The GIT-PIX complexes regulate the chemotactic response of rat basophilic leukaemia cells

Manuela Gavina, Lorena Za, Raffaella Molteni, Ruggero Pardi, Ivan de Curtis

Research output: Contribution to journalArticle

Abstract

Background information. Cell motility entails the reorganization of the cytoskeleton and membrane trafficking for effective protrusion. The GIT-PIX protein complexes are involved in the regulation of cell motility and adhesion and in the endocytic traffic of members of the family of G-protein-coupled receptors. We have investigated the function of the endogenous GIT complexes in the regulation of cell motility stimulated by fMLP (formyl-Met-Leu-Phe) peptide, in a rat basophilic leukaemia RBL-2H3 cell line stably expressing an HA (haemagglutinin)-tagged receptor for the fMLP peptide. Results. Our analysis shows that RBL cells stably transfected with the chemoattractant receptor expressed both GIT1-PIX and GIT2-PIX endogenous complexes. We have used silencing of the different members of the complex by small interfering RNAs to study the effects on a number of events linked to agonist-induced cell migration. We found that cell adhesion was not affected by depletion of any of the proteins of the GIT complex, whereas agonist-enhanced cell spreading was inhibited. Analysis of agonist-stimulated haptotactic cell migration indicated a specific positive effect of GIT1 depletion on trans-well migration. The internalization of the formyl-peptide receptor was also inhibited by depletion of GIT1 and GIT2. The effects of the GIT complexes on trafficking of the receptors was confirmed by an antibody-enhanced agonist-induced internalization assay, showing that depletion of PIX, GIT1 or GIT2 protein caused decreased perinuclear accumulation of internalized receptors. Conclusions. Our results show that endogenous GIT complexes are involved in the regulation of chemoattractantinduced cell motility and receptor trafficking, and support previous findings indicating an important function of the GIT complexes in the regulation of different G-protein-coupled receptors. Our results also indicate that endogenous GIT1 and GIT2 regulate distinct subsets of agonist-induced responses and suggest a possible functional link between the control of receptor trafficking and the regulation of cell motility by GIT proteins.

Original languageEnglish
Pages (from-to)231-244
Number of pages14
JournalBiology of the cell / under the auspices of the European Cell Biology Organization
Volume102
Issue number4
DOIs
Publication statusPublished - 2010

Keywords

  • Adhesion
  • Chemotactic response
  • Endocytosis
  • GIT-PIX complex
  • Migration
  • Spreading

ASJC Scopus subject areas

  • Cell Biology

Fingerprint Dive into the research topics of 'The GIT-PIX complexes regulate the chemotactic response of rat basophilic leukaemia cells'. Together they form a unique fingerprint.

  • Cite this