Gatekeepers of Mitochondrial NAD+

线粒体 NAD 的看门人

基本信息

批准号：
10301237
负责人：
Xiaolu Ang Cambronne
金额：
$ 7.93万
依托单位：
UNIVERSITY OF TEXAS AT AUSTIN
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-09-30 至 2022-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10301237
关键词：
Address Affect Age Back Binding Proteins Bioenergetics Biosensor Cardiovascular Diseases Cell Nucleus Cell Respiration Cells Chromatin Citric Acid Cycle Cultured Cells Cytoplasm Data Detection Development Disease Dyes Etiology Fluorescence Gatekeeping Gene Expression Generations Genetic Health Homeostasis Human Pathology In Vitro Infection Intervention Lead Learning Link Malignant Neoplasms Masks Measurement Measures Metabolic Metabolic Pathway Metabolic syndrome Metabolism Methods Mitochondria Mitochondrial Matrix Modification Molecular Monitor Mutagenesis Myopathy Nerve Degeneration Neurologic Nicotinamide adenine dinucleotide Noise Nuclear Onset of illness Organelles Oxides Pathology Physiological Positioning Attribute Regulation Reporter Reporting Role Series Signal Transduction Structure System Technology Testing Time Tissues Variant Whole Organism base cell type design experimental study improved in vivo insight interest novel strategies response sensor temporal measurement

项目摘要

Project summary We are working on new in vivo methods to study the intermediary metabolite nicotinamide adenine dinucleotide (NAD+). NAD+ is critical for cellular metabolism and health, and its decreased steady-state levels have been linked to human pathologies such as neurodegeneration, cardiovascular disease, metabolic syndrome, and cancer. NAD+ concentrations are highly compartmentalized by cell type, subcellular localization, and its protein- bound or free fractions. A lack of methods that can monitor free NAD+ in cells with spatial and temporal information has hindered our learning about the relevant pools, threshold concentrations, and fluctuations that NAD+ may undergo leading to disease onset. This precludes our ability to identify treatments or approaches to intervene before NAD+ levels are misregulated. In particular, there are no genetically-encoded methods for in vivo mitochondrial NAD+ measurements. Free mitochondrial NAD+ is required for cellular respiration, and diminished levels due to genetics, age, or infection can promote reductive metabolism. Thus, to address this gap we will improve the brightness (Aim 1) of a variant of the current NAD+ sensor, which is able to monitor free mitochondrial NAD+ in cells. We will also generate a series of constructs that will be used to test a new approach for improving the dynamic range of fluorescent readout (Aim 2). Together, data from these the aims will identify a feasible method to robustly obtain free mitochondrial NAD+ measurements in vivo.

项目概要我们正在研究新的体内方法来研究中间代谢物烟酰胺腺嘌呤二核苷酸（NAD+）。 NAD+ 对于细胞代谢和健康至关重要，其稳态水平下降已被证实与神经退行性疾病、心血管疾病、代谢综合征等人类疾病有关癌症。 NAD+ 浓度根据细胞类型、亚细胞定位及其蛋白质进行高度划分。结合或游离分数。缺乏可以在空间和时间上监测细胞中游离 NAD+ 的方法信息阻碍了我们对相关池、阈值浓度和波动的了解 NAD+ 可能会导致疾病发作。这妨碍了我们识别治疗或方法的能力在 NAD+ 水平失调之前进行干预。特别是，目前还没有针对这些疾病的基因编码方法。体内线粒体 NAD+ 测量。细胞呼吸需要游离线粒体 NAD+，并且由于遗传、年龄或感染而导致的水平降低可以促进还原代谢。因此，为了弥补这一差距我们将提高当前 NAD+ 传感器变体的亮度（目标 1），该传感器能够免费监控细胞中的线粒体 NAD+。我们还将生成一系列用于测试新方法的构造用于提高荧光读数的动态范围（目标 2）。这些目标的数据一起将确定一种可靠地获得体内游离线粒体 NAD+ 测量的可行方法。