Chemical tools to decode nuclear lamina-ome

项目摘要 这项应用的目标是成像和识别活细胞中的核膜相互作用蛋白。这个 核层位于内核膜下，由四种主要同源物质组成。 蛋白质，层蛋白A(LA)、层蛋白B1(LB1)、层蛋白B2(LB2)和层蛋白C(LC)。椎板是V型中间体 细丝蛋白，并为哺乳动物的细胞核提供机械支持。然而，Lamins的作用 已经显著扩展到细胞生物学的其他关键方面，包括机械传感dna 修复、染色质调节、基因转录和干细胞调节。在这些过程中，层的作用 核内复杂但尚未完全了解的蛋白质-蛋白质相互作用参与了这一过程。 椎板。LA在这些生物过程中的重要性进一步得到强调，因为更多的 已知有450多个LA突变会导致一系列统称为椎板病的疾病。 应对这一挑战的主要障碍之一是缺乏适当的工具和 生理学相关条件下活细胞膜结合蛋白的成像和鉴定技术 条件。我们最近开发了一种名为LBL1的小分子，它能在整个 细胞蛋白质组。我们对LBL1的识别为我们提供了前所未有的机会来形象和 鉴定活细胞中的内源性层粘连蛋白结合蛋白。这种理解将产生新的 对Lamin功能的深入了解，并可能为开发治疗方法提供新的途径 椎板病症。为了实现这一目标，我们提出了以下两个具体目标：1)开发一个 创新的化学探针标记活细胞中的核层；2)识别和成像LA及其 活细胞中相互作用的伙伴。

Project Summary The goal of this application is to image and identify nuclear lamina-interacting proteins in living cells. The nuclear lamina, located underneath of inner nuclear envelope, is composed of four major homologous proteins, lamin A (LA), lamin B1 (LB1), lamin B2 (LB2) and lamin C (LC). Lamins are type V intermediate filament proteins and provide mechanical support for the mammalian nucleus. However, the roles of lamins have been significant extended into other critical aspects of cell biology including mechanosensing DNA repair, chromatin regulation, gene transcription and stem cell regulation. The function of lamins in these processes are mediated by complex yet incompletely understood protein-protein interactions at the nuclear lamina. The importance of LA in these biological processes is further emphasized by the fact that more than 450 LA mutations are known to cause a wide spectrum of diseases collectively called laminopathies. One of the major obstacles in addressing this challenge is the lack of the appropriate tools and technologies to image and identify the lamina-binding proteins in living cells under physiologically relevant conditions. We recently developed a small molecule called LBL1 that specifically binds lamins in the whole cellular proteome. Our identification of LBL1 provides an unprecedented opportunity for us to image and identify the endogenous lamin-binding proteins in living cells. This understanding will generate novel insights into the lamin functions and potentially provide novel avenues to develop therapies for laminopathies. To achieve this goal, we propose the following two specific aims: 1) To develop an innovative chemical probe to label nuclear lamina in living cells; 2) To identify and image LA and its interacting partners in living cells.