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

Diem H.D. Nguyen, Donna J. Webb, Andrew D. Catling, Qian Song, Anita Dhakephalkar, Michael J. Weber, Kodimangalam S. Ravichandran, Steven L. Gonias

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Abstract

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 that soluble uPA receptor and a peptide derived from the linker region between domains 1 and 2 of the uPA receptor also stimulate cellular migration via a mitogen-activated protein kinase/ERK kinase (MEK)-dependent pathway. Signaling proteins that function upstream of Ras in uPA-stimulated cells remain undefined. To address this problem, we transfected MCF-7 cells to express the noncatalytic carboxylterminal domain of focal adhesion kinase (FAK), FAK(Y397F), kinase-defective c-Src, or Shc FFF, all of which express dominant-negative activity. In each case, ERK phosphorylation and cellular migration in response to uPA were blocked. Both activities were rescued by co-transfecting 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 with Shc in cells that were treated with uPA for 1-2.5 min, probably reflecting the formation of Shc-Grb2/Sos complex; however, by 10 min, co- immunoprecipitation of Sos with Shc was no longer observed. Rapid dissociation of Sos from Shc represents a possible mechanism for the transient phosphorylation of ERK in uPA-treated MCF-7 cells.

Original languageEnglish
Pages (from-to)19382-19388
Number of pages7
JournalJournal of Biological Chemistry
Volume275
Issue number25
DOIs
StatePublished - Jun 23 2000

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