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

Esther van den Bos, Benjamin Ambrosy, Markus Horsthemke, Stefan Walbaum, Anne C. Bachg, Nina Wettschureck, Giulio Innamorati, Thomas M. Wilkie, Peter J. Hanley

Research output: Contribution to journalArticle

Abstract

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 Gαi2), consistent with a reduced leukocyte recruitment previously observed in Gnai2-/- mice, whereas cells lacking Gnai3 (Gαi3) 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 (Gαq) 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 Gαi2 and Gβ2, but not Ca2+ signaling, and membrane protrusive activity is promoted by G proteins that deplete phosphatidylinositol 4,5-bisphosphate.

Original languageEnglish (US)
Pages (from-to)7726-7742
Number of pages17
JournalJournal of Biological Chemistry
Volume295
Issue number22
DOIs
StatePublished - May 28 2020

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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  • Cite this

    van den Bos, E., Ambrosy, B., Horsthemke, M., Walbaum, S., Bachg, A. C., Wettschureck, N., Innamorati, G., Wilkie, T. M., & Hanley, P. J. (2020). Knockout mouse models reveal the contributions of G protein subunits to complement C5a receptor–mediated chemotaxis. Journal of Biological Chemistry, 295(22), 7726-7742. https://doi.org/10.1074/jbc.RA119.011984