Down-regulation of epidermal growth factor receptor correlates with plasminogen activator activity in human A431 epidermoid carcinoma cells.

Abstract

Human A431 epidermoid carcinoma cells in culture exhibit epidermal growth factor (EGF)-induced "down-regulation" of cell-surface and total cellular (Triton X-100 extractable) EGF receptors caused entirely by an enhanced rate (4-fold) of receptor inactivation [Krupp, M. N., Connolly, D. T. & Lane, M. D. (1982) J. Biol. Chem. 257, 11489-11496]. The following observations show that this enhanced rate of EGF receptor inactivation is closely correlated with an increased cellular activity of plasminogen activator (PA), a serine protease. First, EGF-induced down-regulation of cell-surface and total cellular EGF receptors and the concomitant increase in cellular PA activity occur with identical kinetics, the tl/2 for both processes being 3-3.5 hr.Second, the EGF dose-response curves for down-regulation of total cellular EGF receptor and increased PA activity are similar. The EGF concentrations for half-maximal responses of both processes are 10-15 nM and 20 nM, respectively. Third, the removal of EGF from previously down-regulated cells results in the recovery of total cellular EGF binding activity with a concurrent loss of cellular PA activity. Fourth, blocking PA synthesis or activity with cycloheximide or dexamethasone prevents down-regulation of the EGF receptor. Fifth, the addition of leupeptin, an inhibitor of PA and plasmin action, blocks EGF-induced receptor downregulation as well as the increase of PA activity. That EGF receptor down-regulation is independent of plasminogen per se in the culture medium suggests that PA-mediated events may initiate the rapid inactivation of the EGF receptor that occurs during downregulation.The interaction of epidermal growth factor (EGF), a polypeptide hormone, with specific cell-surface receptors initiates numerous biochemical events (1) in target cells, most notably mitogenesis (2). Such factors as the period of exposure to EGF, the concentration of EGF, and the level of functional receptors at the plasma membrane affect the magnitude of ligand-receptor interactions and, thus, the cellular response to EGF (3-5). It has been established that cells can modulate their level of surface EGF receptors in response to EGF and thereby alter the biological response to this potent mitogen (3-5).Like other receptors for polypeptide hormones (6), the EGF receptor exhibits ligand-induced "down-regulation" of cell-surface EGF binding capacity (3-5). This negative modulation of EGF receptor level in vitro is rapid, EGF concentration dependent, and EGF specific and occurs in many cell types (1, 3, 5). Reduced EGF binding capacity is the result of a lowered EGF receptor level rather than an altered affinity of the EGF receptor for its ligand (3,5 Although the mechanism by which ligand-induced downregulation promotes EGF receptor inactivation is unknown, there is some evidence with another cell-surface receptor that proteolysis modulates receptor metabolism. When proteolysis in chicken myogenic cultures is induced by chemical or viral transformation, both the steady-state concentratio...

The plasminogen activator/plasmin system.

“…In addition, proteolytic inactivation of growth factors could involve the PA/plasmin system. Modulation of expression ofreceptors for growth factors, hormones, and neurotransmitters may also be achieved by PA-catalyzed proteolysis, as initially suggested for the EGF receptor (50). Finally, it is conceivable that PAs and PAIs could play a part in controlling the plasticity of synaptic structures.…”

Binding of tissue plasminogen activator to cultured human endothelial cells.

“…However, it is not clear whether large vessel endothelial cells behave similarly. In contrast, u-PA binds to human monocytes and a monocytic cell line (U937) (16,17), normal and virus-transformed fibroblasts (3T3) (18,19), a human epidermoid carcinoma (A43 1) cell line (20)(21)(22), and possibly bovine corneal (23) and capillary (15) endothelial cells.…”

The cytoplasmic amino-terminus of the Latent Membrane Protein-1 of Epstein-Barr Virus: relationship between transmembrane orientation and effector functions of the carboxy-terminus and transmembrane domain

“…Like ligand-activated receptors (i.e. epidermal growth factor and insulin receptors (Gross et al, 1983;Knutson et al, 1985), LMP-1's turnover, but not internalization, requires ongoing protein synthesis (Martin and Sugden, 1991a). The properties of LMP-1's turnover, and the correlation between constitutive activity and turnover (Martin and Sugden, 1991b), suggest LMP-1's turnover is triggered by activated signaling and results in LMP-1's downregulation.…”

“…Like cell surface receptors (Gross et al, 1983;Haigler et al, 1978;Schlessinger et al, 1978;Yarden and Ullrich, 1988), LMP-1 and its functional deletion mutants localize at the cell surface in aggregated patches (Liebowitz et al, 1986;Mann et al, 1985;Martin and Sugden, 1991b), associate with the cytoskeleton (Liebowitz et al, 1987;Mann and Thorley-Lawson, 1987;Martin and Sugden, 1991b), and turn over rapidly in the cell (Baichwal and Sugden, 1987;Mann and Thorley-Lawson, 1987;Martin and Sugden, 1991b). LMP-1's tertiary structure, biochemical properties, interaction with cellular signaling proteins , and self-oligomerization (Eliopoulos and Rickinson, 1998;Eliopoulos and Young, 1998;Floettmann and Rowe, 1997;Gires et al, 1997;Hatzivassiliou et al, 1998) are consistent with LMP-1 functioning as a constitutively active cell surface receptor.…”