Regulation of single chain urokinase binding, internalization, and degradation by a plasminogen activator inhibitor 1-derived peptide.
The internalization and degradation of cell-associated urokinase type plasminogen activator (uPA) through the alpha2-macroglobulin receptor/low density lipoprotein-related receptor (alpha2MR/LRP) represent important steps in the control of plasmin formation. Complexes between two chain urokinase (tcuPA) and plasminogen activator type 1 are degraded rapidly whereas single chain urokinase (scuPA) is not, suggesting that alpha2MR/LRP requires specific epitopes in the serpin for effective function. We report an alternative mechanism that may contribute to this process. The binding of scuPA to LM-TK- cells that lack the uPA receptor was stimulated by the hexapeptide EEIIMD, corresponding to amino acids 350-355 of plasminogen activator type 1, which contacts the sequence RHRGGS, corresponding to amino acids 179-184 in uPA. EEIIMD increased the Bmax of scuPA binding 4-fold with the half-maximal effect achieved at a peptide concentration of 50 microM. Stimulation was dependent on the charge on the COOH-terminal amino acid but not on the NH2 terminus of the peptide. EEIIMD also stimulated the internalization and degradation of scuPA. Both the binding and internalization of scuPA in the presence of EEIIMD were blocked by recombinant, 39-kDa alpha2MR/LRP-associated protein as well as by an anti-alpha2MR/LRP antibody. EEIIMD also stimulated the binding of scuPA to purified alpha2MR/LRP. EEIIMD had no effect on the binding of tcuPA or of complexes between scuPA and its receptor. These results suggest that EEIIMD regulates the binding of scuPA with alpha2MR/LRP. These findings also suggest a potential mechanism by which scuPA can be cleared which is independent of activation by plasmin or binding to uPA receptor.
Zhang
L
L
Departments of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Strickland
D K
DK
Cines
D B
DB
Higazi
A A
AA
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D004705:Endocytosis; D000939:Epitopes / Q000032:analysis; D005341:Fibrinolysin / Q000378:metabolism; D007700:Kinetics; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D046911:Macromolecular Substances; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000378:metabolism / Q000494:pharmacology*; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry* / Q000494:pharmacology*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D011971:Receptors, Immunologic / Q000378:metabolism*; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry* / Q000378:metabolism*
D000939:Epitopes; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D046911:Macromolecular Substances; D009842:Oligopeptides; D010446:Peptide Fragments; D017395:Plasminogen Activator Inhibitor 1; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D055293:Receptors, Urokinase Plasminogen Activator; D011994:Recombinant Proteins; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator