Levin, N. K.; Dahlgren, C.; Forsman, H.; Sundqvist, M.

Signaling by formyl peptide receptor 1 (FPR1), the prototype G protein-coupled receptor (GPCR) expressed in neutrophil leukocytes, is initiated by an activation of a G protein containing a Galphai subunit. FPR1 activation results in an increase in the cytosolic concentration of free calcium ions ([Ca2+]i), and an activation of the superoxide anion producing NADPH oxidase. Receptor downstream signals generated by the danger molecule ATP recognized by the purinergic receptor P2Y2 are transduced by a G protein containing a Galphaq subunit. The neutrophil response induced by ATP also includes a transient rise in [Ca2+]i, but the downstream signals do not activate the NADPH oxidase. ATP can, however, activate this enzyme system through a receptor transactivation mechanism dependent not only on the ATP receptor but also on the free fatty acid receptor FFA2R, provided that this receptor is allosterically modulated. This occurs through a novel mechanism whereby FFA2R is activated from the cytosolic side of the plasma membrane by Galphaq transduced signals generated by the ATP receptor. Furthermore, in neutrophils with a disrupted actin cytoskeleton, ATP (as well as platelet activating factor; recognized by the Galphaq-coupled PAFR) becomes a potent NADPH oxidase activating agonist. At high concentrations of the actin cytoskeleton disrupting drug latrunculin A the activation was only partly reduced by Galphaq inhibition. More importantly, this response was also partly inhibited by pertussis toxin. The effects on the ATP-induced NADPH oxidase activity, of the Galphaq inhibitor and pertussis toxin were more and less pronounced, respectively, when the concentration of latrunculin A was reduced. Taken together, we show that in primary human neutrophils the actin cytoskeleton is part of the regulatory machinery that determines the activation of NADPH oxidase activation and the G protein recruitment profile downstream of activated of Galphaq-coupled GPCRs.