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型层蛋白在核膜的核质面形成二维蛋白质网，即层。编码a型层粘连蛋白的LMNA基因上的突变，以及其他核膜蛋白上的突变，与一系列统称为层粘连病的人类疾病有关。这些疾病的分子病因尚不清楚。emry - dreifuss肌营养不良症（EDMD）是一种无法治愈的、毁灭性的肌肉萎缩疾病，由核膜（INM）蛋白emerin、laminA或外核膜（ONM） kash蛋白nesprin1和nesprin2突变引起。这四种蛋白质通过LINC复合物连接在一起，LINC复合物是INM sun蛋白和ONM kash蛋白之间的进化保守蛋白复合物，它连接了核膜的表面，并将核层物理地连接到哺乳动物细胞的细胞骨架上。综上所述，这些数据表明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的结果。此外，研究表明，INM处Sun1的过度积累是EDMD的病理效应。在R21探索性拨款的资助下，我们于2012年确定了SUN-KASH复合物的核心结构，首次提供了对LINC复合物的分子洞察。在此，我们以这些数据为基础，提出了一个研究项目，该项目应该有助于发现药物靶点，并有望转化为EDMD患者的药物治疗策略。本提案概述的实验将导致A)对人类SUN-KASH相互作用组的全面结构和生化理解，B) LINC复合物锚定在层状蛋白层的结构基础，以及C)洞察LINC复合物组装和拆卸的调节。我们期望对这三个目标的追求将对形成EDMD基础的蛋白质网络产生更好的分子描述，从而揭示可能破坏这些过程的药物靶点。我们进一步预期，这项研究将促进我们对核包膜的理解，这将对大量病理性核包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 overaccumulation 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.