Regulation of Tiam1 nucleotide exchange activity by pleckstrin domain binding ligands

Rho family GTPases play roles in cytoskeletal organization and cellular tra nsformation. Tiam1 is a member of the Dbl family of guanine nucleotide exch ange factors that activate Rho family GTPases. These exchange factors have in common a catalytic Dbl homology and adjacent pleckstrin homology domain. Previous structural studies suggest that the pleckstrin domain, a putative phosphoinositide-binding site, may serve a regulatory function. We identif ied ascorbyl stearate as a compound that binds to the pleckstrin domain of p120 Ras GTPase-activating protein. Furthermore, ascorbyl stearate appears to be a general pleckstrin domain ligand, perhaps by mimicking an endogenou s amphiphilic ligand. Tiam1 nucleotide exchange activity was greatly stimul ated by ascorbyl stearate. Certain phosphoinositides also stimulated Tiam1 activity but were less potent than ascorbyl stearate. Tiam1 contains an add itional N-terminal pleckstrin domain, but only the C-terminal pleckstrin do main was required for activation. Our results suggest that the pleckstrin d omains of Dbl-type proteins may not only be involved in subcellular localiz ation but may also directly regulate the nucleotide exchange activity of an associated Dbl homology domain. In addition, this paper introduces ascorby l stearate as a pleckstrin domain ligand that can modulate the activity of certain pleckstrin domain-containing proteins.