Kv Channel Interacting Proteins (KChIPs) as calcium sensors for Kv4 channels

钾通道在不同的细胞类型中控制静息膜电位和膜兴奋，这从大量的钾通道类型和亚家族中得到了反映。电压门控钾通道(Kv通道)，尤其是Kv4亚家族的电压门控钾通道，在神经元树突状细胞中参与了心肌细胞的复极化和兴奋的受控传播。心肌细胞膜和神经元树突上的Kv4通道与Kv通道相互作用蛋白(KChIP)形成复合体。KChIPs属于神经元钙感受器(NCS)家族的钙结合EF-Hand蛋白。无论是在心肌细胞中，还是在神经元中，胞浆钙的局部波动都是正常细胞功能和可塑性的中心。然而，胞浆钙高于正常范围可能导致病理生理过程，并最终导致细胞死亡。自从KChIP被发现是Kv4通道的特异性β亚单位以来，基于KChIP属于Ncs蛋白家族的事实，KChIP在Kv4通道中的钙传感器作用一直被归因于KChIPs。关于这一角色的暗示确实存在于文学中。然而，KChIP的钙依赖性构象变化是否影响Kv4/KChIP复合体的形成，或者在现有的Kv4/KChIP复合体中，KChIP的钙依赖性构象变化是否直接传递到Kv4通道，目前还没有得到详细的研究。因此，本研究的目的是从分子水平上阐明KChIPs在生理性和病理性钙波动过程中作为Kv4通道钙感受器的作用。用表位标记的Kv4α-和KChIPβ-亚基的荧光测量，以及在不同胞浆钙浓度存在下对Kv4/KChIP介导的电流的电生理测量，结合使用带有突变的钙结合位点的KChIP，应该可以提供关于这个问题的有价值的信息。

Potassium channels control the resting membrane potential and membrane excitation in very different cell types, as reflected by the huge number of potassium channel types and subfamilies. Voltage-gated potassium channels (Kv channels), particularly those of the Kv4 subfamily, are critically involved in cardiomyocyte repolarisation and the controlled spread of excitation in the dendritic trees of neurons. The Kv4 channels in the membrane of cardiomyocytes and neuronal dendrites form complexes with Kv Channel Interacting Proteins (KChIPs). KChIPs belong to the Neuronal Calcium Sensor (NCS) family of calcium-binding EF-hand proteins. Both in cardiomyocytes and in neurons local fluctuations in cytoplasmic calcium are central to normal cell function and plasticity. However, a rise in cytoplasmic calcium above the normal range may lead to pathophysiological processes and eventually to cell death. From the time when the KChIPs were discovered as being specific β-subunits of Kv4 channels, the role of a calcium sensor for Kv4 channels has been attributed to the KChIPs, based on the fact that they belong to the NCS protein family. Hints on such a role do exist in the literature. However, it has not been studied in detail and it remains unproven whether calcium-dependent conformational changes of the KChIPs influence Kv4/KChIP complex formation or whether in an existing Kv4/KChIP complex calcium-dependent conformational changes of the KChIPs are directly transmitted to the Kv4 channel. Therefore, the aim of the present research proposal is the molecular elucidation of the role of KChIPs as calcium sensors for Kv4 channels during physiologic and pathophysiologic calcium fluctuations. Fluorescence measurements with epitope-tagged Kv4 α- und KChIP β-subunits as well as electrophysiological measurements of Kv4/KChIP-mediated currents in the presence of different cytoplasmic calcium concentrations, combined with the use of KChIPs with mutated calcium binding sites, should provide valuable information regarding this issue.