response; tandem mass spectrometry

Abstract Copper is an essential nutrient with critical roles in protein structure and enzyme activity but becomes toxic at elevated levels due to its ability to participate in redox reactions. Copper toxicity is a growing concern as environmental copper concentrations increase due to agricultural and industrial runoff and poorly managed wastewater. The dysregulation of copper homeostasis has been linked to many diseases including anemia, liver damage, and neurodegenerative diseases, but the exact mechanisms are not fully understood. The biochemical response to copper toxicity has been shown to involve neuropeptides, but the full suite of relevant neuropeptides and their expression changes are difficult to characterize due to the complexity of the nervous system. Crustaceans provide a highly relevant model organism with a simple, well-characterized nervous system, facilitating meaningful analysis of environmental stress. Neuropeptides remain challenging to study, however, due to their structural diversity and low in vivo concentrations. As a result, mass spectrometry (MS) has become the preferred method of analyzing neuropeptides. MS is highly sensitive and selective, capable of providing both structural and quantitative information, and requires no prior knowledge of the analytes in the sample. This enables the analyses of not only neuropeptides, but also the chelating molecules responsible for maintaining homeostasis, such as glutathione and metallothionein proteins. To thoroughly probe the biochemical and physiological response to copper toxicity, I propose to: 1) develop and optimize high-throughput, multiplexed mass spectrometry techniques to quantify neuropeptides, 2) globally profile the neuropeptides involved in the stress response to both chronic and acute copper toxicity, and 3) analyze the expression changes of copper chelating molecules in the gills, hemolymph, and hepatopancreas using top-down MS methods. The proposed research will provide a deeper understanding of the signaling molecules and biochemical defense mechanisms involved in responding to excess copper, thereby providing a better understanding of how environmental stressors (such as heavy metals) can have profound impacts on health. Moreover, understanding the stress response can provide unique insights into the many diseases linked to copper toxicity and dysregulation of copper homeostasis.

摘要 铜是一种必需的营养素，在蛋白质结构和酶活性中起着关键作用， 由于其参与氧化还原反应的能力，在升高的水平下变得有毒。铜 随着环境铜浓度的增加， 农业和工业径流以及管理不善的废水。铜的失调 体内平衡与许多疾病有关，包括贫血，肝损伤， 神经退行性疾病，但确切的机制尚未完全了解。的 对铜毒性的生化反应已被证明涉及神经肽，但完整的 相关神经肽组及其表达变化由于 神经系统的复杂性。甲壳类动物提供了高度相关的模式生物 具有简单的，特征良好的神经系统，便于有意义的分析 环境压力然而，神经肽的研究仍然具有挑战性，因为它们 结构多样性和低体内浓度。因此，质谱法（MS）具有 成为分析神经肽的首选方法。MS是高度灵敏和选择性的， 能够提供结构和定量信息，并且不需要事先 了解样品中的分析物。这使得不仅能够分析神经肽， 还有负责维持体内平衡的螯合分子，如谷胱甘肽 和金属硫蛋白。为了彻底探测生物化学和生理反应 铜毒性，我建议：1）开发和优化高通量，多路复用质量 光谱技术来定量神经肽，2）全面分析神经肽 参与慢性和急性铜中毒的应激反应，以及3）分析 铜螯合分子在鳃、血淋巴和肝胰腺中的表达变化 使用自上而下的MS方法。拟议的研究将提供更深入的了解 信号分子和生化防御机制参与响应过量 铜，从而更好地了解环境压力源（如 重金属）对健康有深远的影响。此外，了解压力 反应可以提供独特的见解，许多疾病与铜中毒， 铜稳态失调。