Pertussis toxin produces differential inhibitory effects on basal, P2-purinergic, and chemotactic peptide-stimulated inositol phospholipid breakdown in HL-60 cells and HL-60 cell membranes.
P2-purinergic receptor agonists (UTP) and formylated peptide receptor agonists (FMLP) were found to be equally efficacious in eliciting rapid 6-7-fold increases in inositol polyphosphate accumulation in differentiated HL-60 granulocytes. The activation of this response by either agonist was substantially but incompletely inhibited in cells treated with pertussis toxin. Thus, in cells containing only 1-10% of the control level of non-ADP-ribosylated Gi-2/3, UTP induced rapid 2-fold increases in inositol polyphosphate accumulation whereas smaller 50% increases were observed in FMLP-stimulated cells. Washed membranes prepared from control and toxin-treated HL-60 cells were used to characterize this toxin-insensitive activation of phospholipase C further. The agonist-independent stimulation of phospholipase C by either millimolar Ca2+ or the nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) was only modestly attenuated by toxin treatment. There was a 70-80% decrease in the rate and extent of phospholipase C activity stimulated by GTP per se in the absence of receptor agonists. The rate and extent of FMLP-induced potentiation of GTP-dependent phospholipase C activity were also inhibited by greater than 80% in toxin-treated membranes. Conversely, the potency and efficacy characterizing UTP-induced potentiation of GTP-dependent phospholipase C activity were only modestly attenuated (less than 20% inhibition). The results indicate that P2-purinergic receptors (and perhaps other Ca2(+)-mobilizing receptors) activate both pertussis toxin-sensitive and toxin-insensitive pathways for phospholipase C regulation in phagocytic leukocytes.
Department of Pharmacology, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106.
D S
DS
Cowen
B
B
Baker
G R
GR
Dubyak
D002454:Cell Differentiation; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D006098:Granulocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology*; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism*; D007700:Kinetics; D015473:Leukemia, Promyelocytic, Acute; D008563:Membrane Lipids / Q000378:metabolism*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D037342:Pertussis Toxin; D013873:Thionucleotides / Q000494:pharmacology*; D010738:Type C Phospholipases / Q000378:metabolism; D014544:Uridine Triphosphate / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D007295:Inositol Phosphates; D008563:Membrane Lipids; D013873:Thionucleotides; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009240:N-Formylmethionine Leucyl-Phenylalanine; D006160:Guanosine Triphosphate; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D014544:Uridine Triphosphate