Deciphering the functional role of actin-spectrin-based membrane skeleton in subcellular compartmentalization of signaling proteins and cell signal transduction

项目摘要/摘要： 受体酪氨酸激酶(RTK)、G蛋白偶联受体(GPCRs)和细胞黏附分子(CAM) 是所有真核细胞中的三个主要的细胞表面蛋白家族，共同代表着最初的第一 细胞响应细胞外刺激并启动各种信号通路以 从而调节细胞增殖和分化，促进细胞存活，调节细胞 新陈代谢和细胞间的交流。影响这些信号通路的突变导致许多人类 症状和疾病，如各种类型的神经退行性疾病和癌症。临床部 这些信号蛋白的重要性推动了靶向治疗的发展，旨在阻止 膜受体的激活和下游信号转导。越来越多的证据表明 提示这三个膜蛋白家族之间存在显著的信号串扰。 质膜水平，这些蛋白质可以形成高度组织的膜微或纳米簇，与 独特的生化和生物物理特性，决定了信号结果。然而，分子 膜相关信号蛋白的这种串扰和区隔的机制 启动和维护以调节下行信令的灵敏度和特异度 难以捉摸。我们最近发现的一个新发现的基于肌动蛋白的膜相关周期骨架 (MPS)结构作为神经元中GPCRs和CaM反式激活RTK的信号平台提供了 对这些细胞表面蛋白之间的协同作用如何协调的分子洞察力 上升到下行信令。这项提议目标是将超分辨率成像、细胞和 分子生物学工具和质谱分析，以研究独特的物理分子 通过识别关键的分子相互作用，负责MPS介导的细胞信号传递的机制 负责MPS依赖的信号蛋白簇(即信号)的组装和拆解 蛋白质复合体)，并研究液-液相分离、受体内吞和接触的作用 MPS中质膜和胞内膜结合的胞内细胞器之间的位置 神经元中介导的细胞信号传递。由于基于血影蛋白-肌动蛋白的MPS结构可能存在于其他分化的 例如淋巴细胞等细胞类型，从而控制淋巴细胞的发育和激活，我们的分析将 也被扩展到研究MPS在免疫反应过程中淋巴细胞信号转导中的作用 拟议的研究不仅将拓宽我们对细胞信号转导的基本理解， 神经元和免疫细胞中信号蛋白的膜骨架和相分离行为， 也有助于提出治疗人类疾病的潜在药物靶点，包括神经退行性疾病和 癌症。

Project Summary/Abstract: Receptor tyrosine kinases (RTKs), G-protein-coupled receptors (GPCRs), and cell adhesion molecules (CAMs) are three major families of cell surface proteins in all eukaryotic cells, and together represent the primary first responders for cells to respond an extracellular stimulus and initiates a variety of signaling pathways to subsequently regulate cell proliferation and differentiation, promote cell survival, and modulate cellular metabolism and cell-to-cell communication. Mutations affecting these signaling pathways result in many human syndromes and diseases, such as various types of neurodegenerative disorders and cancer. The clinical importance of these signaling proteins has motivated the development of targeted therapies designed to block the activation of the membrane receptors and the downstream signal transduction. Increasing evidence has suggested that there is significant signaling crosstalk among these three membrane protein families at the plasma membrane level and these proteins can form highly organized membrane micro- or nano-clusters with unique biochemical and biophysical properties, dictating the signaling outcome. However, the molecular mechanisms by which how such crosstalk and compartmentalization of membrane-associated signaling proteins are initiated and maintained to modulate the sensitivity and specificity of the downstream signaling remain largely elusive. Our recent discovery of a newly identified actin-spectrin-based membrane-associated periodic skeleton (MPS) structure being a signaling platform for RTK transactivation by GPCRs and CAMs in neurons provides molecular insights into how the cooperative action among these cell surface proteins can be coordinated to give rise to the downstream signaling. The objective of this proposal is to combine super-resolution imaging, cell and molecular biology tools, and mass spectrometry analyses, to investigate the distinctively physical molecular mechanisms responsible for the MPS-mediated cell signaling, by identifying the key molecular interactions responsible for the MPS-dependent assembly and disassembly of the signaling protein clusters (i.e., signaling protein complexes) and examining the roles of liquid-liquid phase separation, receptor endocytosis, and contact sites between the plasma membrane and intracellular membrane-bound intracellular organelles in the MPS- mediated cell signaling in neurons. As the spectrin-actin based MPS structures likely exist in other differentiated cell types such as lymphocytes and thereby control lymphocyte development and activation, our analyses will also be extended to examine the role of the MPS in lymphocyte signaling during immune responses Our proposed research will not only broaden our fundamental understanding of cell signal transduction controlled by the membrane skeleton and the phase separation behaviors of signaling proteins in neurons and immune cells, but also help suggest potential drug targets for human diseases including neurodegenerative diseases and cancer.