Mechanisms of DUX4 mediated FSHD pathology

DUX4 介导的 FSHD 病理机制

• 批准号: 9098601
• 负责人: Peter L Jones
• 金额: $ 15.68万
• 依托单位: UNIV OF MASSACHUSETTS MED SCH WORCESTER
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-03 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9098601
• 关键词: 4q35; Adult; Affect; Biological Models; Biopsy; C-terminal; Candidate Disease Gene; Cell Death; Cell physiology; Cells; Child; Chromosomes; Clinical; Code; Consensus; Contracts; D4Z4; DNA; DNA Methylation; Development; Disease; Epigenetic Process; Facioscapulohumeral Muscular Dystrophy; Family; Gene Expression; Gene Targeting; Genes; Genetic; Goals; Individual; Laboratories; Lead; Length; Lesion; Libraries; Link; Mediating; Messenger RNA; Modeling; Molecular; Molecular Profiling; Muscle; Muscle Cells; Muscle Fibers; Muscle Weakness; Muscular Dystrophies; Mutation; Myopathy; Pathogenesis; Pathology; Pathway interactions; Patients; Phase; Poly A; Production; Protein Isoforms; Proteins; RNA Splicing; Regulation; Regulatory Element; Research; Research Design; Role; Staging; TP53 gene; Testing; Therapeutic; Time; Variant; base; cohort; cytotoxic; epigenetic regulation; histone modification; in vivo; mRNA Expression; novel therapeutic intervention; novel therapeutics; overexpression; promoter; public health relevance; research study; targeted treatment; telomere; therapy design

DESCRIPTION (provided by applicant): Our proposed experiments will identify mechanisms that cause mis-expression of DUX4 and lead to pathogenesis in facioscapulohumeral muscular dystrophy (FSHD). FSHD is the most prevalent myopathy afflicting both children and adults, but no therapy is known. However, FSHD research has now entered an important new stage as DUX4 has emerged from studies in multiple laboratories, including our own, as the consensus FSHD candidate gene. Current evidence supports a model for FSHD pathology in which stable expression of a polyadenylated DUX4 mRNA splicing variant, termed DUX4-fl (fl = full-length), leads to production of a pathogenic DUX4-FL protein. This model is compatible with our finding, based on analysis of a large new library of myogenic cells and biopsies, that DUX4-fl mRNA and DUX4-FL protein are expressed at a much higher level and in a much higher fraction of myonuclei in FSHD than in healthy control muscle cells. Thus, in FSHD patients, clinically apparent muscle weakness may develop as DUX4-FL-induced pathological changes accumulate over time, whereas in healthy control muscles, the extremely low level of DUX4- FL is apparently insufficient to induce overt pathology. Although the FSHD-associated genetic lesion does not obviously alter protein coding or mRNA sequences, there is strong evidence that aberrant epigenetic regulation underlies DUX4 mis-expression. Our preliminary studies, for example, showed that DUX4-fl expression levels in FSHD myogenic cells could indeed be altered by manipulations of epigenetic status, e.g., DNA methlyation. Based on these epigenetic studies and our complementary analyses of DUX4-FL expression, we propose and will test the hypothesis that epigenetic dysregulation leads to aberrantly increased expression of DUX4-fl in FSHD vs. unaffected myogenic cells and thus to pathology. In Specific Aim 1, we wil identify epigenetic and non-epigenetic mechanisms that regulate DUX4-fl mRNA expression levels in skeletal muscle cells. In Specific Aim 2, we will analyze DUX4-FL expression in biopsies and also use single cell analyses to determine how DUX4-FL expression alters the fate of FSHD vs. unaffected cels. In Specific Aim 3, we will identify gene networks that are differentially regulate by the DUX4 protein isoforms and determine if they are disrupted in DUX4-fl-expressing FSHD cells. The results of our studies, in which we will systematically investigate the expression, regulation, and function of DUX4 isoforms in FSHD and control muscle cells, will provide the vital information needed to guide development of FSHD therapies specifically targeted to DUX4.

描述（由申请人提供）：我们提出的实验将确定导致DUX4误表达并导致facioscapulohumeral肌肉营养不良（FSHD）的发病机理的机制。 FSHD是困扰儿童和成人的最普遍的肌病，但尚无疗法。但是，FSHD研究现在已经进入了一个重要的新阶段，因为DUX4从多个实验室（包括我们自己的）的研究中出现了FSHD候选基因。当前的证据支持FSHD病理学的模型，在该模型中，多腺苷酸化DUX4 mRNA剪接变体的稳定表达称为DUX4-FL（FL =全长），导致致病性DUX4-FL蛋白的产生。该模型与我们的发现兼容，基于对大型肌原性细胞和活检库的分析，即DUX4-FL mRNA和DUX4-FL蛋白在FSHD中比在健康对照肌肉细胞中更高的水平表达，并且在FSHD中的肌核中比例更高。因此，在FSHD患者中，随着DUX4-FL诱导的病理变化的积累，临床上明显的肌肉无力可能会发展出来，而在健康对照肌肉中，DUX4-FL的极低水平显然不足以诱导明显的病理。尽管与FSHD相关的遗传病变显然不会改变蛋白质编码或mRNA序列，但有强有力的证据表明异常表观遗传调节是DUX4错误表达的基础。例如，我们的初步研究表明，FSHD肌原细胞中的DUX4-FL表达水平确实可以通过表观遗传状态（例如DNA甲基化）的操作来改变。基于这些表观遗传学研究和我们对DUX4-FL表达的互补分析，我们提出并将检验以下假设：表观遗传失调导致在FSHD与未受影响的肌原性细胞中dux4-FL的表达异常增加，从而导致了病理学。在特定目标1中，我们将确定调节骨骼肌细胞中DUX4-FL mRNA表达水平的表观遗传和非观测机制。在特定的目标2中，我们将分析活检中的DUX4-FL表达，并使用单细胞分析来确定DUX4-FL表达如何改变FSHD的命运与未受影响的Cels的命运。在特定目标3中，我们将确定由DUX4蛋白同工型差异调节的基因网络，并确定它们是否在表达DUX4-FL的FSHD细胞中被破坏。我们的研究结果（我们将系统地研究FSHD和对照肌肉细胞中DUX4同工型的表达，调控和功能）将提供指导特定针对DUX4的FSHD疗法的重要信息。