Differences in renal metabolism of insulin and cytochrome c

Kidneys degrade small proteins such as cytochrome c (CYT c) by the classic lysosomal pathway. However, because alternate routes for the transport and degradation of protein hormones have been identified in other tissues, we set out to determine whether extralysosomal sites might participate in the renal degradation of insulin. First, we compared the effect of the lysosomal inhibitor NH4Cl on insulin and CYT c degradation by isolated perfused rat kidneys. After kidneys were loaded with radiolabeled proteins to allow for absorption and transport to lysosomes, degradation was measured in the presence or absence of inhibitors. Control kidneys degraded 45 +/- 1.5% of the trapped CYT c per hour, and this was inhibited 62 +/- 1.3% by NH4Cl. In contrast, 86 +/- 2.4% of the trapped insulin was degraded per hour, and this was inhibited 26 +/- 4% by NH4Cl. Next we followed the subcellular distribution of 125I-labeled insulin in kidneys exposed to 125I-labeled insulin in vivo or when isolated and perfused. Under both circumstances the distribution of insulin on a linear sucrose gradient differed from that of the lysosomal enzyme N-acetyl-beta-glucosaminidase. In contrast, [14CH3]CYT c, injected in vivo, distributed over a density similar to the lysosomal marker. Thus important differences exist between the renal metabolism of CYT c, which proceeds in lysosomes, and the renal metabolism of insulin. These include rate of degradation, sensitivity to NH4Cl, and subcellular sites of localization. Accordingly, we suggest that insulin degradation may occur, at least in part, in a different compartment from the classic lysosomal site of protein degradation.