Receptor-stimulated dissociation of GTP[S] from G_i-proteins in membranes of HL-60 cells

Abstract

Formyl peptides stimulate binding of the stable GTP analogue, guanosine 5′-O[γ-thio]triphosphate (GTP[S]), to G-proteins in membranes of myeloid differentiated human leukaemia (HL-60) cells. On the other hand, agonist-activated formyl peptide receptors can also cause rapid and substantial release of GTP[S] bound to HL-60 membrane G-proteins. For fMet-Leu-Phe-stimulated dissociation of labelled GTP[S], an additional guanine nucleotide, in the potency order, unlabelled GTP[S] ⪢ GTP ⪢ guanosine 5′-[β,γ-imino]triphosphate ≥ guanosine 5′-O-[β-thio]diphosphate ≥ GDP > GMP = ATP (no effects at 1 mM), was absolutely necessary. While with unlabelled GTP[S] and GTP similar concentrations were required for control and fMet-Leu-Phe-stimulated release, about 50–100-fold higher concentrations of the other nucleotides were necessary for agonist-stimulated than for basal release of bound GTP[S]. The receptor action appeared to be catalytic, required Mg²⁺ and was pertussis toxin sensitive. The data indicate that binding of GTP[S] to HL-60 membrane G-proteins is reversible and that agonist-activated formyl peptide receptors can interact, either directly or indirectly, with GTP[S]-liganded G_i-proteins, resulting in release of bound GTP[S].