Glycoprotein capture and quantitative phosphoproteomics indicate coordinated regulation of cell migration upon lysophosphatidic acid stimulation

Mol Cell Proteomics. 2010 Nov;9(11):2337-53. doi: 10.1074/mcp.M110.000737. Epub 2010 Jul 16.


The lipid mediator lysophosphatidic acid (LPA) is a serum component that regulates cellular functions such as proliferation, migration, and survival via specific G protein-coupled receptors. The underlying signaling mechanisms are still incompletely understood, including those that operate at the plasma membrane to modulate cell-cell and cell-matrix interactions in LPA-promoted cell migration. To explore LPA-evoked phosphoregulation with a focus on cell surface proteins, we combined glycoproteome enrichment by immobilized lectins with SILAC-based quantitative phosphoproteomics. We performed biological replicate analyses in SCC-9 squamous cell carcinoma cells and repeatedly quantified the effect of 1.5- and 5-min LPA treatment on more than 700 distinct phosphorylations in lectin-purified proteins. We detected many regulated phosphorylation events on various types of plasma membrane proteins such as cell adhesion molecules constituting adherens junctions, desmosomes, and hemidesmosomes. Several of these LPA-regulated phosphorylation sites have been characterized in a biological context other than G protein-coupled receptor signaling, and the transfer of this functional information suggests coordinated and multifactorial cell adhesion control in LPA-induced cell migration. Additionally, we identified LPA-mediated activation loop phosphorylation of the serine/threonine kinase Wnk1 and verified a role of Wnk1 for LPA-induced cell migration in knock-down experiments. In conclusion, the glycoproteome phosphoproteomics strategy described here sheds light on incompletely understood mechanisms in LPA-induced cell migratory behavior.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Cell Line, Tumor
  • Cell Movement / drug effects*
  • Computational Biology
  • ErbB Receptors / genetics
  • ErbB Receptors / metabolism
  • Glycoproteins / chemistry*
  • Humans
  • Lysophospholipids / pharmacology*
  • Molecular Sequence Data
  • Phosphoproteins / chemistry*
  • Proteomics / methods*
  • Receptors, G-Protein-Coupled / metabolism
  • Receptors, Lysophosphatidic Acid / metabolism


  • Glycoproteins
  • Lysophospholipids
  • Phosphoproteins
  • Receptors, G-Protein-Coupled
  • Receptors, Lysophosphatidic Acid
  • ErbB Receptors
  • lysophosphatidic acid