Role of SETD5 in Moyamoya Disease Pathogenesis

SETD5 在烟雾病发病机制中的作用

基本信息

批准号：
10724796
负责人：
Callie S Kwartler
金额：
$ 15.6万
依托单位：
UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-15 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10724796
关键词：
Acetylation Affect Alleles Antibodies Arteries Bilateral Biological Models Brain CRISPR/Cas technology Cell Differentiation process Cell Line Cell Proliferation Cells Cellular Metabolic Process Cerebral Revascularization Cerebrovascular Disorders ChIP-seq Characteristics Child Childhood stroke Co-Immunoprecipitations Complex Consensus Coupled Data Defect Development Developmental Delay Disorders Disease Disease model Distal Epigenetic Process Future Genes Genetic Genetic Transcription Genus Hippocampus Glycolysis Heart Heterozygote Histone Acetylation Human Impairment In Vitro Intellectual functioning disability Internal carotid artery structure Lesion Link Loss of Heterozygosity Metabolic Metabolism Molecular Moyamoya Disease Mus NCOR1 gene Neural Crest Neural Crest Cell Neural tube Oxidative Phosphorylation Pathogenesis Pathogenicity Pathology Pathway interactions Patients Phenotype Procedures Production Proliferating Proteins Role Smooth Muscle Myocytes Somites Specific qualifier value Stains Stenosis Stroke Susceptibility Gene Syndrome Terminator Codon Testing Therapeutic Time Variant Vascular Endothelium Work alpha Actin artery occlusion cell fate specification cell motility cerebrovascular chromatin remodeling de novo mutation genetic variant genome editing genomic locus genomic profiles improved induced pluripotent stem cell insight knock-down loss of function migration model development neurosurgery prevent progenitor stem cell differentiation stem cells young adult

项目摘要

ABSTRACT Moyamoya disease (MMD) occurs when the distal internal carotid arteries are progressively narrowed and eventually occluded, and is a common cause of pediatric stroke. Numerous pathogenic genetic variants have been identified to cause MMD, but a common mechanism of pathogenesis has yet to be defined. Pathology from affected vessels shows the occlusive lesions are comprised of fibroproliferative cells that stain positive for smooth muscle cell (SMC)-specific α-actin (SMA); we therefore propose that SMC migration and proliferation may be drivers of the disease. Multiple genes encoding proteins that participate in chromatin remodeling have been identified to cause MMD, including heterozygous loss of function (LOF) variants in the gene SETD5. SETD5 interacts with the nuclear receptor-corepressor (Ncor) complex to regulate histone acetylation. Our previous work on MMD-causing pathogenic variants in ACTA2 showed that these variants impair SMC differentiation, and the incompletely differentiated cells have increased proliferation and migration and rely on glycolysis for cellular energy production. Importantly, treatments that boost oxidative phosphorylation restored differentiation and reduced migration in mouse SMCs with MMD-causing Acta2 variants, suggesting a potential therapeutic strategy. Based on these results and the list of identified genetic triggers for MMD, we propose a common pathogenic mechanism: aberrant chromatin remodeling during SMC specification leads to cells that proliferate and migrate to occlude the vessels. Here, we will test this hypothesis in cells with LOF variants in SETD5 in two specific aims. 1) We will assess whether LOF variants in SETD5 impact SMC differentiation and phenotype. We will use Crispr/Cas9 gene editing to introduce SETD5 LOF alleles into human induced pluripotent stem cells (iPSCs). We will differentiate these iPSCs alongside isogenic controls into neural crest progenitors and then into SMCs, and will characterize the differentiation, proliferation, migration, and metabolism of the resulting cells. 2) We will assess whether SETD5 impacts chromatin remodeling at loci critical for SMC differentiation. We will introduce a 3xFlag tag at the C-terminus of the SETD5 protein using targeted Crispr/Cas9 gene editing in human iPSCs and will use these cells to identify genomic loci where SETD5 is acting in iPSCs, neural crest progenitors, and SMCs by chromatin immunoprecipitation sequencing. We will assess whether LOF variants in SETD5 affect histone acetylation and gene transcription at the identified loci. Completion of these aims will link SETD5- dependent chromatin remodeling with SMC phenotype and elucidate the molecular mechanisms by which LOF variants in SETD5 cause MMD. The results have the potential to identify therapeutic strategies to treat or prevent MMD in patients with SETD5 LOF variants. Finally, these data will dramatically advance our understanding of a potential common pathway for MMD pathogenesis.

抽象的烟雾病（MMD）发生在远端颈内动脉逐渐变窄并且最终闭塞，并且是许多致病性遗传变异的常见原因。已被确定可引起 MMD，但尚未确定病理学的共同发病机制。受影响的血管显示闭塞性病变由纤维增殖细胞组成，其染色呈阳性因此，我们提出平滑肌细胞（SMC）特异性α-肌动蛋白（SMA）；编码参与染色质重塑的蛋白质的多个基因可能是该疾病的驱动因素。 SETD5 已被确定可引起 MMD，包括 SETD5 基因中的杂合功能丧失 (LOF) 变异。与核受体辅阻遏物 (Ncor) 复合物相互作用来调节组蛋白乙酰化。对 ACTA2 中引起 MMD 的致病性变异的研究表明，这些变异会损害 SMC 分化，并且不完全分化的细胞增殖和迁移增加，细胞依赖糖酵解重要的是，促进氧化磷酸化的治疗恢复了分化和能力。导致 MMD 的 Acta2 变异的小鼠 SMC 的迁移减少，这表明了一种潜在的治疗方法根据这些结果和已确定的 MMD 遗传触发因素列表，我们提出了一个通用策略。致病机制：SMC规范过程中染色质重塑异常导致细胞增殖在这里，我们将在 SETD5 中具有 LOF 变异的细胞中检验这一假设。具体目标 1) 我们将评估 SETD5 中的 LOF 变异是否影响 SMC 分化和表型。将使用 Crispr/Cas9 基因编辑将 SETD5 LOF 等位基因引入人类诱导多能干细胞（iPSC）。我们将这些 iPSC 与同基因对照分化为神经嵴祖细胞，然后分化为神经嵴祖细胞。 SMC，并将表征所得细胞的分化、增殖、迁移和代谢 2)。我们将评估 SETD5 是否影响对 SMC 分化至关重要的位点的染色质重塑。使用针对人类的 Crispr/Cas9 基因编辑在 SETD5 蛋白的 C 末端引入 3xFlag 标签 iPSC 并将使用这些细胞来识别 SETD5 在 iPSC、神经嵴祖细胞、我们将通过染色质免疫沉淀测序评估 SETD5 中的 LOF 变异是否会影响。完成这些目标将连接 SETD5- 的组蛋白乙酰化和基因转录。 SMC 表型依赖性染色质重塑并阐明 LOF 的分子机制 SETD5 的变异会导致 MMD，这些结果有可能确定治疗或预防 MMD 的治疗策略。最后，这些数据将极大地促进我们对 SETD5 LOF 变异患者的理解。 MMD 发病机制的潜在共同途径。