ACKR3 agonism induces heterodimerization with chemokine receptor CXCR4 and attenuates platelet function

Background Platelet receptors ACKR3 and CXCR4 play a crucial role in a variety of cardio-vascular diseases. Like most chemokine receptors, CXCR4 is a G protein coupled receptor that induces platelet activation. In contrast, the atypical chemokine receptor 3 (ACKR3) lacks the ability to activate heterotrimeric G proteins and its activation leads to platelet inhibition and attenuates thrombus formation. In nucleated cells, heterodimerization of ACKR3 with CXCR4 regulates CXCL12-dependent signaling. The aim of our study was to investigate the formation of ACKR3/CXCR4 heterodimers in platelets and the subsequent consequences for platelet function. Methods and results Using a microscopy proximity ligation assay (PLA, Duolink ® ) to screen for CXCR4/ACKR3 heterodimerization inducing compounds, we found that ACKR3 agonism but not conventional platelet agonists or endogen ligands lead to heterodimer formation. To further characterize the formation of ACKR3/CXCR4 heterodimers, we studied the CXCL12-dependent platelet activation via CXCR4. Both, CXCL12-dependent platelet aggregation and collagen-dependent ex vivo thrombus formation were significantly downregulated by ACKR3 agonism. Moreover, platelet intracellular calcium and Akt signaling were increased by CXCL12 and again suppressed by ACKR3-specific agonists. Previously, CXCL12 was shown to decrease platelet cAMP levels via CXCR4. Treatment with a specific ACKR3 agonist counteracted this CXCL12/CXCR4-dependent cAMP decrease. Conclusion Our results reveal that the formation of platelet ACKR3/CXCR4 heterodimers is dependent on ACKR3 rather than CXCR4. Furthermore, ACKR3 agonism induced heterodimerization is associated with mitigating CXCL12/CXCR4-dependent platelet activation possibly by modulating CXCR4-dependent G protein signaling. Our results indicate possible ACKR3 agonist functions and reinforce the potential therapeutic applications of ACKR3 agonists. Abstract Figure Graphical abstract: CXCR4/ACKR3 heterodimerization in platelets.