RPE Lactate Transporters: A Role in Retinal Survival

描述（由申请人提供）：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的运输来维持视网膜稳态。该提案的长期目标是表征RPE乳酸转运蛋白在维持正常视网膜代谢中的作用，并确定这些转运蛋白表达的变化如何有助于视网膜疾病的病理生理学。乳酸盐进出细胞的转运由质子偶联单羧酸转运蛋白（MCT）家族介导。在过去的资助期间，我们确定RPE表达两种MCT;顶膜中的MCT 1和基底膜中的MCT 3。RPE中MCT 1和MCT 3的极化分布提供了乳酸从视网膜向脉络膜的矢量转运机制，从而调节感光细胞周围空间的pH和渗透压。这些转运蛋白的表达或极化的改变预期会导致感光细胞变性。虽然MCT 1广泛表达，但MCT 3优先在RPE中表达。我们克隆了小鼠和人类MCT 3基因，并确定人类基因定位于染色体22q12.3-13.2，这是一个与黄斑变性相关的位点。最近，我们发现RPE中的MCT不是分离的蛋白质，而是与跨膜糖蛋白-CD 147相关。与MCT一样，CD 147在RPE的顶膜和基底外侧膜中丰富。在CD 147敲除小鼠中，RPE中的MCT表达缺失，表明亚基组装是MCT 1和MCT 3靶向质膜所必需的。在这个竞争性的更新应用中，我们提出了一系列假设驱动的实验，旨在进一步了解MCT 3在维持正常视觉功能中的重要性。本项目的具体目标是：（1）确定调节RPE中MCT 1和MCT 3的生物合成、运输和功能活性的机制;（2）表征MCT 3启动子并鉴定调节MCT 3基因发育和组织特异性表达的元件;（3）确定靶向缺失MCT 3基因对感光细胞的存活力和功能活性的影响。

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.