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-8-dependent receptor internalization by monitoring the density of surface 125I-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 the 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-8-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.

[1]  J. Benovic,et al.  Desensitization and Internalization of the m2 Muscarinic Acetylcholine Receptor Are Directed by Independent Mechanisms * , 1995, The Journal of Biological Chemistry.

[2]  A. Chuntharapai,et al.  Regulation of the expression of IL-8 receptor A/B by IL-8: possible functions of each receptor. , 1995, Journal of immunology.

[3]  D. Kelvin,et al.  Interleukin-8 Receptor , 1995, The Journal of Biological Chemistry.

[4]  S. Liggett,et al.  Four Consecutive Serines in the Third Intracellular Loop Are the Sites for β-Adrenergic Receptor Kinase-mediated Phosphorylation and Desensitization of the α2A-Adrenergic Receptor (*) , 1995, The Journal of Biological Chemistry.

[5]  R. Nissenson,et al.  The Cytoplasmic Tail of the G-protein-coupled Receptor for Parathyroid Hormone and Parathyroid Hormone-related Protein Contains Positive and Negative Signals for Endocytosis (*) , 1995, The Journal of Biological Chemistry.

[6]  K. Baker,et al.  Stable expression of a truncated AT1A receptor in CHO-K1 cells. The carboxyl-terminal region directs agonist-induced internalization but not receptor signaling or desensitization. , 1995, The Journal of biological chemistry.

[7]  D. Diviani,et al.  Truncation of the receptor carboxyl terminus impairs agonist-dependent phosphorylation and desensitization of the alpha 1B-adrenergic receptor. , 1994, The Journal of biological chemistry.

[8]  J. Navarro,et al.  The N terminus of interleukin-8 (IL-8) receptor confers high affinity binding to human IL-8. , 1994, The Journal of biological chemistry.

[9]  J. Navarro,et al.  Molecular characterization of a novel rabbit interleukin-8 receptor isotype. , 1994, The Journal of biological chemistry.

[10]  M. Caron,et al.  A highly conserved tyrosine residue in G protein-coupled receptors is required for agonist-mediated beta 2-adrenergic receptor sequestration. , 1994, The Journal of biological chemistry.

[11]  J. Krause,et al.  Functional nonequivalence of structurally homologous domains of neurokinin-1 and neurokinin-2 type tachykinin receptors. , 1993, The Journal of biological chemistry.

[12]  S. Liggett,et al.  Structural basis for receptor subtype-specific regulation revealed by a chimeric beta 3/beta 2-adrenergic receptor. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[13]  T. Vanden Bos,et al.  Importance of the amino terminus of the interleukin-8 receptor in ligand interactions. , 1993, The Journal of biological chemistry.

[14]  R. Mark,et al.  Amino terminus of the interleukin-8 receptor is a major determinant of receptor subtype specificity. , 1992, The Journal of biological chemistry.

[15]  W. Wood,et al.  Characterization of two high affinity human interleukin-8 receptors. , 1992, The Journal of biological chemistry.

[16]  W I Wood,et al.  Structure and functional expression of a human interleukin-8 receptor. , 1991, Science.

[17]  L. Taylor,et al.  The interleukin-8 receptor is encoded by a neutrophil-specific cDNA clone, F3R. , 1991, The Journal of biological chemistry.

[18]  D. Sibley,et al.  Regulation of transmembrane signaling by receptor phosphorylation , 1987, Cell.

[19]  R. Tsien,et al.  A new generation of Ca2+ indicators with greatly improved fluorescence properties. , 1985, The Journal of biological chemistry.

[20]  P. Hargrave,et al.  Localization of binding sites for carboxyl terminal specific anti-rhodopsin monoclonal antibodies using synthetic peptides. , 1984, Biochemistry.

[21]  T. Hunter,et al.  The transforming protein of Moloney murine sarcoma virus is a soluble cytoplasmic protein , 1983, Cell.

[22]  K. Matsushima,et al.  Properties of the novel proinflammatory supergene "intercrine" cytokine family. , 1991, Annual review of immunology.

[23]  J. Thorner,et al.  Model systems for the study of seven-transmembrane-segment receptors. , 1991, Annual review of biochemistry.