The function of the p190 Rho GTPase-activating protein is controlled by its N-terminal GTP binding domains

p190 is a GTPase-activating protein (GAP) for the Rho family of GTPases, Th e GAP domain of p190 is at the C terminus of the protein. At its N terminus , p190 contains a GTP binding domain of unknown significance. We have intro duced a mutation (Ser(36) --> Asn) into this domain of p190 that decreased its ability to bind guanine nucleotide when expressed as a hemagglutinin (H A)-tagged protein in COS cells, In vitro, both the wild type and S36N mutan t HA-p190 proteins showed similar GAP activities toward RhoA, but when expr essed in NIH 3T3 fibroblasts only wild type p190 appeared able to function as a RhoGAP. Wild type HA-p190 induced a phenotype of rounded cells with lo ng, beaded extensions similar to that seen when Rho function is disrupted b y ADP-ribosylation. HA-p190(S36N), although expressed at a similar level to the wild type protein, had no discernible effect on the cells. The beaded extension phenotype induced by wild type HA-p190 required GAP function. A G AP-defective mutant, p190(R1283A), had no effect on cell morphology. Moreov er, the beaded extension phenotype could be suppressed by co-expression of a gain-of-function Rho mutant, RhoA(G14V), or Pac mutant, Rac1(G12V). Activ ation of the Jun kinase (JNK) via muscarinic receptors was inhibited by wil d type HA-p190, but JNK activity was enhanced by the S36N mutant, Go-expres sion of HA-p190 with a fragment containing only the mutated GTP binding dom ain partially inhibited the beaded extension phenotype, suggesting that it may sequester a factor required for p190 function. Taken together these dat a demonstrate that within the cell, the Rho/Rac GAP activity of p190 can be regulated by the N-terminal GTP binding domain.