氯通道、水通道和周期蛋白在细胞周期中的相互作用研究

It’s been confirmed by us and others that ClC-3, a candidate of the volume-sensitive chloride channels associated with the regulation of cell cycle process is up-regulated in expression and/or functional activities, and plays important roles in regulation of cell proliferation, migration and invasion in tumor cells including nasopharyngeal carcinoma cells. Our recent results demonstrated that the activities of ClC-3 were modulated by aquaporin-3 in regulatory volume decrease; ClC-3, as a downstream target of cyclin D1, could take control of G1/S transition. These data suggest that ClC-3 could interact with the function-related proteins to regulate the biological activities of cells. ClC-3, aquaporin-3 and cyclins are key regulators of the cell cycle. However, the interactions between them in regulation of cell cycle progress are far from clarified. In this study, multiple techniques and methods including bioinformatics, molecular dynamics simulation and experimental investigation will be employed to study in the poorly differentiated nasopharyngeal carcinoma cells: 1) the cell cycle phase-related distributions and transportations of ClC-3 and aquaporin-3; 2) the interaction between ClC-3 and aquaporin-3 in multiple levels including transcription, translation and function; 3) the interaction between channel proteins and cyclins and its action mechanism; and 4) finally the mechanism of the cell cycle progress being coordinated by ClC-3, aquaporin-3 and cyclins. This is beneficial to accumulate theoretical knowledge related to their application in cancer prediction, diagnosis, treatment and drug development.

我们及其他人研究证实ClC-3是具有细胞周期调节功能的容积敏感性氯通道，在鼻咽癌等多种肿瘤中异常表达和/或活性改变，影响肿瘤细胞增殖、迁移及侵袭等过程。近期我们发现：ClC-3在细胞调节性容积减小过程中受水通道AQP-3调节；作为细胞周期蛋白D1的下游靶蛋白，参与调控细胞G1/S期过渡。这些结果提示ClC-3可与其他蛋白质相互作用，调节细胞某种功能行为。ClC-3、AQP-3和周期蛋白都是细胞周期调控的关键蛋白，然而对三者在调控细胞周期时的相互作用知之甚少。因此，本课题拟综合利用生物信息学、分子动力学模拟及实验观察手段，首先探讨ClC-3和AQP-3在鼻咽癌细胞内位相分布及其转运，以及在基因转录、翻译和功能上的交互影响，然后研究通道与周期蛋白间的相互作用及其机制，最终分析它们相互协同影响细胞周期进程的途径，为探讨它们及其相互作用在肿瘤预测、诊断、治疗、药物开发等方面的应用积累相关理论知识。

细胞容积调节是细胞应对各种内外环境变化，及参与细胞各种功能过程中维持其体积相对稳定的机制，是细胞生存的先决条件。细胞膜蛋白水通道AQP-3和氯通道ClC-3，在鼻咽癌等多种肿瘤中异常表达和/或活性改变，影响肿瘤细胞增殖、迁移及侵袭等过程。我们采用分子生物学、膜片钳、激光共聚焦显微镜等技术手段，明确了AQP-3和ClC-3之间存在相互作用关系；在细胞容积调节和非细胞容积调节过程中，AQP-3可能作为生物传感器或者调节因子调控ClC-3功能。利用生物信息学和表面等离子体共振成像技术（Surface Plasmon Resonance imaging, SPRi）技术，筛选出了二者间的可能相互作用面；以此为基础，经3D同源建模后发现与ClC-3结合的AQP-3胞外段某区域具有典型的“口袋”结构，适合作为药物靶点。目前，我们正在借助计算机辅助药物设计系统，针对该区域筛选小分子化合物，拟开展研究二者相互作用面对肿瘤增殖、迁移及浸润等影响。