Plasmin-dependent elimination of the growth-factor-like domain in urokinase causes its rapid cellular uptake and degradation.
Urokinase-type plasminogen activator (uPA) and its receptor (uPAR) act in concert to mediate pericellular proteolysis and to stimulate intracellular signalling responsible for cell migration and proliferation. uPA is composed of three domains, a proteolytic domain (PD), a kringle domain (KD) and a growth-factor-like domain (GFD), the last of which mediates the interaction with uPAR. We demonstrate that uPA, associated with the surface of U937 cells, undergoes plasmin-mediated cleavage of the Lys(46)-Ser(47) bond with elimination of the GFD. Using recombinant forms of uPA, we show that a uPA variant lacking the GFD (r-uPADeltaGFD) and unable to associate with uPAR is rapidly cleared from the cell surface. Binding and internalization of r-uPADeltaGFD are markedly decreased in the presence of 39 kDa receptor-associated protein (RAP), the antagonist of several endocytic receptors of the low-density lipoprotein receptor family, suggesting that this protein clearance pathway is used for r-uPADeltaGFD. In contrast with rapidly internalized r-uPADeltaGFD, the intact recombinant single-chain urokinase with wild-type structure (r-uPAwt) bound to uPAR is retained on the cell surface. Soluble uPAR protects uPA from cleavage by plasmin that results in the elimination of GFD, suggesting that uPAR might protect cell-bound urokinase from plasmin-mediated cleavage between the GFD and KD and subsequent degradation.
Poliakov
A
A
Biochemistry Department, Institute of Experimental Cardiology, Cardiology Research Center, Cherepkovskaya Street 15, Moscow 121552, Russia.
Tkachuk
V
V
Ovchinnikova
T
T
Potapenko
N
N
Bagryantsev
S
S
Stepanova
V
V
D001692:Biological Transport / Q000502:physiology; D004705:Endocytosis; D005341:Fibrinolysin / Q000378:metabolism*; D006133:Growth Substances / Q000737:chemistry; D006801:Humans; D018082:Kringles; D010447:Peptide Hydrolases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011956:Receptors, Cell Surface / Q000378:metabolism; D055293:Receptors, Urokinase Plasminogen Activator; D020298:U937 Cells; D014568:Urokinase-Type Plasminogen Activator / Q000737:chemistry / Q000378:metabolism*
D006133:Growth Substances; C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator; D010447:Peptide Hydrolases; D005341:Fibrinolysin; D014568:Urokinase-Type Plasminogen Activator