Significance of the novel Bok-IP3 receptor interaction

肌醇1，4，5-三磷酸受体(IP3Rs)在脊椎动物细胞内信号转导中起着关键作用。 它们是在内质网中形成四聚体钙通道的三个非常相似的蛋白质家族 (呃)膜。在过去的二十年里，我一直在研究IP3R的分子生物学，最近 发现BOK是一个非常鲜为人知的独特功能的Bcl-2蛋白家族成员，它与非常 强烈支持IP3R。Bcl2蛋白家族控制着固有的细胞凋亡途径，是一种 从基础细胞生物学的角度和作为抗癌药物的靶标进行了研究。博克是最 与特征明确的促凋亡蛋白Bax和Bak相似，但其细胞作用仍是谜 极具争议性；然而，共识是BOK定位于ER膜，并在 细胞凋亡信号。在过去的3年里，我的实验室开始表征新的BOK-IP3R相互作用 以及它的意义，但仍有许多工作要做。现在，通过三个具体目标，我计划测试 假设BOK与IP3Rs结合调节IP3R功能，这是BOK在其中发挥作用所必需的 细胞凋亡，严重依赖于新开发的细胞系，其中BOK已使用CRISPR/Cas9-1删除 基于方法。 目的1.BOK在IP3R1上结合位点的解析及IP3R1结合缺陷的研究 抑制BOK-IP3R1相互作用的结构和肽。所获得的结果将为我们深入了解 BOK与IP3R交互，并将生成有助于探测BOK-IP3R交互意义的工具。 目的2.确定BOK与IP3Rs结合是否调节IP3R功能。所获得的结果应该是一种 新的IP3R调控模式，并为BOK影响细胞凋亡的机制提供了洞察力。 目的3.明确BOK在细胞凋亡信号转导中的作用以及BOK-IP3R相互作用对这一作用的重要性。 所获得的结果将与来自其他特定目标的数据相结合，以生成如何BOK的模型 影响细胞凋亡，并将确定BOK-IP3R界面是否为药物靶点。 总而言之，这项提案是第一次努力确定 BOK-IP3R的相互作用，无疑将有助于了解IP3R的功能和BOK的细胞作用。这项工作 这将促进我们对细胞信号和细胞凋亡的理解，并可能确定一个新的治疗靶点。

Inositol 1,4,5-trisphosphate receptors (IP3Rs) play a pivotal role in vertebrate intracellular signaling. They are a family of three very similar proteins that form tetrameric Ca2+ channels in endoplasmic reticulum (ER) membranes. For the last two decades I have been studying the molecular biology of IP3Rs and recently discovered that Bok, a very poorly understood Bcl-2 protein family member with unique features, binds very strongly to IP3Rs. The Bcl-2 protein family controls the intrinsic apoptosis pathway and is intensively investigated, both from a basic cell biology perspective, and as a target for anti-cancer drugs. Bok is most similar to the well-characterized pro-apoptotic proteins Bax and Bak, but its cellular role remains enigmatic and highly controversial; consensus is, however, that Bok is localized to the ER membrane and plays a role in apoptotic signaling. In the last 3 years, my laboratory has begun to characterize the novel Bok-IP3R interaction and its significance, but there is still much work to do. Now, through three Specific Aims, I plan to test the hypotheses that Bok binding to IP3Rs regulates IP3R function and is required for the role that Bok plays in apoptosis, leaning heavily on newly-developed cell lines in which Bok has been deleted using CRISPR/Cas9- based methods. Aim 1. Resolution of the binding site for Bok on IP3R1 and the development of binding-deficient IP3R1 constructs and peptides that inhibit the Bok-IP3R1 interaction. Results obtained will provide insight into how Bok interacts with IP3Rs and will generate tools useful for probing the significance of the Bok-IP3R interaction. Aim 2. Define whether Bok binding to IP3Rs regulates IP3R function. Results obtained should characterize a new mode of IP3R regulation and provide insight into the mechanism by which Bok influences apoptosis. Aim 3. Define the role of Bok in apoptotic signaling and the importance of the Bok-IP3R interaction to this role. Results obtained will be integrated with the data from the other Specific Aims to generate a model for how Bok influences apoptosis and will establish whether the Bok-IP3R interface is a drug target. In summary, this proposal represents the first effort at characterizing the nature and significance of the Bok-IP3R interaction and will undoubtedly shed light on IP3R function and the cellular role of Bok. The work will both advance our understanding of cell signaling and apoptosis, and may identify a new therapeutic target.