Role of p190RhoGAP in beta 2 integrin regulation of RhoA in human neutrophils.
We found that engagement of beta(2) integrins on human neutrophils induced activation of RhoA, as indicated by the increased ratio of GTP:GTP + GDP recovered on RhoA and translocation of RhoA to a membrane fraction. The clustering of beta(2) integrins also induced a time-dependent increase in GDP bound to RhoA, which correlated with beta(2) integrin-induced activation of p190RHOGAP: The activation of p190RhoGAP was completely blocked by [4-amino-5-(4-methylphenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine] (PP1), a selective inhibitor of Src family tyrosine kinases. However, clustering of beta(2) integrins did not increase the basal tyrosine phosphorylation of p190RhoGAP, nor did it affect the amount of p120RasGAP bound to p190RHOGAP: Instead, the beta(2) integrin-induced activation of p190RhoGAP was accompanied by increased tyrosine phosphorylation of a p190RhoGAP-associated protein, p120RasGAP, and accumulation of both p120RasGAP and p190RhoGAP in a membrane fraction. PP1 blocked the beta(2) integrin-induced phosphorylation of p120RasGAP, as well as the translocation of p190RhoGAP and p120RasGAP, but it did not affect the accumulation of RhoA in the membrane fraction. In agreement with the mentioned findings, PP1 also increased the GTP:GTP + GDP ratio recovered on RhoA immunoprecipitated from beta(2) integrin-stimulated cells. Thus, in neutrophils, beta(2) integrin-induced activation of p190RhoGAP requires a signal from a Src family tyrosine kinase, but it does not occur via the signaling pathway responsible for activation of RHOA:
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Division of Experimental Pathology, Department of Laboratory Medicine, Lund University, Malmö University Hospital, Malmö, Sweden. karim.dib@exppat.mas.lu.se
Melander
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Andersson
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D018821:CD18 Antigens / Q000378:metabolism / Q000502:physiology*; D002462:Cell Membrane / Q000201:enzymology / Q000378:metabolism; D002469:Cell Separation; D020690:GTPase-Activating Proteins / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism / Q000502:physiology*; D006801:Humans; D006868:Hydrolysis; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009687:Nuclear Proteins / Q000378:metabolism / Q000502:physiology*; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism; D011720:Pyrazoles / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D012097:Repressor Proteins; D015398:Signal Transduction / Q000187:drug effects; D020729:p120 GTPase Activating Protein / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism*
C108123:4-amino-5-(4-methylphenyl)-7-(tert-butyl)pyrazolo(3,4-d)pyrimidine; C074560:ARHGAP35 protein, human; C513043:ARHGAP5 protein, human; D018821:CD18 Antigens; D020690:GTPase-Activating Proteins; D020662:Guanine Nucleotide Exchange Factors; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011720:Pyrazoles; D011743:Pyrimidines; D012097:Repressor Proteins; D020729:p120 GTPase Activating Protein; C085379:rho GTPase-activating protein; D019000:Phosphotyrosine; D020742:rhoA GTP-Binding Protein