Function of L-Type Ca2+ Channels in B-Lymphocytes

描述（由申请人提供）：钙（Ca2+）作为一种普遍的信使，在生物学中发挥着核心作用，通过调节其细胞内水平，使细胞有效地响应细胞外信号。因此，细胞膜对Ca2+离子的通透性是由称为离子通道的膜蛋白精心调节的。有效免疫反应的建立也需要免疫细胞（如淋巴细胞）细胞内Ca2+升高的延长期。b淋巴细胞是适应性免疫系统的细胞，负责产生抗体。B淋巴细胞中表达的离子通道的多样性比最初预期的要大，包括l型cav通道变体等分子，这些分子最初被认为是所谓的可兴奋细胞（如心脏或神经元细胞）所特有的。虽然这些l型通道在b细胞中的存在已被证实，但它们的功能仍有待阐明。本提案的目的是建立遗传模型系统，使我们能够研究l型通道家族个体成员在b细胞环境中的作用。这包括缺乏l型通道的b细胞系的产生和功能表征，这些b细胞系随后可以与野生型和突变型通道互补。我们还计划通过在现有小鼠模型中研究b细胞限制性基因组缺失对l型通道的影响来分析这些分子在体内的功能。由于免疫反应是机体多个部位众多细胞类型非常复杂的相互作用，这种体内模型代表了细胞系的一种补充方法，使我们能够在完整生物体的背景下研究l型通道在b细胞发育和激活中的潜在作用。公共卫生相关性声明：l型Ca2+通道是一类药物的重要治疗靶点，包括维拉帕米和地尔硫卓，这类药物通常用于治疗高血压等疾病。它已经在体外显示，这些药物也抑制免疫细胞的Ca2+反应，虽然在更高的浓度比这些应用治疗。这表明存在对这些化合物不太敏感的这些通道的免疫特异性变体。因此，设计这些已知通道抑制剂的修饰版本，以特异性地结合和抑制l型通道的免疫系统版本，是免疫调节药物开发的一个有希望的途径。为此，有必要确定这些通道在免疫环境中表达的功能变异，以及它们在免疫应答激活过程中的确切作用。此外，这些发现也可能对患有影响l型通道功能的罕见复杂遗传疾病的年轻患者有益，这些患者已知表现出与免疫缺陷一致的症状。该建议将有助于通过研究这些通道在b淋巴细胞中的作用来回答这些重要问题。

DESCRIPTION (provided by applicant): Calcium (Ca2+) plays a central role in Biology as a universal messenger by allowing cells to efficiently respond to extracellular signals via modulation of its intracellular level. The permeability of cellular membranes for Ca2+ ions is therefore carefully regulated by membrane proteins called ion channels. The mounting of an efficient immune response also requires an extended period of intracellular Ca2+ elevation in immune cells such as lymphocytes. B-lymphocytes are the cells of the adaptive immune system that are responsible for the production of antibodies. The diversity of ion channels expressed in B lymphocytes is greater than originally anticipated, and includes molecules such as L-type CaV-channel variants that were originally assumed to be specific for so-called excitable cells, such as cardiac or neuronal cells. Although the presence of these L-type channels in B-cells is well established, their function remains to be elucidated. The aim of this proposal is to establish genetic model systems allowing us to investigate the role of individual members of the L-type channel family in the B-cell context. This includes the generation and functional characterization of B-cell lines lacking L-type channels that can subsequently be complemented with wildtype and mutant versions of the channels. We also plan to analyze the function of these molecules in vivo by studying the effect of B-cell restricted genomic deletion of L-type channels in an existing mouse model. Because the immune response is a very complex interplay of numerous cell types in multiple locations of the body, this in vivo model represents a complementary approach to the cell lines that will allow us to study the potential role of L-type channels in B-cell development and activation in the context of the complete organism. Public Health Relevance Statement: L-type Ca2+ channels are important therapeutic targets of a class of drugs that include verapamil and diltiazem, which are commonly used to treat conditions such as hypertension in patients. It has been shown in vitro that these drugs also inhibit the Ca2+-response of immune cells, although at higher concentrations than these applied therapeutically. This suggests that immune specific variants of these channels exist that are less sensitive towards these compounds. A promising avenue of immuno-modulatory drug development is therefore to design modified versions of these known channel inhibitors that would specifically bind and inhibit the immune system versions of L-type channels. To this aim, it is essential to identify the functional variants of these channels expressed in the immune context, and their precise role during immune response activation. Furthermore, these findings could also be beneficial to young patients suffering from rare complex genetic disorders affecting L-type channel function, and who are known to show symptoms consistent with immuno-deficiencies. This proposal will contribute to answering these important questions by investigating the role of these channels in B-lymphocytes.