Role of the C terminus of the interleukin 8 receptor in signal transduction and internalization

Interleukin 8 (IL-8) is a potent neutrophil chemoattractant and activator. Two IL-8 receptor subtypes, A and B, are expressed in neutrophils. In this work, we analyzed the role of the C terminus domain of the IL-8 receptor on the signal transduction and receptor internalization mechanisms, The IL-8 receptor A was tagged with an epitope corresponding to the monoclonal antibody 1D4 to monitor the, localization of the IL-8 receptor, We demonstrated IL-S-dependent. receptor internalization by monitoring the density of surface I-125-labeled IL-8 binding sites and by immunofluorescence microscopy, Truncation of the last 27 amino acids of the IL-8 receptor A severely impaired the IL-8-induced internalization of the receptor. Of importance was tire observation that binding of IL-8 to receptors A and B triggered a dramatically faster rate of internalization of receptor B than receptor A, suggesting that the heterologous C termini among receptor subtypes modulate the rate of internalization of IL-8 receptors. However, substitution of the C terminus of the receptor subtype A for the C terminus of receptor B reduced the internalization rate of receptor A. Furthermore, we found that the rate of internalization of IL-8 receptor B triggered by IL-8 was faster than the one induced by the IL-S-related peptide, melanoma growth stimulatory activity, Studies with human neutrophils pretreated with 100 nM IL-8 for 5 min revealed a positive and a negative calcium response mediated by receptors A and B, respectively. In contrast, neutrophils pretreated with melanoma growth stimulatory activity showed positive calcium responses to both receptors A and B. These data suggest that the neutrophil responses mediated by IL-8 are modulated by the rate of internalization of receptors.