Activated heterotrimeric G protein α_i subunits inhibit Rap-dependent cell adhesion and promote cell migration

Our recent work uncovered novel roles for activated Gα_i signaling in the regulation of neutrophil polarity and adhesion. Gα_iGTP-mediated enhancement of neutrophil polarization was dependent on inhibition of cAMP/PKA signaling, whereas reversal of Gβγ-stimulated adhesion was cAMP/PKA independent. To uncover the mechanism for Gα_i regulation of adhesion, we analyzed the effects of constitutively active Gα_i1(Q204L) expression on adhesion driven by constitutively active Rap1a(G12V) or its downstream effector Radil in neutrophil-like HL-60 cells, or in HT-1080 fibrosarcoma cells. In HT-1080 cells, Rap1a(G12V) or Radil cause an increase in cell spreading and adhesion to fibronectin, which are both reversed by Gα_i1(Q204L) but not WT Gα_i1 In contrast, Gα_i1(Q204L) did not reverse Rap1-GTP-interacting adaptor molecule (RIAM)-dependent increases in cell adhesion. This indicates that adhesion regulation by Gα_i-GTP occurs downstream of Rap1a and Radil, but is upstream of components such as integrins and talin that are regulated by both Radil and RIAM. HL-60 neutrophil-like cells expressing Rap1a(G12V) or Radil have an elongated phenotype because of enhanced uropod adhesion as they attempt to migrate on fibronectin. This elongated phenotype driven by Rap1a(G12V) or Radil is reversed by Gα_i1(Q204L), but not by WT Gα_i1 expression, suggesting that Gα_i-GTP also regulates adhesion in immune cells at the level of, or downstream of, Radil. These data identify a novel role of Gα_i-GTP in regulation of cell adhesion and migration. Cell migration involves cycles of adhesion and de-adhesion, and we propose that the dynamic spatiotemporal balance between Gβγ-promoted adhesion and Gα_i-GTP reversal of adhesion is important for this process.