RPE Lactate Transporters: A Role in Retinal Survival

描述(申请人提供)：视网膜色素上皮(RPE)通过调节离子、代谢物和液体进出神经视网膜来维持视网膜的动态平衡。这项建议的长期目标是表征RPE乳酸转运体在维持正常视网膜代谢中的作用，并确定这些转运体的表达变化如何有助于视网膜疾病的病理生理学。乳酸进出细胞的转运是由一系列质子偶联的单羧酸转运体(MCTs)介导的。在过去的资助期间，我们发现RPE表达两种MCTs：MCT1在顶膜，MCT3在基底膜。MCT1和MCT3在RPE中的极化分布为乳酸从视网膜到脉络膜的矢量运输提供了一种机制，从而调节光感受器细胞周围空间的pH和渗透压。这些转运蛋白的表达或极化的改变有望导致光感受器细胞退化。MCT1在RPE中广泛表达，而MCT3在RPE中优先表达。我们克隆了小鼠和人类的MCT3基因，并确定人类的基因定位在染色体22q12.3-13.2上，该基因与黄斑变性有关。最近，我们发现RPE中的MCT不是分离的蛋白，而是与一种跨膜糖蛋白-CD147相关。与MCTs一样，CD147在RPE的顶膜和基底外侧膜中含量丰富。在CD147基因缺失的小鼠中，RPE中MCTs的表达缺失，这表明MCT1和MCT3需要亚单位组装才能靶向质膜。在这一竞争性更新应用中，我们提出了一系列综合的假设驱动实验，旨在加深我们对MCT3在维持正常视觉功能方面的重要性的理解。本项目的具体目标是：(1)确定调控RPE中MCT1和MCT3的生物合成、运输和功能活性的机制；(2)鉴定MCT3启动子的特征，并确定调控MCT3基因发育和组织特异性表达的元件；(3)确定靶向缺失MCT3基因对光感受器细胞活性和功能活性的影响。

DESCRIPTION (provided by applicant): The retinal pigment epithelium (RPE) maintains retinal homeostasis by regulating the transport of ions, metabolites and fluid into and out of the neural retina. The long-term goal of this proposal is to characterize the role of RPE lactate transporters in maintaining normal retinal metabolism and to determine how changes in expression of these transporters could contribute to the pathophysiology of retinal disease. Transport of lactate into and out of cells is mediated by a family of proton-coupled monocarboxylate transporters (MCTs). During the past funding period, we established that the RPE expresses two MCTs; MCT1 in the apical membrane and MCT3 in the basolateral membrane. The polarized distribution of MCT1 and MCT3 in the RPE provides a mechanism for vectorial transport of lactate from the retina to the choroid, thereby regulating the pH and osmolarity in the space surrounding the photoreceptor cells. Alteration in the expression or polarization of these transporters would be expected to lead to photoreceptor cell degeneration. While MCT1 is widely expressed, MCT3 is preferentially expressed in the RPE. We cloned the mouse and human MCT3 genes and determined that the human gene maps to chromosome 22q12.3-13.2, a locus that has been associated with macular degeneration. Recently we have found that MCTs in the RPE are not isolated proteins but rather associate with a transmembrane glycoprotein- CD147. Like MCTs, CD147 is abundant in the apical and basolateral membranes of the RPE. In the CD147 null mice there is a loss of expression of MCTs in the RPE, suggesting that subunit assembly is required for targeting of MCT1 and MCT3 to the plasma membrane. In this competitive renewal application, we have proposed an integrated series of hypothesis driven experiments designed to further our understanding of the importance of MCT3 in maintaining normal visual function. The Specific Aims of this project are: (1) to determine the mechanisms that regulate the biosynthesis, trafficking, and functional activity of MCT1 and MCT3 in the RPE; (2) to characterize the MCT3 promoter and identify elements that regulate developmental and tissue specific expression of the MCT3 gene; (3) to determine the effects of targeted deletion of the MCT3 gene on the viability and functional activity of photoreceptor cells.