Inhibition of Autophagy and Multiple Steps in Asialoglycoprotein Endocytosis by Inhibitors of Tyrosine Protein Kinases (Tyrphostins)

J Biol Chem. 1995 May 26;270(21):12823-31. doi: 10.1074/jbc.270.21.12823.

Abstract

In isolated rat hepatocytes, several tyrosine protein kinase inhibitors (tyrphostins) reduced the autophagic sequestration of electroinjected [3H]raffinose by 40-75% at doses that did not significantly affect cellular ATP levels or plasma membrane integrity. Tyrphostin 46 specifically inhibited autophagy, whereas tyrphostins 1, 25 and 51 also suppressed the receptor-mediated endocytic uptake of 125I-tyramine-cellobiose-asialoorosomucoid, 125I-TC-AOM, by 20-30% and its degradation by 70-90%. Tyrphostins 1 and 51, and the microtubule inhibitor vinblastine, inhibited an early endocytic step (endosome maturation/multivesiculation?), causing accumulation of endocytosed 125I-TC-AOM in a recycling compartment that corresponded to light endosomes (1.10-1.11 g/ml) in sucrose density gradients. In the electron microscope, these endosomes could be recognized as small, peripheral endocytic vesicles and tubules accumulating endocytosed AOM-gold. The serine/threonine protein phosphatase inhibitor okadaic acid inhibited an intermediate endocytic step (detachment of multivesicular endosomes from the tubulovesicular network?), causing accumulation of 125I-TC-AOM in a recycling compartment corresponding to light endosomes (1.10-1.11 g/ml), but with a multivesicular rather than a tubulovesicular morphology. Tyrphostin 25 inhibited endocytosis at a late step (endosome-lysosome fusion?), causing accumulation of 125I-TC-AOM in a non-recycling compartment corresponding to dense, multivesicular endosomes (1.14 g/ml) that had probably detached from the light endosomal network.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / analysis
  • Animals
  • Asialoglycoproteins / metabolism*
  • Autophagy / physiology*
  • Cell Survival / drug effects
  • Dose-Response Relationship, Drug
  • Endocytosis / physiology*
  • Endosomes / physiology
  • Endosomes / ultrastructure
  • Ethers, Cyclic / pharmacology
  • Liver / cytology
  • Liver / physiology*
  • Liver / ultrastructure
  • Lysosomes / physiology
  • Lysosomes / ultrastructure
  • Male
  • Okadaic Acid
  • Organelles / physiology*
  • Organelles / ultrastructure
  • Protein Biosynthesis / drug effects
  • Protein-Tyrosine Kinases / antagonists & inhibitors*
  • Rats
  • Rats, Wistar
  • Vacuoles / physiology
  • Vacuoles / ultrastructure
  • Vanadates / pharmacology
  • Vinblastine / pharmacology

Substances

  • Asialoglycoproteins
  • Ethers, Cyclic
  • Okadaic Acid
  • Vanadates
  • Vinblastine
  • Adenosine Triphosphate
  • Protein-Tyrosine Kinases