Deactivation mechanisms of rod phototransduction

描述（由申请人提供）：视网膜视杆细胞和视锥细胞对光子的吸收激活一系列生化反应（光导级联），产生对光的电反应。级联的激活和失活最终限制了转导信号的振幅和动力学，从而限制了视觉的灵敏度和时间分辨率。本研究的总体目标是了解在完整的小鼠光感受器中关闭光反应的机制。基因靶向技术将被用于操纵一组蛋白质的功能，这些蛋白质被认为在级联失活中起关键作用，并且单个杆状细胞的光反应的最终变化将通过电记录来确定。使用这种方法，我们将解决3个重要问题：(1)视紫红质磷酸化的速度有多快，是什么决定了这个时间进程？(2)阻滞易位对光响应的功能影响是什么？(3) RGS9复合体的光受体特异性剪接变体是否唯一适合于使转导蛋白/PDE失活，以及如何使其失活？4)在光适应过程中加速转导蛋白/PDE失活的机制是什么？这项研究解决了视网膜疾病小组（http://www.nei.nih.gov/strategicplanning/np_retinal.asp#obj）建议的目标之一，即“分析光转导后光适应和恢复的机制，并了解光/暗适应中神经编码的变化。”这项研究将有助于阐明正常视觉过程的初始步骤，以及由失活失败引起的疾病的发病机制，例如某些形式的视网膜色素变性和夜盲症。在更广泛的背景下，这些实验将提供对G蛋白级联失活机制的见解，所有真核细胞都使用G蛋白级联将细胞外信号转导为细胞内反应。

DESCRIPTION (provided by applicant): The absorption of photons in rods and cones of the retina activates a cascade of biochemical reactions (phototransduction cascade) that generates the electrical response to light. The activation and deactivation of the cascade ultimately limits the amplitude and kinetics of the transduced signal, and thus the sensitivity and temporal resolution of vision. The overall goal of this study is to understand the mechanisms that turn off the light response in intact mouse photoreceptors. Gene targeting techniques will be used to manipulate the function of a subset of proteins that have been suggested to play key roles in deactivation of the cascade, and the resulting changes in the photoresponses of single rod cells will be determined by electrical recording. Using this approach, we will address 3 important questions: (1) How rapidly does rhodopsin become phosphorylated, and what determines this time course? (2) What are the functional consequences of arrestin translocation on the photoresponse? (3) Are the photoreceptor-specific splice variants of the RGS9 complex uniquely suited for deactivating transducin/PDE, and how? and 4) What are the mechanisms that speed transducin/PDE deactivation during light adaptation? This research addresses 1 of the objectives recommended by the Retinal Diseases Panel (http://www.nei.nih.gov/strategicplanning/np_retinal.asp#obj), which is to "Analyze the mechanisms underlying light adaptation and recovery following phototransduction and understand the changes in neural coding in light/dark adaptation." This research will help clarify the initial steps in the normal visual process, as well as the pathogenesis of diseases that arise from failures of deactivation, such as in some forms of retinitis pigmentosa and nightblindness. In a broader context, these experiments will provide insights into the mechanisms of deactivation of G protein cascades, which all eucaryotic cells use to transduce extracellular signals into intracellular responses.