Noncanonical G-protein-dependent modulation of osteoclast differentiation and bone resorption mediated by Pasteurella multocida toxin.
Pasteurella multocida toxin (PMT) induces atrophic rhinitis in animals, which is characterized by a degradation of nasal turbinate bones, indicating an effect of the toxin on bone cells such as osteoblasts and osteoclasts. The underlying molecular mechanism of PMT was defined as a persistent activation of heterotrimeric G proteins by deamidation of a specific glutamine residue. Here, we show that PMT acts directly on osteoclast precursor cells such as bone marrow-derived CD14(+) monocytes and RAW246.7 cells to induce osteoclastogenesis as measured by expression of osteoclast-specific markers such as tartrate-resistant acid phosphatase and bone resorption activity. Treatment performed solely with PMT stimulates osteoclast differentiation, showing a receptor activator of nuclear factor-κB ligand (RANKL)-independent action of the toxin. The underlying signal transduction pathway was defined as activation of the heterotrimeric G proteins Gαq/11 leading to the transactivation of Ras and the mitogen-activated protein kinase pathway. Gαq/11 transactivates Ras via its effector phospholipase Cβ-protein kinase C (PKC) involving proline-rich tyrosine kinase 2 (Pyk2). PMT-induced activation of the mitogen-activated protein kinase pathway results in stimulation of the osteoclastogenic transcription factors AP-1, NF-κB, and NFATc1. In addition, Ca(2+)-dependent calcineurin activation of NFAT is crucial for PMT-induced osteoclastogenesis. The data not only elucidate a rationale for PMT-dependent bone loss during atrophic rhinitis but also highlight a noncanonical, G-protein-dependent pathway toward bone resorption that is distinct from the RANKL-RANK pathway but mimics it. We define heterotrimeric G proteins as as-yet-underestimated entities/players in the maturation of osteoclasts which might be of pharmacological relevance.


OBJECTIVE
Pasteurella multocida toxin (PMT) induces degradation of nasal turbinate bones, leading to the syndrome of atrophic rhinitis. Recently, the molecular mechanism and substrate specificity of PMT were identified. The toxin activates heterotrimeric G proteins by a covalent modification. However, the mechanism by which PMT induces bone degradation is poorly understood. Our report demonstrates a direct effect of PMT on osteoclast precursor cells, leading to maturation of bone-degrading osteoclasts. Interestingly, PMT stimulates osteoclastogenesis independently of the cytokine RANKL, which is a key factor in induction of osteoclast differentiation. This implicates a noncanonical osteoclastogenic signaling pathway induced by PMT. The elucidated Gαq/11-dependent osteoclastogenic signal transduction pathway ends in osteoclastogenic NFAT signaling. The noncanonical, heterotrimeric G protein-dependent osteoclast differentiation process may be of pharmacological relevance, as members of this pathway are highly druggable. In particular, modulation of G protein-coupled receptor activity in osteoclast progenitors by small molecules might be of specific interest.
Department I, Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, DepAlbert-Ludwigs-Universität Freiburg, Freiburg, Germany.
Julia
J
Strack
Department I, Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, DepAlbert-Ludwigs-Universität Freiburg, Freiburg, Germany.
Hannah
H
Heni
Department II, Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, DepAlbert-Ludwigs-Universität Freiburg, Freiburg, Germany.
Ralf
R
Gilsbach
Department II, Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, DepAlbert-Ludwigs-Universität Freiburg, Freiburg, Germany.
Lutz
L
Hein
Joachim.Orth@pharmakol.uni-freiburg.de Klaus.Aktories@pharmakol.uni-freiburg.de.
Klaus
K
Aktories
Department I, Institut für Experimentelle und Klinische Pharmakologie und Toxikologie, DepAlbert-Ludwigs-Universität Freiburg, Freiburg, Germany Joachim.Orth@pharmakol.uni-freiburg.de Klaus.Aktories@pharmakol.uni-freiburg.de.
Joachim H C
JH
Orth
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D001862:Bone Resorption; D002460:Cell Line; D019204:GTP-Binding Proteins / Q000378:metabolism*; D054884:Host-Pathogen Interactions; D008264:Macrophages / Q000187:drug effects; D051379:Mice; D010010:Osteoclasts / Q000187:drug effects / Q000378:metabolism*; D016979:Pasteurella multocida / Q000502:physiology*; D012222:Rhinitis, Atrophic / Q000473:pathology*; D015398:Signal Transduction
D001426:Bacterial Proteins; D001427:Bacterial Toxins; C062258:Pasteurella multocida toxin; D019204:GTP-Binding Proteins