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 (encoding Gα), consistent with a reduced leukocyte recruitment previously observed in mice, whereas cells lacking (Gα) exhibited only a slight decrease in cell velocity. Surprisingly, C5a-induced Ca transients and lamellipodial membrane spreading were persistent in macrophages. Macrophages lacking both (Gα) and (Gα) or both (Gα) and (Gα) had essentially normal chemotaxis, Ca signaling, and cell spreading, except /-deficient macrophages had increased cell velocity and elongated trailing ends. Moreover, /-deficient cells did not respond to purinergic receptor P2Y stimulation. Genetic deletion of (Gα) virtually abolished C5a-induced Ca transients, but chemotaxis and cell spreading were preserved. Homozygous (Gβ) deletion was lethal, but mice lacking (Gβ) were viable. macrophages exhibited robust Ca transients and cell spreading, albeit decreased cell velocity and impaired chemotaxis. In summary, complement C5a-mediated chemotaxis requires Gα and Gβ, but not Ca signaling, and membrane protrusive activity is promoted by G proteins that deplete phosphatidylinositol 4,5-bisphosphate.