Proteolytic processing of sodium channel beta1 subunits

钠通道β1亚基的蛋白水解加工

• 批准号: 1643216
• 金额: --
• 依托单位: University of York
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2015
• 资助国家: 英国
• 起止时间: 2015 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1643216
• 关键词: Proteolytic; processing; sodium; channel; beta1

Voltage-gated Na+ channels (VGSCs) are responsible for action potentials in electrically excitable cells, including neurons and muscle cells. VGSCs contain a pore-forming alpha subunit with one or two auxiliary beta subunits. The beta subunits regulate channel activity and are unique amongst other ion channel auxiliary subunits because they are also cell adhesion molecules. VGSCs are therefore not only ion channels, but also cell adhesion complexes. In particular, the beta1 subunit regulates action potential firing, neurite outgrowth and neuronal migration during central nervous system development. Interestingly, beta1 is also expressed in breast cancer cells, where it regulates cellular migration, invasion and metastasis. VGSC beta subunits contain cleavage sites for processing by secretases that are involved in Alzheimer's disease pathology. Secretase activity is also emerging as an important target in breast cancer. However, the functional consequences of proteolytic processing of beta1 are not understood. The aim of this project is to test the hypothesis that beta1 subunits are cleaved by secretases and that this regulates adhesion, neurite outgrowth, cellular migration and electrical activity. We will use a range of sophisticated ensemble and single-molecule microscopy approaches, e.g. confocal microscopy, TIRF microscopy, FRAP to explore the stoichiometry and cycling of beta1 subunits, in neurons and breast cancer cells. We will modulate secretase activity using drugs and by generating beta1 mutants in which the cleavage sites have been modified. We will study the functional consequences of beta1 processing on gene expression using molecular approaches such as chromatin immunoprecipitation (ChIP). Importantly, we will study the effect of proteolytic processing on cellular migration and channel function using cell migration assays and whole cell patch clamp electrophysiological recording. The project will therefore expose the student to a range of cutting-edge cell biology techniques in labs that are leading in this field. As beta1 plays a key role in brain development and in a number of diseases, this project is expected to provide novel mechanistic insights into a potential therapeutic target.

电压门控Na+通道（VGSC）负责电可兴奋细胞（包括神经元和肌肉细胞）中的动作电位。VGSC含有成孔α亚基和一个或两个辅助β亚基。β亚基调节通道活性，并且在其他离子通道辅助亚基中是独特的，因为它们也是细胞粘附分子。因此，VGSC不仅是离子通道，而且是细胞粘附复合物。特别是，β 1亚基调节动作电位放电，神经突生长和神经元迁移在中枢神经系统发育。有趣的是，β 1也在乳腺癌细胞中表达，它调节细胞迁移，侵袭和转移。VGSC β亚基含有用于由参与阿尔茨海默病病理学的分泌酶加工的切割位点。分泌酶活性也成为乳腺癌的重要靶点。然而，β 1的蛋白水解加工的功能后果还不清楚。该项目的目的是测试β 1亚基被分泌酶切割并调节粘附、神经突生长、细胞迁移和电活动的假设。我们将使用一系列复杂的集成和单分子显微镜方法，例如共聚焦显微镜，TIRF显微镜，FRAP来探索神经元和乳腺癌细胞中β 1亚基的化学计量和循环。我们将使用药物调节分泌酶的活性，并通过产生β 1突变体，其中切割位点已被修改。我们将研究β 1加工对基因表达的功能后果，使用分子方法，如染色质免疫沉淀（ChIP）。重要的是，我们将研究蛋白水解加工对细胞迁移和通道功能的影响，使用细胞迁移试验和全细胞膜片钳电生理记录。因此，该项目将使学生在该领域领先的实验室中接触到一系列尖端的细胞生物学技术。由于β 1在大脑发育和许多疾病中起着关键作用，该项目有望为潜在的治疗靶点提供新的机制见解。