Dj. Webb et al., The very low density lipoprotein receptor regulates urokinase receptor catabolism and breast cancer cell motility in vitro, J BIOL CHEM, 274(11), 1999, pp. 7412-7420
The very low density lipoprotein receptor (VLDLr) binds diverse ligands, in
cluding urokinase-type plasminogen activator (uPA) and uPA-plasminogen acti
vator inhibitor-1 (PAI-1) complex. In this study, we characterized the effe
cts of the VLDLr on the internalization, catabolism, and function of the uP
A receptor (uPAR) in MCF-7 and MDA-MB-435 breast cancer cells. When challen
ged with uPA . PAI-1 complex, MDA-MB-435 cells internalized uPAR; this proc
ess was inhibited by 80% when the activity of the VLDLr was neutralized wit
h receptor-associated protein (RAP), To determine whether internalized uPAR
is degraded, we studied the catabolism of [S-35]methionine-labeled uPAR. I
n the absence of exogenous agents, the uPAR catabolism t(1/2) was 8.2 h. uP
A . PAI-1 complex accelerated uPAR catabolism (t(1/2) to 1.8 h), while RAP
inhibited uPAR catabolism in the presence (t(1/2) of 7.8 h) and absence (t(
1/2) of 16.9 h) of uPA . PAI-1 complex, demonstrating a critical role for t
he VLDLr, When MCF-7 cells were cultured in RAP, cell surface uPAR levels i
ncreased gradually, reaching a new steady-state in 3 days, The amount of uP
A which accumulated in the medium also increased. Culturing in RAP for 3 da
ys increased MCF-7 cell motility by 2.2 +/- 0.1-fold and by 4.4 +/- 0.3-fol
d when 1.0 nm uPA was added. The effects of RAP on MCF-7 cell motility were
entirely abrogated by an antibody which binds uPA and prevents uPA binding
to uPAR. MCF-7 cells that were cultured in RAP demonstrated increased leve
ls of activated mitogen-activated protein kinases, Furthermore, the MEK inh
ibitor, PD098059, decreased the motility of RAP-treated cells without affec
ting control cultures. These studies suggest a model in which the VLDLr reg
ulates autocrine uPAR-initiated signaling and thereby regulates cellular mo
tility.