Differential Activation and Regulation of CXCR1 and CXCR2 by CXCL8 Monomer and Dimer

Abstract

CXCL8 (also known as IL-8) activates CXCR1 and CXCR2 to mediate neutrophil recruitment and trigger cytotoxic effect at sites of infection. Under physiological conditions, CXCL8 could exist as monomers, dimers, or a mixture of monomers and dimers. Therefore, both forms of CXCL8 could interact with CXCR1 and CXCR2 with different affinities and potencies to mediate different cellular responses. In the present study, we have used a “trapped” nonassociating monomer (L25NMe) and a nondissociating dimer (R26C) to investigate their activities for human neutrophils that express both receptors and for RBL-2H3 cells stably expressing either CXCR1(RBL-CXCR1) or CXCR2 (RBL-CXCR2). The monomer was more active than the dimer for activities such as intracellular Ca2+ mobilization, phosphoinositide hydrolysis, chemotaxis. and exocytosis. Receptor regulation, however, is distinct for each receptor. The rate of monomer-mediated regulation of CXCR1 is greater for activities such as phosphorylation, desensitization, β-arrestin translocation, and internalization. In contrast, for CXCR2, both monomeric and dimeric CXCL8 mediate these activities to a similar extent. Interestingly, receptor-mediated signal-regulated kinase (ERK) phosphorylation in response to all three CXCL8 variants was more sustained for CXCR2 relative to CXCR1. Taken together, the results indicate that the CXCL8 monomer and dimer differentially activate and regulate CXCR1 and CXCR2 receptors. These distinct properties of the ligand and the receptors play a critical role in orchestrating neutrophil recruitment and eliciting cytotoxic activity during an inflammatory response.

The dynamics of interleukin‐8 and its interaction with human CXC receptor I peptide

“…Site I comprises the initial interaction between the CXCL8 N-loop residues and the CXCR1 receptor N-terminal residues, while site II involves the CXCL8 N-terminal (ELR motif) and the receptor distant extracellular region. [5][6][7]10,[14][15][16] However, a recent biophysical study performed in lipid bilayers using human full-length CXCR1 and CXCL8 did not fully support such findings, suggesting that the binding process between CXCL8 and human CXCR1 appears to be mainly associated with the N-terminal region of the receptor. 17 Furthermore, using biophysical studies, Ravindran et al 7 proposed that upon receptor engagement, wild-type CXCL8 dissociates to form a CXCL8 monomer/receptor complex.…”

“…7,9 As CXCL8 is secreted at high concentrations from injured or cancerous tissues, its local concentration can vary significantly, leading to the existence of both the monomeric and dimeric forms at different locations and over time. 10,11 This differential distribution of the CXCL8 dimeric and monomeric forms may suggest an important role for each of them in the functional activity of this chemokine. For example, one may hypothesise that the formation of CXCL8 dimers at high concentrations may diminish the binding affinity of CXCL8 to its receptors, and thereby serve as a control mechanism to downregulate signaling.…”

“…1,2 CXC chemokines are small proteins (8)(9)(10)(11)(12) kDa) that contain a conserved CXC residue motif (C-cysteine, X-any other residue) proximal to the N-terminal region of the protein. 3 Solution and solid-state structures of several human CXC chemokines have been solved, revealing a common tertiary fold for all members of the family consisting of an unstructured N-terminal loop, antiparellel b-strands and a C-terminal a-helix.…”

The Multifaceted Roles Neutrophils Play in the Tumor Microenvironment

“…CXCL1 was shown to be involved in a paracrine network mediating both metastatic progression and chemoresistance [250]. IL-8 is another chemokine expressed in variety of tumors and is associated with neutrophil recruitment and activation [251]. For example, IL-8 secreted by human fibrosarcoma and prostate carcinoma cells promoted the infiltration of MMP-9 + neutrophils [252] and melanoma secreted IL-8 was shown to increase CD11b/CD18 expression on neutrophils, an indication for their activation [253,254].…”

Interaction of Monomeric Interleukin-8 with CXCR1 Mapped by Proton-Detected Fast MAS Solid-State NMR

“…Most of the differences are in the N-and C-terminals, which bind chemokines and G-proteins, respectively. Both receptors have high affinity for IL-8, with a low nanomolar dissociation constant (22,23). Finding that two similar but distinct receptors bind the same chemokine, and that CXCR1 almost exclusively binds to IL-8 whereas CXCR2 also binds to several other chemokines, provided an early indication of the complexity of the chemokine defense system.…”