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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.

摘要线粒体脑肌病伴线粒体DNA（mtDNA）缺失是一类线粒体疾病导致神经功能障碍，残疾和肌病。ADP特异性琥珀酰辅酶A合成酶（SCS）缺乏，a TCA循环的组成部分，是线粒体脑肌病的原因之一，线粒体DNA缺失。SCS是TCA循环的重要整合点，代谢和翻译后琥珀酰化。先前的研究表明，组成型突变Sucla 2等位基因纯合的胚胎小鼠是胚胎致死的，表现出ADP特异性SCS缺乏和mtDNA缺失，对能量产生不利影响新陈代谢.对于该提议，为了产生成年Sucla 2缺陷动物，采用了限于出生后前脑的Sucla 2敲除策略。初步调查揭示了突变动物在学习和记忆方面表现出缺陷，线粒体DNA缺失时大脑皮层的能量代谢和整体蛋白琥珀酰化耗尽这一建议是基于这样的假设，即Sucla 2的成人表型缺乏是由TCA循环功能障碍和整体翻译后琥珀酰化。Sucla 2-前脑敲除模型将通过一组有针对性的行为测定进行分析，认知领域的缺陷。为了确定发育的潜在差异效应， Sucla 2缺失的时间，产前发生Sucla 2缺陷的条件性敲除也将产生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、自主神经、感觉和认知测定阶段将分析来自前脑条件性敲除模型的皮质，组织病理学变化、代谢组学改变、TCA和ETC酶活性、总体翻译后琥珀酰化和mtDNA含量。一种创新的方法，单核（sn）RNA-seq和snATAC-seq鉴定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改变，将采用不同的表达簇，并使用数据集来告知组织病理学和代谢组学分析。这些研究将为深入了解致病机制提供帮助。 Sucla 2/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。