Determination of pathogenetic mechanisms in cortex-specific Sucla2 deficiency as a model for mitochondrial encephalopathy

确定皮质特异性 Sucla2 缺陷的发病机制作为线粒体脑病模型

基本信息

批准号：
10629432
负责人：
Brett Harrison Graham
金额：
$ 54.06万
依托单位：
INDIANA UNIV-PURDUE UNIV AT INDIANAPOLIS
依托单位国家：
美国
项目类别：
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/10629432
关键词：
ACTL6B gene ATAC-seq Acylation Adolescent Adult Alleles Animals Behavioral Behavioral Assay Binding Biological Assay Brain Cardiomyopathies Cell Nucleus Cells Chemicals Chromatin Citric Acid Cycle Cognitive Cognitive deficits Complex Control Animal Data Data Set Defect Development Dimensions Disease Dissection Electron Transport Embryo Encephalopathies Energy Metabolism Energy-Generating Resources Enzymes Exhibits Food Functional disorder Future Gene Expression Profile Genetic Hippocampus Intellectual functioning disability Investigation Knock-out Learning LoxP-flanked allele Lysine Measures Memory Metabolic Mitochondria Mitochondrial DNA Mitochondrial Diseases Mitochondrial Encephalomyopathies Modeling Molecular Motor Mus Muscle Mutant Strains Mice Myopathy Neurologic Dysfunctions Neurons Nuclear Nuclear RNA Organ Pathogenesis Pathogenicity Pathologic Pathway interactions Pattern Phenotype Physiological Ploidies Pre-Clinical Model Prosencephalon Proteins Proteomics Quantitative Reverse Transcriptase PCR Role Sampling Screening Result Sensory Succinate-CoA Ligases Therapeutic Tissue-Specific Gene Expression Tissues Transgenes Tricarboxylic Acids Validation cell cortex cell type cohort conditional knockout design efficacious treatment enzyme activity epigenomics in vivo innovation insight metabolic phenotype metabolomics mitochondrial dysfunction multiple omics mutant nervous system disorder neuropathology novel therapeutic intervention postnatal prenatal screening transcriptome sequencing transcriptomics

项目摘要

Abstract Mitochondrial encephalomyopathy with mitochondrial DNA (mtDNA) depletion is a class of mitochondrial disease that causes neurological dysfunction with intellectual and developmental disabilities and myopathy. Deficiency of ADP-specific succinyl-CoA synthetase (SCS), a component of the TCA cycle, is one of the causes of mitochondrial encephalomyopathy with mtDNA depletion. SCS is an important integration point of the TCA cycle, intersecting energy metabolism and posttranslational succinylation. Previous studies have demonstrated that embryonic mice homozygous for a constitutive mutant Sucla2 allele are embryonic lethal and exhibit ADP-specific SCS deficiency and mtDNA depletion with detrimental effects on energy metabolism. For this proposal, to generate adult Sucla2 deficient animals, a conditional knockout strategy of Sucla2 limited to the postnatal forebrain is employed. Initial investigations reveal that the mutant animals exhibit a defect in learning and memory and perturbations of energy metabolism and global protein succinylation in the cortex in the absence of mtDNA depletion. This proposal is based on the hypothesis that adult phenotypes of Sucla2 deficiency are caused by a combination of TCA cycle dysfunction and perturbations of global posttranslational succinylation. The Sucla2-forebrain knockout model will be analyzed by a targeted battery of behavioral assays designed to fully characterize different cognitive domains for deficits. To determine the potential differential effects of developmental timing of Sucla2 loss, a conditional knockout with prenatal onset of Sucla2 deficiency will also be generated and subjected to longitudinal broad behavioral and in vivo physiological screening, including basic motor, autonomic, sensory, and cognitive assays in both adolescent and adult stages. Cortex from the forebrain conditional knockout model will be analyzed for histopathological changes, metabolomic alterations, TCA and ETC enzyme activities, global posttranslational succinylation and mtDNA content. An innovative approach using integrated single nuclear (sn) RNA-seq and snATAC-seq to identify differential gene expression patterns and alterations in chromatin accessibility landscape in different cortical cell types defined by distinct expression clusters will be employed and the datasets used to inform histopathological and metabolomic analyses. These studies will provide insights into the pathogenic mechanisms of Sucla2/SCS deficiency and establish a preclinical model to explore and develop novel therapeutic approaches.

抽象的线粒体 DNA (mtDNA) 缺失的线粒体脑肌病是一类线粒体疾病导致智力和发育神经功能障碍残疾和肌病。 ADP 特异性琥珀酰辅酶 A 合成酶 (SCS) 缺乏 TCA 循环的组成部分，是线粒体脑肌病的病因之一线粒体DNA耗尽。 SCS是TCA循环的重要整合点，与能量交叉代谢和翻译后琥珀酰化。先前的研究表明含有组成型突变体 Sucla2 等位基因的纯合胚胎小鼠是胚胎致死的，并且表现出 ADP 特异性 SCS 缺乏和 mtDNA 耗尽，对能量产生不利影响代谢。对于该提案，要产生 Sucla2 缺陷的成年动物，有条件采用仅限于出生后前脑的 Sucla2 敲除策略。初步调查揭示突变动物表现出学习和记忆缺陷以及扰动没有 mtDNA 的情况下皮质中的能量代谢和整体蛋白质琥珀酰化消耗。该提议基于这样的假设：Sucla2 的成体表型缺乏是由 TCA 循环功能障碍和扰动共同引起的全局翻译后琥珀酰化。 Sucla2-前脑敲除模型将是通过一系列有针对性的行为分析进行分析，旨在充分表征不同的特征认知领域的缺陷。确定发育的潜在差异效应 Sucla2 缺失的时机，产前出现 Sucla2 缺乏症的条件性敲除也将生成并进行纵向广泛的行为和体内生理筛选，包括青少年和成人的基本运动、自主、感觉和认知测定阶段。将分析前脑条件敲除模型的皮层组织病理学变化、代谢组学改变、TCA 和 ETC 酶活性、全球翻译后琥珀酰化和 mtDNA 含量。使用集成的创新方法单核 (sn) RNA-seq 和 snATAC-seq 用于识别差异基因表达模式以及不同皮质细胞类型中染色质可及性景观的变化，定义为将采用不同的表达簇和用于通知组织病理学的数据集和代谢组学分析。这些研究将为致病机制提供见解 Sucla2/SCS缺陷的研究并建立临床前模型以探索和开发新的治疗方法。