CXCL17 is an allosteric inhibitor of CXCR4 through a mechanism of action involving glycosaminoglycans

Abstract

CXCL17 is a chemokine principally expressed by mucosal tissues, where it facilitates chemotaxis of monocytes, dendritic cells, and macrophages and has antimicrobial properties. CXCL17 is also implicated in the pathology of inflammatory disorders and progression of several cancers, and its expression is increased during viral infections of the lung. However, the exact role of CXCL17 in health and disease requires further investigation, and there is a need for confirmed molecular targets mediating CXCL17 functional responses. Using a range of bioluminescence resonance energy transfer (BRET)–based assays, here we demonstrated that CXCL17 inhibited CXCR4-mediated signaling and ligand binding. Moreover, CXCL17 interacted with neuropillin-1, a VEGFR2 coreceptor. In addition, we found that CXCL17 only inhibited CXCR4 ligand binding in intact cells and demonstrated that this effect was mimicked by known glycosaminoglycan binders, surfen and protamine sulfate. Disruption of putative GAG binding domains in CXCL17 prevented CXCR4 binding. This indicated that CXCL17 inhibited CXCR4 by a mechanism of action that potentially required the presence of a glycosaminoglycan-containing accessory protein. Together, our results revealed that CXCL17 is an endogenous inhibitor of CXCR4 and represents the next step in our understanding of the function of CXCL17 and regulation of CXCR4 signaling.

Multiplex Detection of Fluorescent Chemokine Binding to CXC Chemokine Receptors by NanoBRET

“…Our obtained K i values of IT1t and AMD3100 for CXCR4 were approximately 1.5-and 30-fold higher, respectively, than previously observed in a NanoBRET-based CXCR4 binding assay versus CXCL12-AF647 [30]. However, a K i value of 10 nM has also been recently reported for AMD3100 in this NanoBRET-based binding assay using CXCL12-AF647 [31], which is only 3-fold lower than observed for the four AZDyelabeled CXCL12 analogs in this study. For burixafor, an IC 50 value of 10 nM in a CXCL12 competition binding assay on human CXCR4 was obtained [32]; this is likely in the same order range as our observed K i value of 25 nM.…”

ACKR5/GPR182 is a scavenger receptor for the atypical chemokine CXCL17, GPR15L and various endogenous peptides

“…Attempts to identify the receptor for CXCL17 remain so far unsuccessful. Recent reports, however, suggested that CXCL17 is a low-affinity agonist for the Mas-related GPR family member X2 (MRGPRX2), a low-selectivity receptor for peptides, and that it could act as an allosteric inhibitor of CXCR4 9,10 . This study establishes GPR182 as a high-affinity receptor also for two biologically active small non-chemokine proteins, GPR15L and CXCL17.…”