The urokinase/PAI-2 complex: a new high affinity ligand for the endocytosis receptor low density lipoprotein receptor-related protein.
The efficient inactivation of urokinase plasminogen activator (uPA) by plasminogen activator inhibitor type 2 (PAI-2) at the surface of carcinoma cells is followed by rapid endocytosis of the uPA-PAI-2 complex. We now show that one pathway of this receptor-mediated endocytosis is mediated via the low density lipoprotein receptor-related protein (LRP) in prostate cancer cells. Detailed biochemical analyses using ligand binding assays and surface plasmon resonance revealed a novel and distinct interaction mechanism between native, human LRP and uPA-PAI-2. As reported previously for PAI-1, inhibition of uPA by PAI-2 significantly increased the affinity of the complex for LRP (K(D) of 36 nm for uPA-PAI-2 versus 200 nm for uPA). This interaction was maintained in the presence of uPAR, confirming the validity of this interaction at the cell surface. However, unlike PAI-1, no interaction was observed between LRP and PAI-2 in either the stressed or the relaxed conformation. This suggests that the uPA-PAI-2-LRP interaction is mediated by site(s) within the uPA molecule alone. Thus, as inhibition of uPA by PAI-2 resulted in accelerated clearance of uPA from the cell surface possibly via its increased affinity for LRP, this represents a mechanism through which PAI-2 can clear proteolytic activity from the cell surface. Furthermore, lack of a direct interaction between PAI-2 and LRP implies that downstream signaling events initiated by PAI-1 may not be activated by PAI-2.
Croucher
David
D
School of Biological Sciences, University of Wollongong, New South Wales 2522.
Saunders
Darren N
DN
Ranson
Marie
M
D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005434:Flow Cytometry; D006801:Humans; D007700:Kinetics; D008024:Ligands; D026503:Low Density Lipoprotein Receptor-Related Protein-1 / Q000378:metabolism*; D008297:Male; D018613:Microscopy, Confocal; D017396:Plasminogen Activator Inhibitor 2 / Q000737:chemistry / Q000502:physiology*; D011471:Prostatic Neoplasms / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D020349:Surface Plasmon Resonance; D013997:Time Factors; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*
D002966:Clathrin; D004791:Enzyme Inhibitors; D008024:Ligands; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D017396:Plasminogen Activator Inhibitor 2; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D014568:Urokinase-Type Plasminogen Activator