Abstract
Desensitization of N-formyl peptide chemoattractant receptors (FPR)
in human neutrophils results in association of these receptors to
the membrane skeleton. This is thought to be the critical event in
the lateral segregation of receptors and guanyl nucleotide-binding
proteins (G proteins) within the plane of the plasma membrane resulting
in an interruption of the signaling cascade. In this study we probed
the interaction of FPR with G protein in human neutrophils that were
desensitized to various degrees. Human neutrophils were desensitized
using the photoreactive agonist N-formyl-met-leu-phe-lys-N epsilon-125I2(p-azidosalicylamino)ethyl-1,3'-
dithiopropionate (fMLFK-125IASD). The interaction of FPR with protein
was studied via a reconstitution assay and subsequent analysis of
FPR-G protein complexes in sucrose density gradients. FPR-G protein
complexes were reconstituted with solubilized FPR from partially
and fully desensitized neutrophils with increasing concentrations
of Gi purified from bovine brain. The respective EC50 values for
reconstitution were similar to that determined for FPR from unstimulated
neutrophils (Bommakanti RK et al., J Biol Chem 267: 7576-7581, 1992).
We conclude, therefore, that the affinity of the interaction of FPR
with G protein is not affected by desensitization, consistent with
the model of lateral segregation of FPR and G protein as a mechanism
of desensitization.
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