Endocytosis and lysosomal delivery of tissue plasminogen activator- inhibitor 1 complexes in Hep G2 cells

D. M. Underhill, D. A. Owensby, P. A. Morton, A. L. Schwartz

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

Receptor-mediated endocytosis of tissue-type plasminogen activator (t-PA)- plasminogen activator inhibitor type 1 (PAI-1) complexes results in their clearance by Hep G2 cells. After complexes are internalized, the t-PA component is degraded. However, neither the locus of intracellular catabolism nor the fate of PAI-1 has been elucidated. To characterize these aspects of t-PA·PAI-1 catabolism, the subcellular distribution of a prebound cohort of ligand molecules was delineated after internalization at 37°C. 125I-t- PA·PAI-1 and t-PA·125I-PAI-1 were compared in separate experiments. After ligand uptake, intracellular vesicles were separated on density gradients. Internalized 125I-t-PA·PAI-1 concentrated initially in endosomes. After 20 minutes of uptake, the complex began to appear in lysosomes. Subsequently, low molecular weight labeled ligand fragments were detected in culture media. A panel of lysosomotropic agents, including primaquine, chloroquine, ammonium chloride, and a combination of leupeptin and pepstatin A, inhibited degradation. When t-PA·125I-PAI-1 rather than 125I-t-PA·PAI-1 was internalized, strikingly different results were observed. Although the kinetics of internalization and the intracellular itinerary were indistinguishable for the differently labeled complexes, the 125I-PAI-1 component of t-PA·125I-PAI-1 resisted rapid degradation. After a rapid loss of t-PA, the 125I-PAI-1 moiety persisted in lysosomes for up to 180 minutes. Thus, internalized t-PA·PAI-1 is targeted to lysosomes in which PAI-1 is relatively more stable than t-PA.

Original languageEnglish
Pages (from-to)2746-2754
Number of pages9
JournalBlood
Volume80
Issue number11
StatePublished - Jan 1 1992
Externally publishedYes

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