Mechanisms behind the clustering of BK and calcium channels

项目摘要 大钾离子通道（BK）是一类非典型的钾离子通道，因其电导率大而得名 通过电压和钙的协同作用。BK通道在几乎所有器官中都有表达，在神经系统中具有重要作用。 神经、胃肠、生殖、视觉、内分泌、泌尿、淋巴、骨骼肌的功能，以及 心血管系统BK通道的激活需要钙与细胞质C-受体的变构结合。 通道的终端。因此，空间定位在质膜和接近钙源 是调节BK通道功能的两个关键因素。我以前的工作使用超分辨率显微镜， 表明，而不是均匀分布在质膜，BK通道组织成 大型集群此外，我证明了这些BK通道簇组织成更高的层次结构， 钙离子通道与电压门控钙通道（CaV）的排列，CaV是其激活的主要钙源。 结合电生理学和药理学，我确定这种BK-CaV共聚类是BK-CaV的关键。 BK通道在生理超极化电压下的激活。然而， BK通道簇的形成和与特定CaV通道的共簇仍然是神秘的。研究 我的实验室现在以我的发现为基础，专注于确定BK-BK和BK背后的分子机制， CaV通道聚集和聚集在BK功能微调中的作用。利用我们的专业知识， 超分辨率显微镜，定量荧光，离子通道生物物理学和脂质代谢，我们有 确定了三个研究领域，这将促进我们对BK通道背后机制的理解 聚类研究领域1将重点研究介导形成和维持的机制 BK-BK簇-为此，我们将系统地评估BK通道C-末端的作用和 BK β和γ辅助亚基。研究领域2将集中在介导BK-CaV通道共- 聚类-为此，我们已经确定AKAP 150和RIB 2作为两个强有力的候选者， 通道，可以作为共同聚类的基础。研究领域3将集中在一个探索不足的主题， 膜脂质组成的BK-CaV通道的集群-我将利用我的专业知识在研究 磷酸肌醇和胆固醇对离子通道的调节，以引导这两种信号传导的研究 脂质参与离子通道聚集。总的来说，这项研究计划旨在提供新的见解 BK通道簇是如何在钙源附近形成和维持的。从长远来看，这一建议 这将为研究BK通道在健康和疾病中的调节机制开辟新的途径。

Project Summary Big potassium (BK) channels, named after their "Big K+" conductance, are atypical potassium channels activated synergistically by voltage and calcium. Expressed in nearly all organs, BK channels have important roles in the function of the nervous, gastrointestinal, reproductive, visual, endocrine, urinary, lymphatic, skeletal muscle, and cardiovascular systems. BK channel activation requires the allosteric binding of calcium to the cytoplasmic C- terminal of the channel. Therefore, spatial localization at the plasma membrane and proximity to calcium sources are two critical factors regulating BK channel function. My previous work using super-resolution microscopy has demonstrated that rather to be homogeneously distributed at the plasma membrane, BK channels organize into large clusters. In addition, I demonstrated that these BK channel clusters organize into a higher hierarchical arrangement with voltage-gated calcium channels (CaV), which are their primary calcium source for activation. Combining electrophysiology and pharmacology, I established that this BK-CaV co-clustering is key for the activation of BK channels at physiological hyperpolarized voltages. Yet, the molecular mechanisms that mediate BK channel cluster formation and the co-clustering with specific CaV channels remain mysterious. Research in my lab now builds on my discoveries and focus on identifying the molecular mechanisms behind BK-BK and BK- CaV channel clustering and the role of clustering in the fine-tuning of BK function. Leveraging our expertise in super-resolution microscopy, quantitative fluorometry, ion channel biophysics, and lipid metabolism, we have identified three research areas that will advance our understanding of the mechanisms behind BK channel clustering. Research area 1 will focus on the study of the mechanisms mediating the formation and maintenance of BK-BK clusters – for this, we will systematically evaluate the role of the BK channel C-terminal and the role of BK β and γ auxiliary subunits. Research Area 2 will focus on the mechanisms mediating BK-CaV channel co- clustering – for this, we have identified AKAP150 and RIB2 as two strong candidates that interact with both channels and can underlie the co-clustering. Research Area 3 will focus in a poorly explored topic, the role of membrane lipid composition in the clustering of BK-CaV channels – I will leverage my expertise in the study of ion-channel modulation by phosphoinositides and cholesterol to lead the research on how these two signaling lipids are involved in ion channel clustering. Overall, this research program is designed to provide new insights into how BK channel clusters are formed and maintained near calcium sources. In the long-term, this proposal will open new avenues for the study of mechanisms modulating BK channel function in health and disease.