Urokinase-type plasminogen activator stimulates the Ras/extracellular signal-regulated kinase (ERK) signaling pathway and MCF-7 cell migration by a mechanism that requires focal adhesion kinase, Src, and Shc - Rapid dissociation of GRB2/SOS-SHC complex is associated with the transient phosphorylation of ERK in urokinase-treated cells

Urokinase-type plasminogen activator (uPA) stimulates MCF-7 cell migration by binding to the UPA receptor and activating the Ras-extracellular signal- regulated kinase (Ras-ERK) signaling pathway. Studies presented here show t hat soluble uPA receptor and a peptide derived from the Linker region betwe en domains 1 and 2 of the uPA receptor also stimulate cellular migration vi a a mitogen-activated protein kinase/ERK kinase (MEK)-dependent pathway. Si gnaling proteins that function upstream of Res in uPA-stimulated cells rema in undefined. To address this problem, we transfected MCF-7 cells to expres s the noncatalytic carboxylterminal domain of focal adhesion kinase (FAK), FAK(Y397F), kinase-defective c-Src, or Shc FFF, all of which express domina nt-negative activity. In each case, ERK phosphorylation and cellular migrat ion in response to uPA were blocked. Both activities were rescued by co-tra nsfecting the cells to express constitutively active MEK1, indicating that FAK, c-Src, and Shc are upstream of MEK. Shc was tyrosine phosphorylated in uPA-treated cells. The level of phosphorylated Shc was increased within 1 min and remained increased for at least 30 min. Sos co-immunoprecipitated w ith Shc in cells that were treated with uPA for 1-2.5 min, probably reflect ing the formation of Shc-Grb2/Sos complex; however, by 10 min, co-immunopre cipitation of Sos with Shc was no longer observed. Rapid dissociation of So s from Shc represents a possible mechanism for the transient phosphorylatio n of ERK in uPA-treated MCF-7 cells.