Proteasome function in the aging retina

描述(申请人提供)：建议的工作重点是蛋白酶体。这种酶复合体负责大多数细胞蛋白质的降解，包括负责细胞活力和功能的基本过程。我们将询问与年龄相关的蛋白酶体功能缺陷是否会破坏视网膜色素上皮(RPE)的功能，以及蛋白酶体功能降低的后果是什么。一个重点将是定义蛋白酶体在RPE中的角色，其中考虑两个角色。由于蛋白酶体控制着对细胞生存至关重要的过程，蛋白酶体功能的丧失可能通过功能障碍的蛋白质积累或关键细胞通路的中断而导致RPE细胞死亡。或者，如果蛋白酶体参与吞噬的杆状外节(ROS)的降解，蛋白酶体功能的降低可能导致ROS蛋白加工的减少。第二个重点将是阐明导致蛋白酶体功能随年龄变化而丧失的生化机制。将追求以下目标：(1)确定RPE中蛋白酶体的蛋白分解能力。(A)确定蛋白酶体在被吞噬的杆状外节降解中的作用。(B)确定应激源对RPE蛋白酶体功能的影响。(2)测定视网膜蛋白酶体功能的增龄性变化。(A)量化与年龄有关的蛋白酶体功能和表达的变化。(B)确定蛋白酶体亚基的翻译后修饰。我们的研究方法将包括生物化学、细胞生物学、免疫学、分子生物学和蛋白质组学方面的一系列技术，使用啮齿动物衰老模型、转基因小鼠和培养细胞。目前对蛋白酶体在视网膜中的功能知之甚少，因此这些研究将为视网膜的基本生理学和衰老的影响提供重要的贡献。

DESCRIPTION (provided by applicant): The proposed work focuses on the proteasome. This enzyme complex is responsible for most cell protein degradation, including fundamental processes responsible for cell viability and function. We will ask if age-related defects in proteasome function disrupt the function of the retinal pigment epithelium (RPE), and what are the consequences of reduced proteasome function. One focus will be on defining the role of the proteasome in the RPE, where two roles are considered. Since proteasome controls processes that are essential for cell survival, a loss of proteasome function could cause RPE cell death through the accumulation of dysfunctional proteins or the disruption of critical cell pathways. Alternatively, if proteasome participates in the degradation of phagocytosed rod outer segments (ROS), decreased proteasome function could contribute to diminished processing of ROS proteins. A second focus will be to elucidate the biochemical mechanism responsible for the age-dependent loss in proteasome function. The following Aims will be pursued: (1) Define the proteolytic capabilities of the proteasome in the RPE. (a) Define the role of the proteasome in the degradation of phagocytosed rod outer segments. (b) Determine the effect of stressors on RPE proteasome function. (2) Determine age-related changes in retinal proteasome function. (a) Quantify age-related alterations in proteasome function and expression. (b) Identify post-translational modifications in proteasomal subunits. Our investigative approach will include a range of techniques in biochemistry, cell biology, immunology, molecular biology, and proteomics using rodent models of aging, transgenic mice, and cultured cells. Little information is known about proteasome function in the retina, so these studies will provide an important contribution to basic retinal physiology and the effects of aging.