Knockout mouse models reveal the contributions of G protein subunits to complement C5a receptor-mediated chemotaxis

G protein-coupled receptor signaling is required for the navigation of immune cells along chemoattractant gradients. However, chemoattractant receptors may couple to more than one type of heterotrimeric G protein, each of which consists of a Gα, Gβ, and Gγ subunit, making it difficult to delineate the critical signaling pathways. Here, we used knockout mouse models and time-lapse microscopy to elucidate Gα and Gβ subunits contributing to complement C5a receptor-mediated chemotaxis. Complement C5a-mediated chemokinesis and chemotaxis were almost completely abolished in macrophages lacking Gnai2 (encoding Gα_i2), consistent with a reduced leukocyte recruitment previously observed in Gnai2^-/- mice, whereas cells lacking Gnai3 (Gα_i3) exhibited only a slight decrease in cell velocity. Surprisingly, C5a-induced Ca²⁺ transients and lamellipodial membrane spreading were persistent in Gnai2^-/- macrophages. Macrophages lacking both Gnaq (Gα_q) and Gna11 (Gα₁₁) or both Gna12 (Gα₁₂) and Gna13 (Gα₁₃) had essentially normal chemotaxis, Ca²⁺ signaling, and cell spreading, except Gna12/Gna13-deficient macrophages had increased cell velocity and elongated trailing ends. Moreover, Gnaq/Gna11-deficient cells did not respond to purinergic receptor P2Y₂ stimulation. Genetic deletion of Gna15 (Gα₁₅) virtually abolished C5a-induced Ca²⁺ transients, but chemotaxis and cell spreading were preserved. Homozygous Gnb1 (Gβ₁) deletion was lethal, but mice lacking Gnb2 (Gβ₂) were viable. Gnb2^-/- macrophages exhibited robust Ca²⁺ transients and cell spreading, albeit decreased cell velocity and impaired chemotaxis. In summary, complement C5a-mediated chemotaxis requires Gα_i2 and Gβ₂, but not Ca²⁺ signaling, and membrane protrusive activity is promoted by G proteins that deplete phosphatidylinositol 4,5-bisphosphate.