title: 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/Sps-Shc complex is associated with the transient phosphorylation of ERK in urokinase-treated cells.
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.
authors.affiliation: Departments of Biochemistry and Molecular Biology, Pathology, and Microbiology, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
authors.forename: D H
authors.initials: DH
authors.lastname: Nguyen
authors.forename: D J
authors.initials: DJ
authors.lastname: Webb
authors.forename: A D
authors.initials: AD
authors.lastname: Catling
authors.forename: Q
authors.initials: Q
authors.lastname: Song
authors.forename: A
authors.initials: A
authors.lastname: Dhakephalkar
authors.forename: M J
authors.initials: MJ
authors.lastname: Weber
authors.forename: K S
authors.initials: KS
authors.lastname: Ravichandran
authors.forename: S L
authors.initials: SL
authors.lastname: Gonias
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D002465:Cell Movement / Q000502:physiology*; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D051380:GRB2 Adaptor Protein; D006801:Humans; D020935:MAP Kinase Signaling System / Q000502:physiology*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000502:physiology; D011506:Proteins / Q000378:metabolism; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000502:physiology*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D051380:GRB2 Adaptor Protein; C495948:GRB2 protein, human; D011506:Proteins; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496050:PTK2 protein, human; D016392:Proto-Oncogene Proteins pp60(c-src); D014568:Urokinase-Type Plasminogen Activator