RPE Lactate Transporters: A Role in Retinal Survival

描述（由申请人提供）：视网膜色素上皮（RPE）通过调节进出神经视网膜的离子、代谢物和液体的运输来维持视网膜稳态。本提案的长期目标是表征RPE乳酸转运体在维持正常视网膜代谢中的作用，并确定这些转运体表达的变化如何有助于视网膜疾病的病理生理。乳酸进出细胞的运输是由质子偶联的单羧酸转运体（mct）家族介导的。在过去的资助期间，我们确定了RPE表达两个mct；MCT1位于根尖膜，MCT3位于基底外膜。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，从而调节了光感受器细胞周围空间的pH和渗透压。这些转运蛋白表达或极化的改变可能导致光感受器细胞变性。虽然MCT1广泛表达，但MCT3优先在RPE中表达。我们克隆了小鼠和人类MCT3基因，并确定人类基因定位于染色体22q12.3-13.2，一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CD147相关。与mct一样，CD147在RPE的根尖和基底外膜中含量丰富。在CD147缺失的小鼠中，RPE中mct的表达缺失，这表明亚基组装是MCT1和MCT3靶向质膜所必需的。在这个竞争性更新应用中，我们提出了一系列完整的假设驱动实验，旨在进一步了解MCT3在维持正常视觉功能中的重要性。该项目的具体目标是：(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)表征MCT3启动子，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.