rho family GTPase activating proteins p190, bcr and rhoGAP show distinct specificities in vitro and in vivo.
rho family GTPases link extracellular signals to changes in the organization of cytoskeletal actin. Serum stimulation of quiescent Swiss 3T3 fibroblasts leads to rho-dependent actin stress fibre formation and focal adhesions, whilst several growth factors initiate signalling pathways leading to rac-dependent actin polymerization at the plasma membrane, and membrane ruffling. The product of the breakpoint cluster region gene bcr, rho GTPase accelerating protein (rhoGAP) and rasGAP-associated p190 share structurally related rho GAP domains, and possess GAP activity for rho family members in vitro. We have directly compared the activities of the isolated GAP domains of these three proteins in regulating different rho family GTPases, both by in vitro assays and by microinjection, to address their possible physiologic functions. We show that bcr accelerates the GTPase activity of rac, but not rho in vitro, and inhibits rac-mediated membrane ruffling, but not rho-mediated stress fibre formation, after microinjection into Swiss 3T3 fibroblasts. In vitro, rhoGAP has a striking preference for G25K as a substrate, whilst p190GAP has marked preferential activity for rho. Furthermore, p190 preferentially inhibits rho-mediated stress fibre formation in vivo. Our data suggest that p190, rhoGAP and bcr play distinct roles in signalling pathways mediated through different rho family GTPases.
Ridley
A J
AJ
Institute for Cancer Research, London, UK.
Self
A J
AJ
Kasmi
F
F
Paterson
H F
HF
Hall
A
A
Marshall
C J
CJ
Ellis
C
C
D016475:3T3 Cells; D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000818:Animals; D004268:DNA-Binding Proteins; D019204:GTP-Binding Proteins / Q000378:metabolism*; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D051379:Mice; D008845:Microinjections; D009687:Nuclear Proteins / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010982:Platelet-Derived Growth Factor / Q000037:antagonists & inhibitors; D011505:Protein-Tyrosine Kinases; D011506:Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051562:Proto-Oncogene Proteins c-bcr; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D015398:Signal Transduction; D013379:Substrate Specificity; D020744:rac GTP-Binding Proteins
C513041:Arhgap35 protein, mouse; C513042:Arhgap5 protein, mouse; D004268:DNA-Binding Proteins; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D010750:Phosphoproteins; D010982:Platelet-Derived Growth Factor; D011506:Proteins; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C085379:rho GTPase-activating protein; D011505:Protein-Tyrosine Kinases; C496614:Bcr protein, mouse; D051562:Proto-Oncogene Proteins c-bcr; D019204:GTP-Binding Proteins; D020744:rac GTP-Binding Proteins