The LTB4-BLT1 axis regulates the polarized trafficking of chemoattractant GPCRs during neutrophil chemotaxis

Neutrophils sense and respond to diverse chemotactic cues using G-protein coupled receptors (GPCRs). However, the precise trafficking dynamics of chemoattractant GPCRs during neutrophil activation and chemotaxis remain unclear. Here, using small molecule inhibitors and CRISPR-based knockouts, we establish that two primary chemoattractant GPCRs - formyl peptide receptor 1 (FPR1) and C5a receptor 1 (C5aR) - internalize in a CDC42-Actin-dependent manner. Using live-cell imaging, we demonstrate that upon stimulation FPR1 rapidly clusters, re-distributes along the plasma membrane to the trailing edge, where it internalizes and is directionally trafficked towards the front of migrating primary human neutrophils. In contrast to FPR1 and C5aR, the leukotriene B4 (LTB4) receptor (BLT1), which relays LTB4 signals in response to primary chemoattractants during neutrophil chemotaxis, fails to internalize upon physiological stimulation with LTB4, fMLF or C5a. Importantly, we report that blocking the LTB4-BLT1 axis or downstream Myosin activation enhances the internalization of FPR1 and C5aR, thus reducing downstream signaling and impairing chemotaxis to primary chemoattractants. The polarized trafficking of chemoattractant GPCRs and its regulation by the BLT1-mediated Myosin activation therefore drives persistent chemotactic signaling in neutrophils.