EMBRYONIC FIBROBLASTS THAT ARE GENETICALLY DEFICIENT IN LOW-DENSITY-LIPOPROTEIN RECEPTOR-RELATED PROTEIN DEMONSTRATE INCREASED ACTIVITY OF THE UROKINASE RECEPTOR SYSTEM AND ACCELERATED MIGRATION ON VITRONECTIN

Low density lipoprotein receptor-related protein (LRP) mediates the en docytosis of diverse ligands, including urokinase plasminogen activato r (uPA) and its receptor, uPAR, which have been implicated in cellular migration. The purpose of this study was to determine whether LRP aff ects cellular migration, Murine embryonic fibroblasts (MEF) that are L RP-deficient due to targeted gene disruption and exotoxin selection (M EF-2), heterozygous fibroblasts (PEA-10), and wild-type fibroblasts (M EF-1) were compared. When cultures were denuded of cells in a 1-mm-wid e strip, all three cell types migrated into the denuded area. The MEF- 2 cells migrated nearly twice as rapidly as the MEF-1 cells or PEA-10 cells, The difference in migration velocity was duplicated in culture wells that were precoated with serum or vitronectin and partially dupl icated in wells coated with fibronectin but not in wells coated with t ype I collagen or Matrigel. uPA was detected in MEF-2 conditioned medi um (CM) at a concentration of 0.30 +/- 0.02 nm, which was 13-fold high er than the level detected in MEF-1 CM or PEA-10 CM, suggesting one po tential mechanism for the enhanced migration of MEF-2 cells. uPAR was also increased on MEF-2 cells by 4-5-fold, as determined by PI-PLC rel ease, and by 2.5-fold, as determined by a uPA/uPAR activity assay, Man nosamine treatment, which down-regulates cell-surface uPAR, decreased MEF-2 migration by 40% without significantly affecting MEF-1 migration , MEF-2 CRI, which is uPA-rich, increased the rate of MEF-1 migration, and MEF-1 CM did not, These studies demonstrate alterations in cellul ar migration and in the activity of the uPA/uPAR system which accompan y complete deficiency of LRP expression in fibroblasts. We propose tha t uPA and uPAR form an autocrine loop for promoting fibroblast migrati on and that LRP counteracts the activity of this system.