The luminal brush border of the renal proximal tubule is highly specialized for protein reabsorption by endocytosis. This endosomal compartment is an important site of hormone and iron metabolism, as well as a portal of entry for nephrotoxins. New methods in cell biology, including freeflow electrophoresis and flow cytometry, allow us to isolate elements of the proximal tubular endosomal pathway with diverse structural and functional properties. These elements include clathrin coated vesicles, endosomal vesicles, endosomal vacuoles, and dense apical tubules. Our hypothesis is that compartments of the renal cortical endosomal pathway have different protein and lipid components, and these components determine their function in the pathway. Endosomes undergo efficient and highly specific fusion events with select compartments of intracellular vesicles. To delineate the molecular machinery which governs fusion events, we have reconstituted a cell free fusion assay in vitro. We will test whether small molecular weight G-- proteins mediate fusion specificity. Understanding the structure and function of the renal proximal tubular endosomal pathway may explain some forms of nephrotoxicity. Several lines of evidence suggest that disruption of the endosomal pathway is an important mediator of aminoglycoside nephrotoxicity. To determine whether the changes in the endosomal pathway associated with aminoglycoside treatment are mediators of nephrotoxicity or an epiphenomenon we will correlate endosomal changes with documented patterns of aminoglycoside injury. Hence we will determine the role of endosomal disruption in nephrotoxic injury.
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