Desensitization of Cone Visual Signaling Pathways

描述（由申请人提供）：脊椎动物视杆细胞和视锥细胞的光传导由一系列精确计时的事件组成，这些事件是光感受器在广泛的光强度下发挥作用所必需的。杆状细胞在昏暗的光线下工作，在明亮的光线下很容易饱和，而锥状细胞恢复得更快，能够适应更强的光线。第二信使，如cGMP和Ca2+，已经被认为在视杆细胞和视锥细胞的光传导级联和适应中发挥重要作用。另一种第二信使cAMP在光感受器中以光依赖和昼夜节律依赖的方式进行调节，可能对适应也很重要。例如，cAMP合成的变化与杆状细胞和视锥细胞的适应缺陷有关，而杆状细胞和视锥细胞的适应缺陷会改变内视网膜的信号。有趣的是，cAMP的下游靶点PKA对光感受器细胞蛋白的磷酸化作用尚未得到很好的研究。我们已经确定视网膜特异性G蛋白偶联受体激酶GRK1和GRK7都是体外PKA的底物，它们在视杆细胞和视锥细胞的恢复和适应中起着关键作用。我们还表明，PKA的磷酸化降低了这些激酶磷酸化其底物（体外视蛋白）的能力。在这项提议中，我们提供了新的证据，证明这两种激酶在体内都被PKA磷酸化，并且磷酸化受光调节。我们建议使用小鼠作为模型来研究PKA磷酸化GRK1对其在恢复和适应中的作用。我们的数据介绍了光感受器细胞特异性GRKs调控的新机制，它可能影响光传导的多个方面。

DESCRIPTION (provided by applicant): Phototransduction in vertebrate rods and cones consists of a series of precisely timed events that are necessary for photoreceptors to function under a broad range of light intensities. While rods operate under dim light and are easily saturated in response to bright light, cones recover more rapidly and are able to adapt to much more intense light. Second messengers, such as cGMP and Ca2+, are already well-known to play important roles in the phototransduction cascade and adaptation in rods and cones. Another second messenger, cAMP, which is regulated both in a light- and a circadian-dependent manner in photoreceptors, may also be important for adaptation. For example, changes in cAMP synthesis are associated with defects in adaptation in rods and cones that alter signaling to the inner retina. Interestingly, phosphorylation of photoreceptor cell proteins by PKA, the downstream target of cAMP, has not been well-studied. We have determined that the retina-specific G protein-coupled receptor kinases, GRK1 and GRK7, which play critical roles in recovery and adaptation in rods and cones, are both substrates for PKA in vitro. We have also shown that phosphorylation by PKA reduces the ability of these kinases to phosphorylate their substrates, the opsins in vitro. In this proposal, we provide new evidence that both kinases are phosphorylated by PKA in vivo and that phosphorylation is regulated by light. We propose to use the mouse as a model to study the functional consequences of GRK1 phosphorylation by PKA on its role in recovery and adaptation. Our data introduce a novel mechanism for the regulation of photoreceptor cell- specific GRKs that may influence multiple facets of phototransduction. PUBLIC HEALTH RELEVANCE: Rods and cones are the cells in the vertebrate retina that mediate our visual response to light. Rods are responsible for dim light vision, whereas cones function in bright light and are responsible for color vision. Significant differences between rods and cones have been observed in the sensitivity and kinetics of the light responses as well as their susceptibility to genetically and environmentally induced disease processes. This proposal is designed to analyze the contributions of GRK1, a kinase that is important in visual signaling in both rods and cones and that has been implicated in disease such as stationary night blindness. These studies are expected to lead to a better understanding of the mechanisms in the retina used to adapt to changing light conditions.