Abstract
Binding of the urokinase-type plasminogen activator (uPA) to its receptor activates diverse cell signaling pathways. How these signals are integrated so that cell physiology is altered remains unclear. In this study, we demonstrated that migration of MCF-7 breast cancer cells and HT-1080 fibrosarcoma cells on serum-coated surfaces is stimulated by agents that activate ERK, including uPA, epidermal growth factor, and constitutively active MEK1. The promigratory activity of these agents was entirely blocked not only by the MEK-specific antagonist PD098059, but also by antagonists of the Rho-Rho kinase pathway, including Y-27632 and dominant-negative RhoA (RhoA-N19). uPA did not significantly increase the level of GTP-bound RhoA, suggesting that the constitutive activity of the Rho-Rho kinase pathway may be sufficient to support ERK-stimulated cell migration. Paradoxically, Y-27632 and RhoA-N19 increased ERK phosphorylation in MCF-7 cells, providing further evidence that ERK activation alone does not promote cell migration when Rho kinase is antagonized. When MCF-7 cell migration was stimulated by ERK-independent processes such as expression of the beta(3) integrin subunit or changing the substratum to type I collagen, Y-27632 and RhoA-N19 failed to inhibit the response. This study supports a model in which the Ras-ERK and Rho-Rho kinase pathways cooperate to promote cell migration. Neutralizing either pathway is sufficient to block the response to agents that stimulate cell migration by activating ERK.
Highlights
The urokinase-type plasminogen activator1 is a serine proteinase that activates diverse cell signaling pathways by binding to its cell-surface receptor [1,2,3]
We demonstrated that migration of MCF-7 breast cancer cells and HT-1080 fibrosarcoma cells on serum-coated surfaces is stimulated by agents that activate ERK, including urokinase-type plasminogen activator (uPA), epidermal growth factor, and constitutively active MEK1
In many cell types, binding of uPA to uPAR results in activation of the mitogen-activated protein kinase ERK (7, 10 –13). This signaling response may be of central importance because the level of activated ERK controls many physiologic processes, including cell growth, differentiation, apoptosis, and migration, and in cancer, invasion and metastasis [1]. uPA-induced ERK activation in MCF-7 breast cancer cells is entirely dependent on the small GTPase Ras [7], which presumably functions through the downstream effectors Raf and MEK [14]
Summary
The urokinase-type plasminogen activator (uPA) is a serine proteinase that activates diverse cell signaling pathways by binding to its cell-surface receptor (uPAR) [1,2,3]. In many cell types, binding of uPA to uPAR results in activation of the mitogen-activated protein kinase ERK (7, 10 –13) This signaling response may be of central importance because the level of activated ERK controls many physiologic processes, including cell growth, differentiation, apoptosis, and migration, and in cancer, invasion and metastasis [1]. The function of Tyk and phosphatidylinositol 3-kinase in smooth muscle cell migration [19] raises the possibility that distinct signaling pathways may be preeminent in determining physiologic responses to uPA in different cell types. Recent studies suggest that RhoA, RhoC, and Rho kinase may be important in cell migration and cancer invasion (24 –29)
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