Regulation of L-type calcium channel inactivation

钙离子通过钙通道进入大脑神经元，钙通道是由膜电压变化激活的，这一过程对从基因转录到膜兴奋性等功能至关重要。事实上，钙作为一种信号分子的重要性决定了系统需要精确调节钙进入的时间和程度。“l型”（Cav1）钙通道在大脑中广泛表达，并受钙调蛋白（CaM）调控，钙调蛋白是一种通过“钙依赖性失活”（CDI）或“钙依赖性促进”（CDF）过程修饰Cav1钙内流的分子。最近的研究表明，通过与通道C末端结合以降低CDI的辅助蛋白，可以显著增加cav1介导的钙内流。两个例子是支架蛋白致密素，它与C端结合以招募特殊的激酶CaMKII，第二个例子是钙敏感蛋白caldendrin，它在物理上取代Cav1通道上的CaM。我们有一种新的Cav1.3通道促进形式的初步数据，该形式持续bb50秒（称为L-CDF），随后是电路活动的简短输入特征序列。此外，Cav1.3 L-CDF依赖于densin和CaMKII的表达。这很重要，因为这些蛋白都在CA1海马锥体细胞中表达，在这里，短暂的突触输入后钙通道活性的“延迟促进”增强了钙的流入。我们假设，辅助蛋白致密蛋白和胼胝体蛋白是延迟促进的基础，促进了锥体细胞中钙流入的大量增加。我们将应用分子生物学、生物化学和电生理学等技术来研究模型系统和啮齿动物脑组织切片中表达的l型钙通道，以确定致密素和caldendrin在产生Cav1通道L-CDF中的作用。总之，这项工作代表了我们实现长期目标的第一步，即了解l型钙通道如何被调节以控制内部钙和第二信使途径的激活。Cav1通道在从大脑到肌肉和心脏的所有组织中的广泛表达，将确保该结果对于理解钙内流在多种情况下是如何控制的立即重要，推进对体内可兴奋细胞的功能如何由这些蛋白质-蛋白质相互作用调节的知识。**

Calcium ions enter neurons in the brain through calcium channels activated by a change in membrane voltage, a process that is critical to functions ranging from gene transcription to membrane excitability. Indeed, the importance of calcium as a signaling molecule dictates the need for systems to precisely regulate the timing and extent of calcium entry. The “L-type” (Cav1) class of calcium channels is widely expressed in the brain and is regulated by calmodulin (CaM), a molecule that modifies Cav1 calcium influx through a process of “calcium-dependent inactivation” (CDI) or “calcium-dependent facilitation” (CDF). Recent work reveals that Cav1-mediated calcium influx can be substantially increased by accessory proteins that bind to the channel C terminus to reduce CDI. Two examples are the scaffolding protein densin that binds to the C terminus to recruit the specialized kinase CaMKII, and a second is the calcium sensitive protein caldendrin that physically displaces CaM on the Cav1 channel. We have preliminary data for a novel form of Cav1.3 channel facilitation that lasts for >5 seconds (termed L-CDF) following a brief train of input characteristic of circuit activity. Moreover, Cav1.3 L-CDF depends on expression of densin and CaMKII. This is important, as each of these proteins are expressed in CA1 hippocampal pyramidal cells, where a “delayed facilitation” of calcium channel activity after a brief train of synaptic input enhances calcium influx. We hypothesize that the accessory proteins densin and caldendrin underlie the delayed facilitation that promotes a large increase in calcium influx in pyramidal cells. We will apply techniques ranging from molecular biology to biochemistry to electrophysiology to study L-type calcium channels expressed in a model system and in slices of rodent brain tissue to determine the role of densin and caldendrin in producing Cav1 channel L-CDF. Together this work represents the first step of attaining our long term goal of*understanding how L-type calcium channels are regulated to control*internal calcium and activation of second messenger pathways. The widespread expression of Cav1 channels in all tissues ranging from brain to muscle and heart will ensure that the results will be immediately important to understanding how calcium influx is controlled in multiple contexts, advancing knowledge of how the function of excitable cells in the body is regulated by these protein-protein interactions. **