CXCR1 and CXCR2 Activation and Regulation

Abstract

CXCL8 (interleukin-8) interacts with two receptors, CXCR1and CXCR2, to activate leukocytes. Upon activation, CXCR2 internalizes very rapidly relative to CXCR1 (ϳ90% versus ϳ10% after 5 min). The C termini of the receptors have been shown to be necessary for internalization but are not sufficient to explain the distinct kinetics of down-regulation. To determine the structural determinant(s) that modulate receptor internalization, various chimeric and point mutant receptors were generated by progressively exchanging specific domains or amino acids between CXCR1 and CXCR2. The receptors were stably expressed in rat basophilic leukemia 2H3 cells and characterized for receptor binding, intracellular Ca Chemokines are a family of structurally related peptides that regulate inflammation through cell-surface G protein-coupled receptors on leukocytes. These peptides mediate diverse biological and biochemical activities, including endothelial adhesion, directed migration, and activation of cytotoxic activities such as the respiratory burst and exocytosis (1, 2). Chemokines have been classified into four families (C, CC, CXC, and CX3C) based on the number and positions of the N-terminally conserved cysteine residues. Most chemokines activate more than one chemokine receptor, and many chemokine receptors are activated by multiple chemokines (3). CXCL8 activates two receptors, CXCR1 and CXCR2. CXCR1 is specific for CXCL8, whereas CXCR2 also interacts with CXCL1, CXCL2, CXCL3, CXCL5, CXCL6, and CXCL7 (4). Upon activation, both receptors couple to pertussis toxin-sensitive G protein to mediate phosphoinositide (PI) 2 hydrolysis, intracellular Ca 2ϩ mobilization, chemotaxis, and exocytosis. CXCR1 (but not CXCR2) activates phospholipase D and mediates respiratory burst, suggesting that the two receptors may play different physiological roles (5, 6). Like many G protein-coupled receptors, both receptors become phosphorylated, desensitized, and internalized upon exposure to CXCL8. Over 95% of CXCR2 internalizes in the first 2-5 min of activation compared with ϳ10% of CXCR1 (7-10). CXCR2 also recovers more slowly (ϳ35% after 90 min) to the cell surface than does CXCR1 (ϳ100% after 90 min) upon removal of CXCL8 (7,8,(11)(12)(13)(14). This difference in receptor trafficking appears to be an important factor in the distinct ability of CXCR1 and CXCR2 to mediate leukocyte activation and regulation in response to CXCL8 (9, 10).To date, the molecular basis for the differential regulation of the CXCL8 receptors remains unclear. CXCR1 and CXCR2 are highly homologous (77%) (15, 16). The most divergent regions are the N terminus, the fourth transmembrane domain (TMD), the second extracellular loop (ECL), and the C terminus (15-18). Although both receptors internalize via a phosphorylation/ arrestin/dynamin-dependent mechanism,

Chemokine Signaling and the Regulation of Bidirectional Leukocyte Migration in Interstitial Tissues

“…One potential mechanism for this effect is the downregulation or desensitization of the receptor and/or the contribution of specific G-protein coupled receptor kinases (GRKs) (Bennett et al, 2011; Ribas et al, 2007). There is evidence that ligation of human CXCR2 leads to a more sustained response than CXCR1, consistent with our finding that random motility through Cxcr2 is more sustained than the directed response through Cxcr1 at the zebrafish wound (Nasser et al, 2007; Richardson et al, 2003; Rose et al, 2004). Additionally, it has been reported that human CXCR1 and CXR2 bind to different GRKs (Raghuwanshi et al, 2012), which could also mediate different neutrophil responses after activation.…”

Novel Human Cytomegalovirus Viral Chemokines, vCXCL-1s, Display Functional Selectivity for Neutrophil Signaling and Function

“…These data point to different thresholds for the different neutrophil functions. Calcium flux and integrin expression have low thresholds where any degree of stimulation will activate them (69). Other functional outcomes (i.e., migration, signaling, or CCL22 expression) are more complex.…”

New paradigms in chemokine receptor signal transduction: Moving beyond the two-site model

“…2A, B) [83–86]. Similar pharmacological and structural studies expanded these principles to other chemokine-CKR pairs, including CCR1 [81, 87, 88], CCR2 [18, 89, 90], CCR3 [91, 92], CCR5 [18, 88, 93–96], CXCR1 [97–99], CXCR2 [97, 98], CXCR3 [100], CXCR4 [82, 101], CX3CR1 [102]. …”