Determination of the clinical relevance of Parkinson disease-associated intronic enhancer of the alpha-synuclein gene, in a novel mouse deletion model

在新型小鼠缺失模型中确定帕金森病相关的α-突触核蛋白基因内含子增强子的临床相关性

• 批准号: 10665271
• 负责人: Hanseok Ko
• 金额: $ 24.56万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-15 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10665271
• 关键词: Age; Age Months; Automobile Driving; Behavioral; Biological; Biological Assay; Cells; Chromatin; Cis-Acting Sequence; Clinical; Data; Disease; Disease Progression; Engineering; Enhancers; Equilibrium; Exhibits; Exposure to; Fluorescent in Situ Hybridization; Functional disorder; Gene Amplification; Genes; Genetic; Genetic Transcription; Genetic study; Genotype; Heterozygote; Histologic; Homozygote; Human; Immunohistochemistry; Inflammation; Injections; Introns; Knockout Mice; Lewy Bodies; Midbrain structure; Modeling; Motor; Mus; Mutate; Mutation; Nerve Degeneration; Neurons; Onset of illness; Orthologous Gene; Oxidopamine; Parkinson Disease; Parkinsonian Disorders; Pathologic; Pathology; Phenotype; Population; RNA Splicing; Reporter; Risk; Risk Factors; Risk Reduction; Role; SNCA gene; Severities; Severity of illness; Smell Perception; Symptoms; Techniques; Testing; Tetanus Helper Peptide; Tetracyclines; Titrations; Transcript; Transcriptional Regulation; Transgenic Mice; Untranslated RNA; Variant; Viral Vector; Work; alpha synuclein; behavioral phenotyping; clinically relevant; disease phenotype; disorder risk; dopaminergic neuron; glial activation; molecular phenotype; mouse model; mutant; neuron loss; neurotoxic; novel; novel therapeutics; olfactory bulb; phenotypic data; pre-formed fibril; promoter; protein misfolding; risk variant; single molecule; sporadic Parkinson' s Disease; therapeutic development

We will evaluate the biological necessity of an intronic enhancer of SNCA and determine its relevance to Parkinson disease (PD). SNCA is frequently mutated in familial PD; it encodes alpha-synuclein (α-syn), the primary constituent of Lewy bodies (LB). LB formation/accumulation is a pathological hallmark of PD and is driven by protein misfolding promoted by SNCA structural mutation, gene amplification, or genetic/environmental insults that elevate α-syn levels. Noncoding variants predicted to impact SNCA transcriptional regulatory control are also risk factors for sporadic PD. We recently identified PD-associated variants in SNCA intron 4. These variants lie within a dopaminergic (DA) neuron open chromatin region (OCR) that we have shown interacts with the SNCA promoter and is a cis-regulatory enhancer in catecholaminergic neurons. We predict that this sequence, and the variants therein, impact SNCA transcriptional control and modulate PD risk. Hypothesis - Deletion of the Snca enhancer in mice will reduce Snca transcription in PD-relevant cells with potential impact on their viability/distribution, and on motor/non-motor PD phenotypes (Aim 1); deletion will reduce DA neuron vulnerability to PD-relevant insults and ameliorate the onset and severity of disease (Aim2). Several highly effective strategies exist to elicit Parkinsonian pathology in mice. Tetracycline (tet)-dependent expression of PD mutant and wild-type (WT) forms of SNCA in DA neurons result in marked and progressive loss of ventral midbrain DA neuron populations, consistent with PD pathology. Likewise, intrastriatal injection of α-syn pre-formed fibrils (PFF) also result in a mouse model that exhibits progressive DA neuronal loss. Consistent with the known role of Snca levels impacting PD risk and progression, α-syn deficient mice are protected from PFF-induced neurodegeneration and from tet-dependent expression (via viral vector) of the PD missense mutant (hA53T) or WT SNCA. α-syn null mice are also protected from the neurotoxic effects of other PD promoting (and Snca-elevating) insults, like MPTP, 6-OHDA, and LPS. We will test whether cell-dependent titration of Snca levels, similarly, ameliorates risk and progression in a new mouse model. We have engineered mouse lines lacking this enhancer (Snca Enhdel) and provide preliminary evidence that this sequence impacts Snca transcription. Using established techniques, we propose to assay the effect of Snca Enhdel on prodromal, behavioral, and motor phenotypes (Aim 1a), as well as on PD-relevant neuron viability/distribution and microglial activation in the midbrain and olfactory bulb (Aim 1b) via immunohistochemistry and single molecule fluorescence in situ hybridization. Similarly, we will determine whether Snca Enhdel reduces onset and progression of prodromal/motor/non-motor disease phenotypes (Aim 2a) and impacts DA neuron vulnerability (Aim 2b) when exposed to PD-relevant insults (intrastriatal injection of SNCA PFF and adenoviral-delivery of SNCA hA53T). We will establish the extent to which modulating Snca transcription impacts risk of PD relevant pathology in mice, and its potential as a novel therapeutic avenue.

我们将评估SNCA内含子增强子的生物学必要性，并确定其相关性