RETROGRADE PROTEIN TRANSPORT IN RENAL EPITHELIAL CELLS

蛋白质跨膜转运的机制 仍然是现代细胞生物学研究的中心焦点。 细胞 膜被认为是一种选择性的蛋白质屏障， 除此之外，外源蛋白要么被排除在细胞之外，要么进入细胞。 这是一种“默认途径”，迅速达到溶酶体降解的高潮。 外源蛋白质进入细胞内部是罕见的 到目前为止，似乎仅限于少数细菌毒素， 靶向序列设法逃脱溶酶体降解。 在 与任何先前对外源蛋白质易位的描述相反， 我们最近在大鼠的近端组织中发现了一种丰富的15.5 kDa的蛋白质， 是α 2 μ-微球蛋白的蛋白水解裂解产物的小管 (A2)主要由肝脏合成的19 kDa蛋白质。 a2是 蛋白质超家族成员（包括视黄醇结合蛋白） 被认为是作为疏水配体的转运蛋白， 长链脂肪酸。 这种肝蛋白在 肾脏的近端小管可能是第一个生理上 "逆行蛋白质转运"一个重要例子， 天然合成的外源蛋白质在细胞质中积累 另一种细胞类型。 我们认为肾小球滤过后， 近曲小管细胞通过一种新的途径代谢A2， 积累大量的蛋白水解切割产物（A2片段） 在细胞质中。 为了验证这一假设，我们建议：（1） 描述了A2的吸收和转运的过程，由近端 （2）确定这一过程是否由 内吞作用（受体介导、液相或吸附性内吞作用） （3）评估A2与疏水性的结合是否 配体是A2摄取所需的，或修饰A2摄取;（4）鉴定细胞的 负责处理A2的隔室， 发生A2的积累而不是降解，并且（5）确定 是否独特的结构特征决定了A2是如何被处理的 近端小管细胞 这些研究可以提供新的见解 一种以前未知的将蛋白质从外移到内的途径 牢房 这一信息可能与有针对性的交付有关 各种药剂或药物对肾脏的影响 此外，A2结合脂肪 在体外作为脂肪酸结合蛋白，在体内作为"脂肪酸结合样蛋白"。 作为一种转运蛋白，A2可以调节脂肪酸的氧化速率 在正常和病理生理状态如租赁缺血期间。

The mechanism(s) by which proteins are translocated across membranes remains a central focus of research in modern cell biology. The cell membrane is thought to act as a selective protein barrier and with few exceptions, exogenous proteins are either excluded from the cell or enter a 'default pathway' that rapidly culminates with lysosomal degradation. Entry of an exogenous protein into the cell's interior is a rare event and thus far, appears restricted to a few bacterial toxins with unique targeting sequences that manage to escape lysosomal degradation. In contrast to any previous description of exogenous protein translocation, we recently found an abundant, 15.5 kDa protein in the rat proximal tubule that is a proteolytic cleavage product of alpha2mu-microglobulin (A2), a 19 kDa protein synthesized predominantly by the liver. A2 is a member of a protein superfamily (including retinol binding protein) thought to function as transport proteins for hydrolyphobic ligands such as long chain fatty acids. Accumulation of this hepatic protein within the proximal tubule of the kidney may be the first physiologically significant example of 'retrograde protein transport' in which a naturally synthesized, exogenous protein is accumulated in the cytosol of another cell type. We suggest that following glomerular filtration, the proximal tubule cell metabolizes A2 by a novel pathway, ultimately accumulating large amount of a proteolytic cleavage product (A2-fragment) in the cytosol. To evaluate this hypothesis, we propose to: (1) describe the process of A2 uptake and translocation by the proximal epithelial cell; (2) determine whether this process is mediated by endocytosis (receptor-mediated, fluid-phase, or adsorptive endocytosis) or simple diffusion; (3) evaluate whether binding of A2 to a hydrophobic ligand is required for, or modifies A2 uptake; (4) identify the cellular compartment(s) responsible for processing A2 so that cytosolic accumulation, rather than degradation, of A2 occurs and (5) determine whether unique structural features determine how A2 is processed by the proximal tubule cell. These studies could provide new insights regarding a previously unknown pathway for moving proteins from outside to inside the cell. This information could have relevance for targeted delivery of various agents or drugs to the kidney. In addition, A2 binds fatty acids in vitro and act as a 'fatty acid binding like-protein' in vivo. As a transport protein, A2 could modulate fatty acid oxidation rates during both normal and pathophysiologic states such as rental ischemia.