Structure-Function of the Nuclear Envelope Bridge and its Role in Laminopathies

描述（由申请人提供）：在哺乳动物细胞中，A型和B型核纤层蛋白在核膜的核质面形成二维蛋白质网络，即核纤层。沿着编码A型核纤层蛋白的LMNA基因散布的突变以及其他核膜蛋白内的突变与统称为核纤层蛋白病的广泛的人类疾病相关。这些疾病的分子病因学仍然未知。Emery-Dreifuss肌营养不良症（EDMD）是最突出的核纤层蛋白病，是一种不可治愈的、破坏性的肌肉消耗性疾病，由内核膜（INM）蛋白Emerin、核纤层蛋白A或外核膜（ONM）KASH-蛋白nesprins 1和2中的突变引起。这四种蛋白质通过LINC复合物连接，LINC复合物是INM SUN-蛋白质和ONM KASH-蛋白质之间的进化保守蛋白质复合物，其桥接核膜的表面并将核纤层物理连接到哺乳动物细胞的细胞骨架。总之，数据表明EDMD是异常核定位的结果，或者是通过LINC复合物的异常机械信号传导的结果。此外，已显示Sun 1在INM处的过度积累是EDMD的病理效应子。通过探索性R21赠款的资助，我们在2012年确定了SUN-KASH复合物的核心结构，为LINC复合物提供了第一个分子见解。在这里，我们建立在这些数据的基础上，并提出了一个研究计划，应该有助于发现药物靶点，希望将转化为EDMD患者的药物治疗策略。该提案概述了将导致A）对人类SUN-KASH相互作用组的全面结构和生物化学理解，B）LINC复合物锚定到核纤层蛋白层的结构基础，以及C）对LINC复合物组装和拆卸的调节的洞察的实验。我们期望，对这三个目标的追求将产生对形成EDMD基础的蛋白质网络的更好的分子描述，从而揭示破坏这些过程的可能药物靶点。我们进一步预计，这项研究将促进我们对核膜的理解，这将对大量的病理性核膜疾病产生切实的影响。

DESCRIPTION (provided by applicant): In mammalian cells, A- and B-type lamins form a two-dimensional protein meshwork, the lamina, at the nucleoplasmic face of the nuclear envelope. Mutations scattered along the LMNA gene, which encodes A-type lamins, as well as mutations within other nuclear envelope proteins are associated with a broad range of human diseases collectively called laminopathies. The molecular etiology of these diseases remains unknown. Emery-Dreifuss muscular dystrophy (EDMD), the most prominent laminopathy, is an incurable, devastating muscular wasting disease, caused by mutations in either, the inner nuclear membrane (INM) protein emerin, laminA, or the outer nuclear membrane (ONM) KASH-proteins nesprins 1 and 2. The four proteins are connected via LINC complexes, evolutionary-conserved protein complexes between INM SUN-proteins and ONM KASH-proteins that bridge the faces of the nuclear envelope and physically connect the nuclear lamina to the cytoskeleton of mammalian cells. Taken together, the data suggests that EDMD is the result of aberrant nuclear positioning or, alternatively, aberrant mechanical signaling through the LINC complex. Furthermore, it has been shown that the over-accumulation of Sun1 at the INM is the pathological effector of EDMD. With funding through an exploratory R21 grant we have determined the core structure of the SUN-KASH complex in 2012, providing the first molecular insight into LINC complexes. Here, we build on this data and suggest a research program that should aid in the discovery of drug targets that hopefully will translate into a medication strateg for EDMD patients. This proposal outlines experiments that will lead A) to a comprehensive structural and biochemical understanding of the human SUN-KASH interactome, B) a structural basis for LINC complex anchorage to the lamin layer, and C) insight into the regulation of LINC complex assembly and disassembly. We expect that the pursuit of these three aims will yield a much better molecular description of the protein network that forms the basis of EDMD, and consequently will unveil possible drug targets that disrupt these processes. We further anticipate that this research will advance our understanding of the nuclear envelope in general, which will have a tangible impact on the vast array of pathological nuclear envelope disorders.