Multiple Factors Confer Specific Cdc42 and Rac Protein Activation by Dedicator of Cytokinesis (DOCK) Nucleotide Exchange Factors

Abstract

To understand the molecular basis for DOCK-GTPase specificity, we have determined the crystal structure of DOCK2 DHR2 in complex with Rac1. DOCK2 DHR2 and DOCK9 DHR2 exhibit similar tertiary structures and homodimer interfaces and share a conserved GTPase-activating mechanism. Multiple structural differences between DOCK2 DHR2 and DOCK9 DHR2 account for their selectivity toward Rac1 and Cdc42. Key determinants of selectivity of Cdc42 and Rac for their cognate DOCK DHR2 are a Phe or Trp residue within ␤3 (residue 56) and the ability of DOCK proteins to exploit differences in the GEF-induced conformational changes of switch 1 dependent on a divergent residue at position 27. DOCK proteins, therefore, differ from DH-PH GEFs that select their cognate GTPases through recognition of structural differences within the ␤2/␤3 strands.

The role of Rac1 in the regulation of NF-kB activity, cell proliferation, and cell migration in non-small cell lung carcinoma

“…For example, VAV2 has been shown to activate Rac1, Cdc42 and RhoA, 36 whereas DOCK6 has been shown to activate Rac1 and Cdc42. 37,38 Thus, the interaction of DN-Rac1 with GEFs such as VAV2 and DOCK6 might suppress the activities of RhoA and Cdc42, in addition to Rac1. Therefore, G 2 arrest induced by DN-Rac1 might be due to the inhibition of pathways mediated by multiple small GTPases in addition to Rac1.…”

Pharmacological inhibition of Dock5 prevents osteolysis by affecting osteoclast podosome organization while preserving bone formation

“…The DHR2 domain of mouse Dock5 (residues 1212-1642, Dock5-DHR2, as defined for Dock2 in ref. 13) was expressed in Escherichia coli as a 6 Â His-MBP fusion with a TEV protease cleavage site. The tag was cleaved and the protein was purified to homogeneity ( Supplementary Fig.…”

“…Nucleotide exchange was assayed by fluorescence kinetics using a highly purified Dock5 construct encompassing its previously described DHR2 (DOCK homology region 2) catalytic and dimerization domain 13 and the GTPases Rac1, Rac2, RhoA and Cdc42 ( Supplementary Fig. 1a,b).…”

CDC-42 and RAC-1 regulate opposite chemotropisms inNeurospora crassa

“…Key determinants of selectivity of Cdc42 and Rac1 for their cognate GEFs is conferred through specific residues in the GEF-binding domains of both GTPases. A tryptophan or tyrosine residue at position 56 is specific for CDC-42, whereas a phenylalanine residue at position 56 and an alanine residue at position 27 are specific for Rac (Gao et al, 2001;Karnoub et al, 2001;Kulkarni et al, 2011). Recent studies on the filamentous fungus N. crassa identified an important role for CDC-42 in initiating cell polarity, whereas RAC-1 was predominantly involved in the maintenance of polarized tip growth (AraujoPalomares et al, 2011).…”