Rapid Remodeling of Invadosomes by Gi-coupled Receptors: DISSECTING THE ROLE OF Rho GTPases

J Biol Chem. 2016 Feb 26;291(9):4323-33. doi: 10.1074/jbc.M115.695940. Epub 2016 Jan 6.

Abstract

Invadosomes are actin-rich membrane protrusions that degrade the extracellular matrix to drive tumor cell invasion. Key players in invadosome formation are c-Src and Rho family GTPases. Invadosomes can reassemble into circular rosette-like superstructures, but the underlying signaling mechanisms remain obscure. Here we show that Src-induced invadosomes in human melanoma cells (A375M and MDA-MB-435) undergo rapid remodeling into dynamic extracellular matrix-degrading rosettes by distinct G protein-coupled receptor agonists, notably lysophosphatidic acid (LPA; acting through the LPA1 receptor) and endothelin. Agonist-induced rosette formation is blocked by pertussis toxin, dependent on PI3K activity and accompanied by localized production of phosphatidylinositol 3,4,5-trisphosphate, whereas MAPK and Ca(2+) signaling are dispensable. Using FRET-based biosensors, we show that LPA and endothelin transiently activate Cdc42 through Gi, concurrent with a biphasic decrease in Rac activity and differential effects on RhoA. Cdc42 activity is essential for rosette formation, whereas G12/13-mediated RhoA-ROCK signaling suppresses the remodeling process. Our results reveal a Gi-mediated Cdc42 signaling axis by which G protein-coupled receptors trigger invadosome remodeling, the degree of which is dictated by the Cdc42-RhoA activity balance.

Keywords: CDC42; G protein-coupled receptor (GPCR); Rac (Rac GTPase); Rho (Rho GTPase); biosensor; calcium intracellular release; fluorescence resonance energy transfer (FRET); imaging; invadopodia; phosphatidylinositide 3-kinase (PI 3-kinase).

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Biomarkers / metabolism
  • Cell Line, Tumor
  • Endothelins / metabolism*
  • Extracellular Matrix / metabolism
  • Extracellular Matrix / pathology
  • Fluorescence Resonance Energy Transfer
  • Humans
  • Hydrolysis
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Lysophospholipids / metabolism*
  • Melanoma / enzymology
  • Melanoma / metabolism*
  • Melanoma / pathology
  • Microscopy, Confocal
  • Microscopy, Fluorescence
  • Neoplasm Proteins / agonists
  • Neoplasm Proteins / antagonists & inhibitors
  • Neoplasm Proteins / genetics
  • Neoplasm Proteins / metabolism
  • Podosomes / enzymology
  • Podosomes / metabolism*
  • Podosomes / pathology
  • RNA Interference
  • Receptors, G-Protein-Coupled / antagonists & inhibitors
  • Receptors, G-Protein-Coupled / genetics
  • Receptors, G-Protein-Coupled / metabolism
  • Receptors, Lysophosphatidic Acid / agonists*
  • Receptors, Lysophosphatidic Acid / antagonists & inhibitors
  • Receptors, Lysophosphatidic Acid / genetics
  • Receptors, Lysophosphatidic Acid / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Time-Lapse Imaging
  • cdc42 GTP-Binding Protein / agonists*
  • cdc42 GTP-Binding Protein / antagonists & inhibitors
  • cdc42 GTP-Binding Protein / genetics
  • cdc42 GTP-Binding Protein / metabolism
  • rac1 GTP-Binding Protein / agonists
  • rac1 GTP-Binding Protein / antagonists & inhibitors
  • rac1 GTP-Binding Protein / genetics
  • rac1 GTP-Binding Protein / metabolism*
  • rhoA GTP-Binding Protein / antagonists & inhibitors
  • rhoA GTP-Binding Protein / genetics
  • rhoA GTP-Binding Protein / metabolism

Substances

  • Biomarkers
  • Endothelins
  • GPR26 protein, human
  • Luminescent Proteins
  • Lysophospholipids
  • Neoplasm Proteins
  • RAC1 protein, human
  • Receptors, G-Protein-Coupled
  • Receptors, Lysophosphatidic Acid
  • Recombinant Proteins
  • RHOA protein, human
  • cdc42 GTP-Binding Protein
  • rac1 GTP-Binding Protein
  • rhoA GTP-Binding Protein
  • lysophosphatidic acid