The role of ZIP12 in zinc homeostasis and associated neurodegenerative pathologies

ZIP12 在锌稳态和相关神经退行性疾病中的作用

• 批准号: 10452802
• 负责人: Robert Edward Dempski
• 金额: $ 22.74万
• 依托单位: WORCESTER POLYTECHNIC INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10452802
• 关键词: Adult; Alzheimer' s Disease; Alzheimer' s disease brain; Binding Proteins; Brain; CRISPR/Cas technology; Carrier Proteins; Cations; Cell Line; Cells; Code; Cytoplasm; Development; Dimerization; Disease; Fostering; Functional disorder; Genes; Genetic Polymorphism; Health; Homeostasis; Human; Incidence; Inductively Coupled Plasma Mass Spectrometry; Institutes; Intestines; Ion Channel; Ion Channel Gating; Ion Transport; Iron; Kinetics; Knock-out; Knowledge; Late Onset Alzheimer Disease; Ligands; Link; Longevity; Maintenance; Mass Spectrum Analysis; Measures; Mediating; Membrane; Membrane Proteins; Memory; Mental Depression; Metabolism; Metals; Micronutrients; Mission; Modeling; Molecular; Molecular Chaperones; Molecular Sieve Chromatography; Morphology; Mutate; Mutation; Nerve Degeneration; Neuraxis; Neurodegenerative Disorders; Neurons; Neuropathy; Pathology; Play; Property; Proteins; Proteome; Public Health; Research; Role; Schizophrenia; Signal Transduction; Single Nucleotide Polymorphism; Structure; Surface; Synapses; Testing; Touch sensation; Transition Elements; United States National Institutes of Health; Variant; Zinc; base; caudate nucleus; experience; experimental study; improved; innovation; malformation; mind control; neonatal brain development; nervous system disorder; neuroblastoma cell; novel strategies; rare variant; response; transcriptome sequencing; uptake; voltage; zinc-binding protein

Zinc is an essential micro-nutrient that participates in catalytic and structural functions touching nearly every metabolic process in the cell. While alterations in neuronal Zn2+ distribution have been associated with multiple disease states including Alzheimer’s Disease (AD), ALS, schizophrenia, and depression, the molecular mechanism of Zn2+ homeostasis in neurons is largely unexplored. The Zinc and Iron-reg- ulated transport Proteins (ZIP) mediate the entrance of first row transition metals into the cytoplasm. hZIP12, one of the fourteen human ZIPs, is a Zn2+ uptake transporter expressed in the brain. hZIP12 cSNPs (coding single nucleotide polymorphisms) have been associated with AD and abnormalities in the caudate nucleus. Importantly, our previous structural functional studies of the homologous hZIP4 enables postulation of the likely functional alteration introduced by observed cSNP. This proposal is based on the hypothesis that studies of hZIP12 and variants associated with neurodegenerative diseases will serve as an entrance point to build testable mechanistic models of neuronal Zn2+ homeostasis. Aim one tests the hypothesis that hZIP12 mutations associated with neuronal diseases impact transport activity and/or sur- face expression. hZIP12-mediated Zn2+ transport will be measured in neuronal SH-SY5Y cells expressing wild type and mutated proteins. Membrane targeting and dimerization studies will characterize predicted structural effects of observed cSNPs. The structural and functional consequence of three types of vari- ants will be examined: 1) one cSNP linked to AD, 2) two cSNPs associated with morphological alterations of the caudate nucleus; and 3) two cSNPs hypothesized to alter metal selectivity. Aim two tests the hy- pothesis that hZIP12 dysfunction modifies cellular Zn2+ pools and metal distribution. Cytosolic Zn2+ levels, as well as transmembrane transporter expression, will be assessed under early steady state and non- toxic Zn2+ levels. To this end, targeted as well as unbiased global approaches will be employed: 1) size exclusion chromatography-inductively coupled plasma mass spectrometry (SEC-ICP-MS) profiling of the proteome will be combined with mass spectrometry (LC-MS/MS) to identify Zn2+ binding proteins. 2) The expression of compensatory genes participating in the response will be evaluated via unbiased approaches (RNA-Seq) as well as directed Q-PCR of genes known to be involved in Zn2+ homeostasis/distribution. Results from these studies will initiate the definition of molecular and subcellular mechanisms of Zn2+ homeostasis in the brain, provide a detailed description of hZIP12 function in neuronal Zn2+ subcellular distribution, and elucidate the role of hZIP12 in relevant neurodegenerative diseases.

锌是必不可少的微营养素，几乎参与催化和结构功能几乎接触 细胞中的每个代谢过程。虽然神经元Zn2+分布的改变已关联 具有多种疾病状态，包括阿尔茨海默氏病（AD），ALS，精神分裂症和抑郁症， 神经元中Zn2+稳态的分子机制在很大程度上是意外的。锌和铁reg- 启用的转运蛋白（ZIP）介导了第一行转换金属进入细胞质的入口。 HZIP12是十四个人的拉链之一，是在大脑中表达的Zn2+摄取转运蛋白。 HZIP12 CSNP（编码单核苷酸多态性）与AD和异常有关 尾状核。重要的是，我们先前对同源HZIP4的结构功能研究使 假定观察到的CSNP引入的可能功能改变。该建议基于 假设HZIP12和与神经退行性疾病相关的变体的研究将作为 建立神经元Zn2+稳态的可测试机械模型的入口点。瞄准一个测试 假设HZIP12与神经元疾病相关的突变会影响运输活性和/或Sur- 面部表情。 HZIP12介导的Zn2+转运将在表达的神经元SH-SY5Y细胞中测量 野生型和突变的蛋白质。膜靶向和二聚化研究将表征预测的 观察到的CSNP的结构效应。三种类型的变量的结构和功能结果 将检查蚂蚁：1）一个与AD相关的CSNP，2）两个与形态变化相关的CSNP 尾状核； 3）假设有两个CSNP可以改变金属的选择性。瞄准两个测试 HZIP12功能障碍修饰剂细胞Zn2+池和金属分布的假顿。胞质Zn2+水平， 以及转运蛋白的表达，将在早期稳态和非 - 有毒Zn2+水平。为此，将雇用针对性和公正的全球方法：1）大小 排除色谱 - 感应耦合等离子质谱法（SEC-ICP-MS）分析 蛋白质组将与质谱（LC-MS/MS）结合使用，以识别Zn2+结合蛋白。 2） 参与响应的补偿基因的表达将通过无偏见进行评估 接近（RNA-seq）以及已知参与Zn2+的基因的定向Q-PCR 稳态/分布。这些研究的结果将启动分子和 大脑中Zn2+稳态的亚细胞机制提供了HZIP12的详细描述 在神经元Zn2+亚细胞分布中的功能，并阐明了HZIP12在相关的作用 神经退行性疾病。