Specific recognition of Rac2 and Cdc42 by DOCK2 and DOCK9 guanine nucleotide exchange factors.
Recognition of cognate Rho GTPases by guanine-nucleotide exchange factors (GEF) is fundamental to Rho GTPase signaling specificity. Two main GEF families use either the Dbl homology (DH) or the DOCK homology region 2 (DHR-2) catalytic domain. How DHR-2-containing GEFs distinguish between the GTPases Rac and Cdc42 is not known. To determine how these GEFs specifically recognize the two Rho GTPases, we studied the amino acid sequences in Rac2 and Cdc42 that are crucial for activation by DOCK2, a Rac-specific GEF, and DOCK9, a distantly related Cdc42-specific GEF. Two elements in the N-terminal regions of Rac2 and Cdc42 were found to be essential for specific interactions with DOCK2 and DOCK9. One element consists of divergent amino acid residues in the switch 1 regions of the GTPases. Significantly, these residues were also found to be important for GTPase recognition by Rac-specific DOCK180, DOCK3, and DOCK4 GEFs. These findings were unexpected because the same residues were shown previously to interact with GTPase effectors rather than GEFs. The other element comprises divergent residues in the beta3 strand that are known to mediate specific recognition by DH domain containing GEFs. Remarkably, Rac2-to-Cdc42 substitutions of four of these residues were sufficient for Rac2 to be specifically activated by DOCK9. Thus, DOCK2 and DOCK9 specifically recognize Rac2 and Cdc42 through their switch 1 as well as beta2-beta3 regions and the mode of recognition via switch 1 appears to be conserved among diverse Rac-specific DHR-2 GEFs.
Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724; Program in Genetics and Medical Scientist Training Program, Stony Brook University, Stony Brook, New York 11790.
Michael A
MA
Kwofie
Cold Spring Harbor Laboratory, Cold Spring Harbor, New York 11724. Electronic address: skowrons@cshl.edu.
Jacek
J
Skowronski
D000595:Amino Acid Sequence; D020690:GTPase-Activating Proteins; D005786:Gene Expression Regulation; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D020764:cdc42 GTP-Binding Protein / Q000737:chemistry*; D020744:rac GTP-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D000097766:RAC2 GTP-Binding Protein
D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D011994:Recombinant Proteins; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins; D000097766:RAC2 GTP-Binding Protein; C402110:DOCK2 protein, human; C466321:DOCK9 protein, human