Internalization of insulin into rat liver Golgi elements. Evidence for vesicle heterogeneity and the path of intracellular processing.

Abstract

In this study, we used Percoll gradient centrifugation to subfractionate Golgi fractions (Golgi light, G1; intermediate, Gi; heavy, Gh).Marker enzyme and insulin receptor distribution as well as the pattern of '251-insulin internalization were examined.In Gi and Gh subfractions, the relative specific activity (RSA) for galactosyltransferase was maximum (RSA = 70) in the low density region of the gradient corresponding to the major protein peak; whereas the maximum for N-acetyl-P-D-glucosaminidase (RSA = 4.1) and acid phosphatase (RSA = 4.0) was in the high density region near the minor protein peak.Two peaks of insulin binding were seen in low and high density regions, respectively.In G1 subfractions, no peak of lysosomal enzyme activity was seen and only one peak of insulin binding in the low density galactosyltransferase-enriched region was observed.'2SI-Insulin was rapidly (1 min) internalized exclusively into the galactosyltransferase region but by 10 min was predominantly in the acid phosphatase region in both Gi and Gh.In G1, no such shift in radiolabel distribution was seen.Radiolabel reflected intact '251-insulin and radioautography demonstrated grains in vesicles containing lipoprotein-like particles in all gradient regions.These studies have demonstrated vesicle heterogeneity within Gi and Gh and two intracellular pools of insulin receptors.One pool was associated with Golgilike vesicles of low density and a second with unique vesicles, having lysosome-like features, of higher density.The sequence of '251-insulin internalization (first into low and then into higher density vesicles) provides evidence for the intracellular pathway of insulin processing, and is compatible with a role for insulin in Golgi function.