PERMEABILITY MEDIATED BY CONNEXIN CHANNELS

通过缝隙连接传递细胞间信号的生理学机制是 仍然是个谜。尽管在理解上取得了长足的进步 连接通道的生物化学、遗传学和生物合成， 它们最显著的功能特性-- 信号分子的渗透性--目前尚不清楚。哪一个 化学信号通过连接蛋白通道，有多好？ 影响深远的根本性问题。 缝隙连接通道(由连接蛋白组成)形成调控 介导离子和小分子的细胞间直接转移的途径 分子。对该途径的研究受到其在 孔的两端都在细胞内，大多数人不能接触到 探索渠道选择性的操作。的研究 连接蛋白的渗透倾向于集中在小原子离子或大原子离子上 荧光示踪剂。然而，它是特定之间的选择性 具有重要生物学意义的信号分子。另外， 由于进入途径是通过细胞质，所以很难识别调节作用 直接作用于通道以改变渗透性的因素，而不是 而不是通过蜂窝组件。 长期的目标是理解分子的运作 这条细胞间通讯的路径。方法是 研究重组系统中的连接蛋白通道 可以充分探索选择性渗透特性。本研究 使用连接蛋白32和连接蛋白26形成的通道 来自天然组织的免疫尿素场和表达载体 提供不可用信息的特性良好的系统 来自细胞研究。拟议的实验解决了这些问题 问题：连接蛋白通道是否在信号分子中进行选择 (例如，第二信使)通过特定的分子亲和力？什么 是不同分子的不同的选择性 缝隙连接蛋白？分子大小和电荷如何影响 弥漫在海峡中？什么是直接调制器？ 缝隙连接蛋白通道及其作用机制？ 通过对实验上可获得的连接蛋白通道的研究 系统中，人们希望了解 交汇点通信。缝隙连接非常普遍，以至于 阐明连接蛋白通道的选择性将具有深远的意义 对整个细胞和发育生物学的影响。 目前已知的连接蛋白有12种。遗传性缺陷 连接蛋白32导致周围神经病变，而在连接蛋白43中 心脏发育缺陷。毫无疑问，还有许多其他的综合征 全部或部分由连接蛋白通道功能缺陷引起。 连接蛋白通道的功能缺陷将反映在 细胞间对细胞质的异常渗透性(高或低) 分子。拟议的研究解决了这一问题的基础 可能会发生。

The physiology of intercellular signaling through gap junctions is still a mystery. In spite of considerable progress in understanding the biochemistry, genetics and biosynthesis of junctional channels, the character of their most salient functional property - permeability to signaling molecules - remains unknown. Which chemical signals go through connexin channels, and how well, are fundamental issues with far-reaching impact. Gap junction channels (composed of connexin) form regulated pathways mediating direct intercellular transfer of ions and small molecules. Study of the pathway is constrained by its location in situ; both ends of the pore are intracellular, inaccessible to most manipulations that explore channel selectivity. Studies of connexin permeation tend to focus on small atomic ions or large fluorescent tracers. Yet, it is the selectivity among specific signaling molecules that is of key biological importance. Also, since access is via cytoplasm, it is difficult to identify modulatory factors acting directly on the channel to alter permeability, rather than via cellular components. The long-term objective is to understand the molecular operation of this pathway of intercellular communication. The approach is to study connexin channels in a reconstituted system where their selective permeation properties can be fully explored. This study uses channels formed by connexin32 and connexin26 immunopurifield from native tissues and expression vectors in a well-characterized system that yields information not available from cellular studies. The proposed experiments address these questions: Do connexin channels select among signaling molecules (e.g., second messengers) by specific molecular affinities? What are the distinctive molecular selectivities of the different connexins? How do molecular size and charge affect what can permeate the channels? What are the direct modulators of connexin channels, and their mechanisms of action? By study of connexin channels in an experimentally accessible system, one hopes to understand the fundamental properties of junctional communication. Gap junctions are so widespread that elucidation of connexin channel selectivity will have profound consequences throughout cellular and developmental biology. There are currently 12 known connexins. Genetic defects in connexin32 cause a peripheral neuropathy, and in connexin43 defects of cardiac development. No doubt many other syndromes arise in toto or in part from defects in connexin channel function. A functional ~defect~ of connexin channels will be reflected in an abnormal intercellular permeability (high or low) to a cytoplasmic molecule. The proposed studies address the basis for how this may occur.