CXCR1 and CXCR2 display receptor bias for shared chemokine agonists.

G protein-coupled receptors (GPCRs) mediate diverse signaling outputs through their proximal transducers: G proteins, GPCR kinases, and β-arrestins. Although ligand bias at chemokine receptors (CKRs), where ligands for the same receptor display distinct signaling patterns, is well recognized, receptor bias, where the same agonist at different receptors yields distinct transducer engagement, remains poorly understood. We compared endogenous chemokine ligands (CXCL1, CXCL5, CXCL7, CXCL8) at the highly homologous CXCR1 and CXCR2 using biosensor assays to measure Gαi activation, β-arrestin1/2 recruitment, GPCR kinase 2/3/5/6 translocation, and receptor internalization. Our data reveal qualitatively different signaling patterns, most notably where CXCL1 acts as a G protein-biased partial agonist at CXCR1 but as a balanced full agonist at CXCR2. These signaling differences correlate with receptor internalization but not subcellular ERK activation patterns measured using compartment-specific biosensors. Collectively, our findings demonstrate receptor bias in CKR signaling, transducer activation, and compartmentalized kinase activation in translating chemokine identity into discrete functional outcomes. SIGNIFICANCE STATEMENT: Ligand bias, where different ligands for the same receptor display different signaling patterns, is now well appreciated. However, there are only a few examples of receptor bias, where the same agonist generates distinct signaling profiles at different receptors. This study used biosensors and compartmental ERK biosensors to show that shared ligands of CXCR1 and CXCR2 differentially engage G proteins, β-arrestins, and G protein-coupled receptor kinases, revealing receptor bias in CXCL1-mediated signaling that extends beyond prior characterizations.