Determinants of Rod and Cone Response Characteristics

描述（由申请人提供）：视色素磷酸化对正常视力至关重要。缺乏这种生化反应可导致夜盲症和感光细胞凋亡。磷酸化使视紫红质和大多数g蛋白偶联受体从下游信号通路中分离。大多数g蛋白偶联受体的磷酸化位点尚不清楚。在之前的资助期间，我们开发了一种基于质谱的方法，使我们能够确定完整视网膜中视觉色素上发生的磷酸化和去磷酸化反应的位点和动力学。我们利用该方法建立了多种生理条件下小鼠和斑马鱼视觉色素磷酸化的位点和动力学。

DESCRIPTION (provided by applicant): Phosphorylation of visual pigments is essential for normal vision. Deficiencies in this biochemical reaction can cause night blindness and photoreceptor apoptosis. Phosphorylation uncouples rhodopsin and most G-protein coupled receptors from downstream signaling pathways. The sites of phosphorylation on most G-protein coupled receptors are not known. During the previous funding period we developed a mass- spectrometry based method that enables us to determine the sites and kinetics of phosphorylation and dephosphorylation reactions that occur on visual pigments in intact retinas. We have used the method to establish the sites and kinetics of phosphorylation on mouse and zebrafish visual pigments under a variety of physiological conditions. We propose to use the method to pursue four new specific aims. 1. Evaluate rhodopsin phosphorylation during long-term illumination and identify relationships between visual pigment phosphorylation, visual cycle and visual sensitivity. 2. Evaluate factors that influence phosphorylation of cone visual pigments. 3. Identify structural characteristics that determine site selection and kinetics of phosphorylation by rhodopsin kinase. 4. Evaluate the influence of recoverin and Calcium on visual pigment phosphorylation. These studies will provide the most precise data yet available about the kinetics and site preferences for phosphorylation of G-protein coupled receptors.