In vivo dynamics of Rac-membrane interactions

Mol Biol Cell. 2006 Jun;17(6):2770-9. doi: 10.1091/mbc.e06-01-0005. Epub 2006 Apr 5.


The small GTPase Rac cycles between the membrane and the cytosol as it is activated by nucleotide exchange factors (GEFs) and inactivated by GTPase-activating proteins (GAPs). Solubility in the cytosol is conferred by binding of Rac to guanine-nucleotide dissociation inhibitors (GDIs). To analyze the in vivo dynamics of Rac, we developed a photobleaching method to measure the dissociation rate constant (k(off)) of membrane-bound GFP-Rac. We find that k(off) is 0.048 s(-1) for wtRac and approximately 10-fold less (0.004 s(-1)) for G12VRac. Thus, the major route for dissociation is conversion of membrane-bound GTP-Rac to GDP-Rac; however, dissociation of GTP-Rac occurs at a detectable rate. Overexpression of the GEF Tiam1 unexpectedly decreased k(off) for wtRac, most likely by converting membrane-bound GDP-Rac back to GTP-Rac. Both overexpression and small hairpin RNA-mediated suppression of RhoGDI strongly affected the amount of membrane-bound Rac but surprisingly had only slight effects on k(off). These results indicate that RhoGDI controls Rac function mainly through effects on activation and/or membrane association.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • 3T3 Cells
  • Animals
  • Cell Membrane / physiology*
  • Cell Membrane / ultrastructure
  • Computer Simulation
  • Diffusion
  • Genes, Reporter
  • Guanine Nucleotide Dissociation Inhibitors / deficiency
  • Guanine Nucleotide Dissociation Inhibitors / genetics
  • Guanine Nucleotide Dissociation Inhibitors / metabolism
  • Kinetics
  • Mice
  • Microscopy, Confocal
  • Models, Theoretical
  • Plasmids
  • Protein Transport
  • Recombinant Fusion Proteins
  • Recombinant Proteins / metabolism
  • rac GTP-Binding Proteins / genetics
  • rac GTP-Binding Proteins / metabolism*
  • rho-Specific Guanine Nucleotide Dissociation Inhibitors


  • Guanine Nucleotide Dissociation Inhibitors
  • Recombinant Fusion Proteins
  • Recombinant Proteins
  • rho-Specific Guanine Nucleotide Dissociation Inhibitors
  • rac GTP-Binding Proteins