Molecular domains determining gap junction channel properties

缝隙连接通道是细胞间特殊的膜孔结构，允许一个细胞中的离子和小信号分子传递给相邻细胞。缝隙连接通道的构件是一组称为连接蛋白(Cx)的跨膜蛋白。在啮齿动物中发现了20种不同类型的连接蛋白。六个相同的连接蛋白或不同的连接蛋白形成一个圆柱形的半通道，称为半通道。两个半槽的背靠背连接形成了一个完整的缝隙连接通道。通过缝隙连接通道的通信对于同步、协调的细胞活动和组织中细胞之间的小分子交换至关重要。缝隙连接通道的孔大小、打开/闭合的开关控制以及化学敏感性因构成通道的连接蛋白亚型而异。这一建议旨在揭示连接蛋白分子的哪一部分负责神经元连接蛋白Cx36的不同功能特性。我们计划将Cx36的几个部分改造成CX50，一种眼晶状体连接蛋白，以形成嵌合体蛋白。这些嵌合体蛋白将被表达到细胞中，用于缝隙连接通道特性的功能测试。进一步细分确定的阳性部位将用于追踪显示独特功能特性所需的最低氨基酸残基(S)。这项研究将有助于我们理解控制这些重要通道开启/关闭的分子机制，以及通道传导离子能力的分子决定因素。全面了解神经元中缝隙连接分子的结构-功能关系，对于理解这些通道在大脑同步活动中的功能至关重要。

Gap junction channels are specialized membrane pore structures between cells, allowing ions and small signalling molecules in one cell to pass on to the neighbour cells. The building blocks for gap junction channels are a group of transmembrane proteins called connexins (Cx). 20 different types of connexins are identified in rodents. Six identical connexins or different connexins form a cylinder-shaped half channel known as a hemichannel. Back-to-back linking of two hemichannels forms a complete gap junction channel. Communication through gap junction channels is essential for synchronized, coordinated cellular activities and exchanging small molecules between cells in tissues. The gap junction channel pore size, the switch control for opening/closing, and the chemical sensitivities are different depending on the connexin subtypes that organize the channel. This proposal aims to reveal which part of the connexin molecule is responsible for the distinct functional properties of the neuronal connexin, Cx36. We plan to engineer several parts of Cx36 into Cx50, an eye lens connexin, to form chimera proteins. These chimera proteins will be expressed into cells for functional test of gap junction channel properties. Further subdivision of positively identified parts will be used to track down the minimum amino acid residue(s) required to display unique functional properties. The proposed research will assist us in understanding the molecular mechanisms in the control of opening/closing of these important channels and the molecular determinants for the ability of the channel to conduct ions. A comprehensive knowledge of structure-function relationships for gap junction molecules in neurons is critical in understanding the functions of these channels in synchronized brain activities.