Characterizing Tissue Specific Regulation of Mutant Lamin Protein Degradation

项目摘要 椎板病是一种广泛的疾病，包括多系统疾病，如衰老， 哈钦森病、吉尔福德早衰症和组织特异性疾病，如埃默里-德赖富斯肌 营养不良和腓骨肌萎缩症的神经病变。所有这些不同的疾病都可能是由 然而，LMNA基因中的突变如何引起这种不同的疾病， 不完全理解。已经发现核纤层蛋白的突变形式聚集，并且这种聚集是 与受损的细胞功能和疾病表型相关。因此，我们假设某些 由于不能进行组织特异性质量控制 降解这些突变形式的机制，导致蛋白质聚集和细胞毒性。我们将测试 这个假设使用果蝇作为我们的模型系统。果蝇让我们 操纵特定组织中的基因表达，使实验能够区分不同的核纤层蛋白质 突变在特定组织中降解。苍蝇有两个人类LMNA基因的同源物，LamC和 Lam Dm 0，这些基因的突变模拟人类疾病。此外，引入等效 导致人类疾病的LamC突变导致核纤层蛋白聚集和疾病样 表型此外，AMPK信号，这是下调肌肉活检从病人，已被证实， 发现减少突变LamC聚集在苍蝇肌肉。另一方面，我们发现Lam Dm 0 p38 MAPK（p38 Kb）和CASA复合物调节衰老过程中肌肉中的聚集， Lam Dm 0的降解。LamC和Lam Dm 0之间的一个差异是，虽然两种核纤层蛋白都是 在肌肉中表达，只有Lam Dm 0在神经元中表达。由于LamC和Lam Dm 0具有不同的组织 特异性，我们还可以评估特定的蛋白质质量控制机制是否能够靶向某些突变体 每种组织中的核纤层蛋白。这将使我们能够确定疾病组织特异性是否是由于 蛋白质质量控制机制，以降解某些突变形式的核纤层蛋白，导致肌肉 例如，营养不良而不是神经病。因此，我们将1）确定p38 Kb和CASA是否 复合物调节特定形式的突变LamC蛋白的聚集，以及这如何有助于 肌肉缺陷，2）表征Lam Dm 0中的特定疾病突变如何影响肌肉中的聚集， 3）确定AMPK和p38 Kb是否在相同或不同的途径中调节突变核纤层蛋白 聚合来我们希望我们提出的研究将为核纤层蛋白的突变形式如何导致 在疾病状态下，如果激活不同的蛋白质质量控制机制可以证明是有效的， 治疗机制

Project Summary Laminopathies are a wide range of disorders that include both multisystem disorders such as the aging disorder Hutchinson Guilford Progeria and tissue specific disorders such as Emery-Dreifuss Muscular Dystrophy and the neuropathy Charcot-Marie-Tooth disease. All of these different disorders can be caused by mutations in the LMNA gene, however, how mutations in one gene give rise to such different disorders is incompletely understood. Mutant forms of lamin protein have been found to aggregate, and this aggregation is associated with impaired cellular function and disease phenotypes. Therefore, we hypothesize that certain tissues are susceptible to specific lamin mutations due to the inability of tissue specific quality control mechanisms to degrade those mutant forms, leading to protein aggregation and cellular toxicity. We will test this hypothesis using the fruit fly Drosophila melanogaster as our model system. Drosophila allow us to manipulate gene expression in specific tissues, enabling experiments to tease apart how different lamin protein mutations are degraded in specific tissues. Flies have two homologues of the human LMNA gene, LamC and Lam Dm0, and mutations in these genes mimic human disorders. In addition, introducing the equivalent mutations that cause human disease into LamC results in lamin protein aggregation and disease-like phenotypes. Furthermore, AMPK signaling, which is downregulated in muscle biopsies from patients, has been found to reduce mutant LamC aggregation in the fly muscle. On the other hand, we find that Lam Dm0 aggregates in the muscle during aging, and that p38 MAPK (p38Kb) and the CASA complex regulate the degradation of Lam Dm0. One difference between LamC and Lam Dm0 is that while with both lamins are expressed in muscle, only Lam Dm0 is expressed in neurons. As LamC and Lam Dm0 have different tissues specificities, we can also assess if specific protein quality control mechanisms are able to target certain mutant forms of lamin in each tissue. This will allow us to determine if disease tissue specificity is due to the failure of protein quality control machinery to degrade certain mutant forms of lamin protein, resulting in muscular dystrophy rather than neuropathy, for example. Therefore, we will 1) determine if p38Kb and the CASA complex regulate the aggregation of specific forms of mutant LamC proteins and how this contributes to muscle defects, 2) characterize how specific disease mutations in Lam Dm0 affect aggregation in muscles and neurons, and 3) determine if AMPK and p38Kb act in the same or different pathways to regulate mutant lamin aggregation. We expect that our proposed study will provide new insights into how mutant forms of lamin result in a disease state and if activating different protein quality control mechanisms could prove to be an effective therapeutic mechanism.