Characterization of interleukin-8 receptors in human neutrophil membranes: regulation by guanine nucleotides.
Interleukin-8 (IL-8) is a polymorphonuclear leukocyte (PMN) chemoattractant and activator which mediates its effects through specific cell-surface receptors. Indirect evidence indicates that guanine nucleotide regulatory proteins (G proteins) are necessary for transmembrane signaling. The present study characterizes IL-8 receptors in isolated PMN membrane fractions and shows direct regulation of these receptors by guanine nucleotides. The binding of [125I]IL-8 to subcellular fractions of PMNs showed specific binding in a low-density membrane fraction containing alkaline phosphatase, but not in primary or secondary granules. The binding of [125I]IL-8 was rapid and reversible. The equilibrium dissociation constant (Kd) of the receptor ranged from 5.0-12.4 nM and there were 1.58-5.90 . 10(10) receptors/mg protein. The dose-response curves for the competitive binding of three different forms of IL-8 to the receptor labeled by [125I]IL-8 corresponded with their ability to produce chemotaxis and granule exocytosis in PMNs. Treatment of membranes with the nonhydrolyzable analogs of GTP, GMP-PNP and GTP gamma S, inhibited the binding of [125I]IL-8. GMP-PNP decreased the affinity of the IL-8 receptor by approx. 2-fold without altering the total receptor number. These findings demonstrate that IL-8 receptors in PMN membranes are of high affinity and are convertible to a low-affinity state in the presence of guanine nucleotides, suggesting a direct role for G proteins in transmembrane signaling by this cytokine.
Barnett
M L
ML
Arthritis Unit, Harvard Medical School, Massachusetts General Hospital, Boston 02114.
Lamb
K A
KA
Costello
K M
KM
Pike
M C
MC
D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism; D006150:Guanine Nucleotides / Q000494:pharmacology*; D006801:Humans; D016209:Interleukin-8 / Q000378:metabolism; D007457:Iodine Radioisotopes; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D011971:Receptors, Immunologic / Q000737:chemistry / Q000187:drug effects*; D023062:Receptors, Interleukin-8A
D006150:Guanine Nucleotides; D016209:Interleukin-8; D007457:Iodine Radioisotopes; D011971:Receptors, Immunologic; D023062:Receptors, Interleukin-8A