Down-regulation of Rac activity during beta 2 integrin-mediated adhesion of human neutrophils.
In human neutrophils, beta2 integrin engagement mediated a decrease in GTP-bound Rac1 and Rac2. Pretreatment of neutrophils with LY294002 or PP1 (inhibiting phosphatidylinositol 3-kinase (PI 3-kinase) and Src kinases, respectively) partly reversed the beta2 integrin-induced down-regulation of Rac activities. In contrast, beta2 integrins induced stimulation of Cdc42 that was independent of Src family members. The PI 3-kinase dependence of the beta2 integrin-mediated decrease in GTP-bound Rac could be explained by an enhanced Rac-GAP activity, since this activity was blocked by LY204002, whereas PP1 only had a minor effect. The fact that only Rac1 but not Rac2 (the dominating Rac) redistributed to the detergent-insoluble fraction and that it was independent of GTP loading excludes the possibility that down-regulation of Rac activities was due to depletion of GTP-bound Rac from the detergent-soluble fraction. The beta2 integrin-triggered relocalization of Rac1 to the cytoskeleton was enabled by a PI 3-kinase-induced dissociation of Rac1 from LyGDI. The dissociations of Rac1 and Rac2 from LyGDI also explained the PI 3-kinase-dependent translocations of Rac GTPases to the plasma membrane. However, these accumulations of Rac in the membrane, as well as that of p47phox and p67phox, were also regulated by Src tyrosine kinases. Inasmuch as Rac GTPases are part of the NADPH oxidase and the respiratory burst is elicited in neutrophils adherent by beta2 integrins, our results indicate that activation of the NADPH oxidase does not depend on the levels of Rac-GTP but instead requires a beta2 integrin-induced targeting of the Rac GTPases as well as p47phox and p67phox to the plasma membrane.
Dib
Karim
K
Department of Laboratory Medicine, Lund University, Malmö University Hospital, Entrance 78, SE-205 02 Malmö, Sweden. Karim.Dib@exppat.mas.lu.se
Melander
Fredrik
F
Axelsson
Lena
L
Dagher
Marie-Claire
MC
Aspenström
Pontus
P
Andersson
Tommy
T
D018821:CD18 Antigens / Q000502:physiology*; D002448:Cell Adhesion; D015536:Down-Regulation; D006801:Humans; D007700:Kinetics; D009504:Neutrophils / Q000166:cytology*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D021381:Protein Transport; D015398:Signal Transduction; D020764:cdc42 GTP-Binding Protein / Q000378:metabolism; D020744:rac GTP-Binding Proteins / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism*; D000097766:RAC2 GTP-Binding Protein
D018821:CD18 Antigens; D019869:Phosphatidylinositol 3-Kinases; D020764:cdc42 GTP-Binding Protein; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; D000097766:RAC2 GTP-Binding Protein