Molecular mechanisms for regulation of HCN channels by TRIP8b subunits

描述(由申请人提供)：神经元对突触输入产生动作电位的基本能力在很大程度上取决于其树突的电特性。调节树突电学性质的一个关键离子通道是超极化激活的环核苷酸门控(HCN)离子通道。HCN通道在兴奋性的阈值下工作，因此通道的数量、定位、电压依赖和环核苷酸依赖的微小变化可以对细胞的兴奋性产生巨大影响。我们的长期目标是了解HCN通道如何控制神经元兴奋性的分子机制。最近在神经元中发现了HCN通道的一个辅助亚基，称为TRIP8b。TRIP8b对HCN通道的转运、电压依赖和环核苷酸依赖有深远的影响。在这项授权中，我们建议研究TRIP8b如何与HCN通道结合并调节通道功能的分子机制。我们的方法将是结合X射线结晶学，以获得原子分辨结构信息，与电生理学和荧光相结合，研究其天然膜环境中的功能通道。我们的实验将揭示HCN2C-末端区域与TRIP8b在原子分辨率上的相互作用的结构，相互作用的化学计量比，以及TRIP8b调节HCN2的环核苷酸依赖和电压依赖的门控的机制。这些发现将为我们理解HCN通道的结构和调控提供重要的进展，因为它们存在于神经元中，并进一步阐明它们在脑生理和病理中的作用。 与公共健康相关：离子通道是大脑的晶体管，因此控制着从我们的感觉到我们的思想的一切。超极化激活的环核苷酸门控(HCN)通道从根本上参与了大脑神经元的电兴奋性，其功能障碍与癫痫等神经系统疾病有关。我们的长期目标是了解HCN通道如何控制神经元兴奋性的分子机制，以使我们能够开发针对神经疾病的靶向治疗。

DESCRIPTION (provided by applicant): The fundamental ability of a neuron to fire action potentials in response to synaptic input is largely determined by the electrical properties of its dendrites. One key ion channel that regulates the electrical properties of dendrites is the hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channel. HCN channels operate at the threshold of excitability so small changes in the number, localization, voltage dependence, and cyclic nucleotide dependence of the channels can have a dramatic impact on the excitability of the cell. Our long term goal is to understand the molecular mechanisms for how HCN channels control neuronal excitability. Recently an auxiliary subunit of HCN channels in neurons was discovered, called TRIP8b. TRIP8b has a profound effect on the trafficking, voltage dependence, and cyclic nucleotide dependence of the HCN channels. In this grant, we propose to study the molecular mechanism for how TRIP8b binds to the HCN channel and regulates channel function. Our approach will be to combine x-ray crystallography, for atomic resolution structural information, with electrophysiology and fluorescence to study the functional channel in its native membrane environment. Our experiments will reveal the structure of the interaction between the HCN2 C-terminal region and TRIP8b at atomic resolution, the stoichiometry of the interaction, and the mechanism for TRIP8b regulation of the cyclic nucleotide-dependent and voltage- dependent gating of HCN2. These findings will provide a significant advance in our understanding of the structure and regulation of HCN channels as they exist in the neuron and further illuminate their role in the physiology and pathology of the brain. PUBLIC HEALTH RELEVANCE: Ion channels are the transistors of the brain and thereby control everything from our senses to our thoughts. Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels are fundamentally involved in the electrical excitability of the neurons in our brain, and their dysfunction is responsible for some neurological diseases such as epilepsy. Our long term goal is to understand the molecular mechanisms for how HCN channels control neuronal excitability to enable us to develop targeted therapies for neurological diseases.