Role of System LAmino Acid Transporter in Podocyte Cell Biology

我们研究了肾小球中的两个L氨基酸转运蛋白系统LAT2和LAT3。LAT2及其功能配体4F2hc不仅表达于近端小管，也表达于足细胞，但定位于足细胞胞浆，提示LAT2在正常成熟足细胞中不具有氨基酸转运蛋白的功能。然而，有趣的是，LAT2在人IgA肾病和ANCA相关性肾小球肾炎的细胞新月体病变中大量表达。我们利用大鼠新月体肾炎模型进一步研究了LAT2在新月体形成过程中的作用。MTOR途径下游的S6核糖体蛋白和4E-BP1的磷酸化明显增强，先于细胞新月体中LAT2和4F2hc的表达。LAT2基因转染的细胞株中p70S6K和4E-BP1的磷酸化明显增强，但在LAT功能抑制剂的存在下，这种磷酸化作用明显被抑制。综上所述，我们将…进一步认为，LAT2和4F2hc的上调可能通过生长因子和细胞因子激活的mTOR信号促进肾小球壁细胞质膜上LAT2和4F2hc的表达，从而通过增加细胞内支链氨基酸而导致mTOR信号的二次诱导而参与新月体的形成。另一方面，LAT3在人和小鼠肾脏足细胞足突的顶膜上有明显的表达。培养的小鼠足细胞呈L系统氨基酸转运系统。食物饥饿使小鼠LAT3蛋白和mRNA表达上调。人的致命肾脏发现LAT3开始在S形体肾小球表达。最后，缺失LAT3的斑马鱼形态异型体显示肾小球塌陷，肾小球基底膜增厚，尽管裂隙隔保存完好。染料注射实验表明斑马鱼LAT3变异体的超滤屏障被破坏。我们的数据表明，LAT3在肾小球发育中起着关键作用，其机制是LAT3将亮氨酸从鲍曼间隙运输到足细胞胞质，从而通过哺乳动物雷帕霉素-2激活的靶点来调节蛋白质合成系统。较少

We investigated two system L amino acid transporters, LAT2 and LAT3 in the glomerulus. LAT2 and its functional ligand, 4F2hc, was expressed not only in the proximal tubules but also in the podocytes, however, its localization was revealed in the podocyte cytoplasm, suggesting that LAT2 does not function as amino acid transporter in normal mature podocytes. However, interestingly, LAT2 was dearly expressed in the cellular crescent lesion of human IgA nephropathy and ANCA-related glomerulonephritis. We further studied the role of LAT2 in the process of crescent formation by using rat crescent glomerulonephritis model. Increased phosphorylation of S6 ribosomal protein and 4E-BP1, down stream of mTOR pathway, was clearly demonstrated, preceding increase of LAT2 and 4F2hc expression in cellular crescents. LAT2 cDNA-transfected cell line exhibited predominant phosphorylation of p70S6K and 4E-BP1, which was clearly abrogated in the presence of LAT functional inhibitor. Taken together, we conc … More luded that upregulation of LAT2 and 4F2hc could be involved in the crescent formation via the mechanism by which mTOR signaling activated by growth factors and cytokines accelerated LAT2 and 4F2hc expression in the plasma membrane of glomerular parietal cells that leads secondary induction of mTOR signaling through increase of intra cellular branched-chain amino acids. On the other hand, LAT3 was clearly expressed in apical membrane of podocyte foot processes in human and mouse kidney. Cultured mouse podocytes exhibited system L amino acid transport system. Mouse LAT3 protein and mRNA were upregulated by food starvation. The human fatal kidney revealed LAT3 commencing expressing in the S-shaped body glomeruli Finally, zebrafish morphant lacking lat3 revealed collapsed glomerulus with thickened glomerular basement membrane, despite well-preserved slit diaphragm. Dye-injection experiment indicated the disruption of ultrafiltration barrier in zebrafish lat3 morphant Our data suggest that LAT3 plays a crucial role in glomerular development via the mechanism by which LAT3 transports leucine from the Bowman's space to podocyte cytoplasm, leading to the regulation of the protein synthesis system through mammalian target of rapamycin-2 activation. Less