Nuclear mechanics and mechanotransduction in muscular laminopathies

描述(申请人提供)：超过三分之一的扩张型心肌病病例是由遗传突变引起的，其中5%至10%的突变与LMNA基因有关，LMNA基因编码核膜蛋白LMNA和层蛋白A和C。重要的是，LMNA基因的突变还导致广泛的其他疾病，包括Emery-Dreifuss肌营养不良症、肢体带状肌营养不良症和家族性部分脂肪营养不良症。尽管最近取得了进展，但不同的层粘连蛋白突变导致的肌肉特异性缺陷的机制(S)仍然难以捉摸。这一建议的中心假设是，层粘连蛋白突变可通过两种可能重叠的机制导致骨骼肌和心肌疾病：(I)层粘连蛋白A和C的结构功能丧失，导致机械应激组织中更脆弱的细胞核破裂；(Ii)扰乱(机械敏感)信号通路，导致肌肉细胞功能受损。特定的LMNA突变可能会不同地影响层蛋白功能的这些不同方面，导致广泛的疾病表型。我的长期目标是了解几乎无处不在表达的Lamins突变导致肌肉特异性表型的分子机制(S)，并探索核结构受损和细胞对机械应力的敏感性改变在多大程度上导致肌肉特异性表型。在第一个目标中，我们将检验这一假设，即改变核力学会导致机械应力组织中核破裂的增加。通过使用一种遗传报告试验，在三种肌层病变的小鼠模型中，甚至可以检测到心肌细胞核膜完整性的短暂受损，我们可以直接评估核膜蛋白的突变是否导致心脏组织核破裂的发生率增加。在第二个目标中，我们将确定椎板病中核力学受损与肌肉表型严重程度之间的关系。利用表达一组含有不同肌肉受累的Lamin突变的果蝇模型，我们将把这些突变对果蝇幼虫完整肌肉组织中细胞核力学性质的影响与成年果蝇肌肉缺陷的严重程度联系起来。在第三个目标中，我们将研究导致扩张型心肌病的层蛋白突变与特定的信号通路--肌钙蛋白相关转录因子A(MRTF-A)之间的相互作用。我们将探索我们最近发现的层蛋白A/C缺陷和突变细胞中MRTF-A核转位受损的机制(S)，并评估MRTF-A信号受损对细胞功能的影响。研究Lamin突变对细胞核结构和细胞信号的影响将有助于我们更好地理解这些蛋白的正常和组织特异性功能，并导致对扩张型心肌病、Emery-Dreifuss肌营养不良症和其他椎板病的分子机制的新见解，可能为这些疾病的治疗提供新的靶点。

DESCRIPTION (provided by applicant): More than one-third of all cases of dilated cardiomyopathy are caused by inherited mutations, with 5% to 10% of these mutations being linked to the LMNA gene, which encodes the nuclear envelope proteins lamin A and C. Importantly, mutations in the LMNA gene are also responsible for a broad spectrum of other diseases, including Emery-Dreifuss muscular dystrophy, limb-girdle muscular dystrophy and familial partial lipodystrophy. Despite recent advances, the mechanism(s) responsible for the often muscle-specific defects caused by different lamin mutations remains elusive. The central hypothesis of this proposal is that lamin mutations can cause skeletal and cardiac muscle disease through two, possibly overlapping mechanisms: (i) loss of structural function of lamins A and C, leading to rupture of the more fragile nucleus in mechanically stressed tissues; (ii) disturbing (mechanosensitive) signaling pathways that results in impaired function of muscle cells. Specific LMNA mutations may differentially affect these distinct aspects of lamin function, resulting in a broad spectrum of disease phenotypes. My long term goal is to understand the molecular mechanism(s) by which mutations in the nearly ubiquitously expressed lamins can lead to muscle-specific phenotypes and to explore to what extent impaired nuclear structure and altered cellular sensitivity to mechanical stress contribute to the muscle-specific phenotypes. In the first aim, we will test the hypothesis that altered nuclear mechanics result in increased nuclear rupture in mechanically stressed tissue. By using a genetic reporter assay that can detect even transiently compromised nuclear envelope integrity in cardiac myocytes in three mouse models of muscular laminopathies, we can directly assess whether mutations in nuclear envelope proteins cause increased rates of nuclear rupture in cardiac tissue. In the second aim, we will determine the relationship between impaired nuclear mechanics and the severity of muscular phenotypes in laminopathies. Using drosophila melanogaster models expressing a panel of lamin mutations with variable muscle involvement, we will relate effects of the mutations on the mechanical properties of nuclei in intact muscle tissue in drosophila larvae with the severity of muscle defects in adult flies. In the third aim, we will investigate the interplay between lamin mutations responsible for dilated cardiomyopathy and a specific signaling pathway, myocardin-related transcription factor A (MRTF-A). We will explore the mechanism(s) responsible for the impaired nuclear translocation of MRTF-A in lamin A/C-deficient and mutant cells we recently discovered and assess the functional consequences of impaired MRTF-A signaling on cellular function. Studying the effects of lamin mutations on nuclear structure and cellular signaling will improve our understanding of normal and tissue-specific functions of these proteins and lead to new insights into the molecular mechanisms responsible for dilated cardiomyopathy, Emery-Dreifuss muscular dystrophy and other laminopathies, potentially providing new targets for the treatment of these diseases.