Mitogenic effects of urokinase on melanoma cells are independent of high affinity binding to the urokinase receptor

Mitogenic effects of urokinase on melanoma cells are independent of high affinity binding to the urokinase receptor

Gaubius Instituut TNO
Koopman, J.L.
Slomp, J.
de Bart, A.C.W.
Quax, P.H.A.
Verheijent, J.H.

1998

The structural and functional properties of the urokinase-type plasminogen activator (u-PA) that are involved in the mitogenic effect of this proteolytic enzyme on human melanoma cells M14 and IF6 and the role of the u-PA receptor (u-PAR) in transducing this signal were analyzed. Native u- PA purified from urine induced a mitogenic response in quiescent IF6 and M14 cells that ranged from 25 to 40% of the mitogenic response obtained by fetal calf serum. The half-maximum response in M14 and IF6 cells was reached at u- PA concentrations of approximately 35 and 60 nM, respectively. Blocking the proteolytic activity of u-PA resulted in a 30% decrease of the mitogenic effect, whereas inhibition of plasmin activity did not alter the mitogenic effect. No mitogenic response was elicited by low molecular weight u-PA, lacking the growth factor domain and the kringle domain. The ATF domain of u- PA induced a mitogenic response that was similar to complete u-PA. Defucosylated ATF and recombinant u-PA purified from Escherichia coli lacking all post-translational modifications did not induce a mitogenic response. Blocking the interaction of u-PA with u-PAR, using a specific monoclonal antibody, did not alter the mitogenic effect induced by u-PA. The binding of radiolabeled u-PA to M14 and IF6 cells was characterized by high affinity binding mediated by u-PAR and low affinity binding to an unknown binding site. These results demonstrate that proteolytically inactive u-PA is able to induce a mitogenic response in quiescent melanoma cells in vitro by a mechanism that involves the ATF domain but is independent of high affinity binding to u-PAR. Furthermore, it suggests that u-PA is able to bind with low affinity to a hitherto unidentified membrane associated protein that could be involved in u-PA-induced signal transduction.

Cell Division
Humans
Melanoma
Mitogens
Plasminogen Activator Inhibitor 1
Protein Binding
Receptors, Cell Surface
Signal Transduction
Tissue Plasminogen Activator
Tumor Cells, Cultured
Urinary Plasminogen Activator
Escherichia coli
Rickettsia sp. PAR

http://resolver.tudelft.nl/uuid:ee227f0d-479d-4dae-bbc3-7cdd9665bc7f

https://doi.org/10.1074/jbc.273.50.33267

234852

0021-9258

Journal of Biological Chemistry, 273 (273), 33267-33272

article