Canonical and Noncanonical G-Protein Signaling Helps Coordinate Actin Dynamics To Promote Macrophage Phagocytosis of Zymosan

Abstract

Both chemotaxis and phagocytosis depend upon actin-driven cell protrusions and cell membrane remodeling. While chemoattractant receptors rely upon canonical G-protein signaling to activate downstream effectors, whether such signaling pathways affect phagocytosis is contentious. Here, we report that G␣ i nucleotide exchange and signaling helps macrophages coordinate the recognition, capture, and engulfment of zymosan bioparticles. We show that zymosan exposure recruits F-actin, G␣ i proteins, and Elmo1 to phagocytic cups and early phagosomes. Zymosan triggered an increase in intracellular Ca 2؉ that was partially sensitive to G␣ i nucleotide exchange inhibition and expression of GTP-bound G␣ i recruited Elmo1 to the plasma membrane. Reducing GDP-G␣ i nucleotide exchange, decreasing G␣ i expression, pharmacologically interrupting G␤␥ signaling, or reducing Elmo1 expression all impaired phagocytosis, while favoring the duration that G␣ i remained GTP bound promoted it. Our studies demonstrate that targeting heterotrimeric G-protein signaling offers opportunities to enhance or retard macrophage engulfment of phagocytic targets such as zymosan.

Proteogenomic Characterization of Human Early-Onset Gastric Cancer

“…The subtype 2 network model showed that phagosome activity in antigen-presenting cells and TCR signaling in T cells could be modulated by mutations in CXCR5 and its downstream G-proteins (GNAI3, GNB3-5, and GNG4) via their associations (green edges in Figure 4D) with phosphorylated proteins in these processes (NCF2/4 and CYBA/B in phagosomes and CD8A, CD247, LCK, and PLCG in T cell signaling). In support of this finding, the association of G-proteins with phagocytosis (Huang et al, 2014) was previously reported to modulate inflammatory responses. The subtype 4 network model ( Figure 4E) showed that the activity of actin cytoskeleton, which is primarily regulated by RHOA and RAC1 signaling, could be modulated by two mutated genes, PLK4 and NEK3, in its upstream pathways via their associations with corresponding protein were selected as the signatures for subtypes 2 (light blue) and 4 (dark green).…”

Gαi2 Signaling Regulates Inflammasome Priming and Cytokine Production by Biasing Macrophage Phenotype Determination

“…Importantly, these types of macrophages have significant developmental differences at the transcriptional level (65), and thus are likely convergently affected by signaling inputs during development. Despite inconsistencies with Li et al (63), the proinflammatory M1 nature of G184S KI BMDMs revealed in this study are consistent with our previous findings that unprimed G184S KI macrophages have enhanced phagocytic capacity (29,66). Our data are also consistent with a study on RGS10deficient macrophages (38).…”

“…Ga i signaling has previously been shown to modulate macrophage chemotaxis, phagocytosis, and activation (6,28,29). Although several Ga i -coupled GPCRs show opposing isoform/ligand-specific effects on macrophage polarization, the role of Ga i in macrophage polarization is unknown.…”

SWAP70 Organizes the Actin Cytoskeleton and Is Essential for Phagocytosis

“…We also often observed concentric ring-like structures of SWAP70 (Figure 1G; Movies S5 and S6). This observation raised the possibility that SWAP70 would align with the F-actin filaments previously described to form around the phagocytic cup following uptake of non-opsonized zymosan (Goodridge et al., 2012, Huang et al., 2014, Liebl and Griffiths, 2009). We performed multi-color STED microscopy to address this possibility and observed perfect alignment of the F-actin filaments with SWAP70 (Figures 1H–1J; Movie S7).…”