High constitutive activity of the human formyl peptide receptor

The formyl peptide receptor (FPR) couples to pertussis toxin (PTX)-sensitive Gi-proteins to activate chemotaxis and exocytosis in neutrophils. PTX reduces not only formyl peptide-stimulated but also agonist-independent ("basal") G(i)-protein activity, suggesting that the FPR is constitutively active. We aimed at identifying an inverse FPR agonist, i.e. a compound that suppresses constitutive FPR activity. In Sf9 insect cell membranes, the G-protein heterotrimer G(i)alpha(2)beta(1)gamma(2) reconstituted N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)stimulated guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) binding and GTP gamma S-sensitive high affinity [H-3]FMLP binding. The FPR "antagonist" cyclosporin H (CsH) potently and efficiently reduced basal GTP gamma S binding in Sf9 membranes. Another FPR antagonist, N-t-butoxycarbonyl-L-phenylalanyl-L-leucyl-L-phenylalanyl-L-leucyl-L-phenylalanine did not inhibit basal GTP gamma S binding but blocked the inhibitory effect of CsH on GTP gamma S binding. Na+ reduced basal GTP gamma S binding and eliminated the inhibitory effect of CsH. Similar effects of FMLP, CsH, and Na+ as in Sf9 membranes were observed with FPR expressed in the mammalian cell line HEK293. Our data show that the human FPR possesses high constitutive activity. CsH is an inverse FPR agonist and stabilizes the FPR in an inactive state. Na+ also stabilizes the FPR in an inactive state and, thereby, diminishes inverse agonist efficacy.