EGF-stimulated migration in ovarian cancer cells is associated with decreased internalization, increased surface expression, and increased shedding of the urokinase plasminogen activator receptor

E. Henic, M.K. Sixt, S. Hansson, G. Høyer Hansen, B. Casslén, Gynecologic Oncology 101 (2006) 28–39.

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Abstract
OBJECTIVES: The EGFR is expressed in malignant ovarian tumor tissue, and tissue content of EGFR has been directly associated with poor prognosis in patients with ovarian cancer. The uPA system plays a role in pericellular proteolysis, cell migration, invasion, and is over-expressed in ovarian cancer. This study explored the effects of EGF on uPAR expression in the ovarian cancer cell line OVCAR-3. METHODS: We used OVCAR-3 cells and the following methods: cell migration assay, time-lapse video microscopy, real-time PCR, assays for cellular binding of 125I-uPA and cellular degradation of 125I-uPA:PAI-1 complex, biosynthetic labeling using 35S-methionin, Western blot, Northern blot, and ELISAs for uPA, PAI-1, and uPAR. RESULTS: EGF up-regulates both protein and mRNA not only for uPAR, but also for the ligand uPA and its inhibitor PAI-1. Cell surface uPAR, in control as well as EGF-stimulated cells, is present only in the intact, not the cleaved, form. Ligand binding experiments showed an increase of endogenously occupied uPAR, whereas non-occupied receptor sites were not increased. In addition, EGF treatment resulted in decreased degradation of radiolabeled uPA:PAI-1 complex. This suggests decreased internalization of uPAR, since the complex is internalized together with uPAR. Like EGF, colchicine, which inhibits endocytosis, increased cell surface expression of uPAR. In addition, we found an immediate increase of uPAR after exposing the cells to EGF and this was accompanied by a transient increase of cell migration. The increase of cell surface uPAR in response to EGF is accompanied by increased release of the soluble form of uPAR (suPAR) to the medium as well as by increased cell migration. Both uPAR and suPAR increased in cells treated with the endocytosis inhibitor colchicine even though cell migration was inhibited, suggesting that the mechanism of uPAR shedding is not related to cell migration. CONCLUSION: Increased cell surface uPAR in response to EGF stimulation results from mobilization of uPAR from detergent-resistant domains, increased expression of uPAR mRNA, and decreased internalization and degradation of uPAR. Both the anti-uPAR antibody R3, which inhibits binding of uPA, and the EGFR phosphorylation inhibitor Iressa inhibited cell migration in response to uPA as well as to EGF, suggesting that EGFR and uPAR are engaged in the same multiprotein assembly on the cell surface.
Publishing Year
Date Published
2006-04-01
Journal Title
Gynecologic Oncology
Volume
101
Issue
1
Page
28 - 39
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Henic E, Sixt MK, Hansson S, Høyer Hansen G, Casslén B. EGF-stimulated migration in ovarian cancer cells is associated with decreased internalization, increased surface expression, and increased shedding of the urokinase plasminogen activator receptor. Gynecologic Oncology. 2006;101(1):28-39. doi:10.1016/j.ygyno.2005.09.038
Henic, E., Sixt, M. K., Hansson, S., Høyer Hansen, G., & Casslén, B. (2006). EGF-stimulated migration in ovarian cancer cells is associated with decreased internalization, increased surface expression, and increased shedding of the urokinase plasminogen activator receptor. Gynecologic Oncology, 101(1), 28–39. https://doi.org/10.1016/j.ygyno.2005.09.038
Henic, Emir, Michael K Sixt, Stefan Hansson, Gunilla Høyer Hansen, and Bertil Casslén. “EGF-Stimulated Migration in Ovarian Cancer Cells Is Associated with Decreased Internalization, Increased Surface Expression, and Increased Shedding of the Urokinase Plasminogen Activator Receptor.” Gynecologic Oncology 101, no. 1 (2006): 28–39. https://doi.org/10.1016/j.ygyno.2005.09.038.
E. Henic, M. K. Sixt, S. Hansson, G. Høyer Hansen, and B. Casslén, “EGF-stimulated migration in ovarian cancer cells is associated with decreased internalization, increased surface expression, and increased shedding of the urokinase plasminogen activator receptor,” Gynecologic Oncology, vol. 101, no. 1, pp. 28–39, 2006.
Henic E, Sixt MK, Hansson S, Høyer Hansen G, Casslén B. 2006. EGF-stimulated migration in ovarian cancer cells is associated with decreased internalization, increased surface expression, and increased shedding of the urokinase plasminogen activator receptor. Gynecologic Oncology. 101(1), 28–39.
Henic, Emir, et al. “EGF-Stimulated Migration in Ovarian Cancer Cells Is Associated with Decreased Internalization, Increased Surface Expression, and Increased Shedding of the Urokinase Plasminogen Activator Receptor.” Gynecologic Oncology, vol. 101, no. 1, Elsevier, 2006, pp. 28–39, doi:10.1016/j.ygyno.2005.09.038.

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