Servicenavigation

Project Details

Back

Projekt Print View

H-Ras, a new regulator of bone integrity - the mechanistic base of osteoporosis triggered by a constitutive active H-Ras GTPase

Subject Area Orthopaedics, Traumatology, Reconstructive Surgery
Term from 2019 to 2023
Project identifier Deutsche Forschungsgemeinschaft (DFG) - Project number 408077919
 
Final Report Year 2024

Final Report Abstract

The protooncogene Hras is a ubiquitously expressed central regulator of fundamental biological processes such as proliferation, differentiation, apoptosis and senescence. Hras receives a wide diversity of extra-/intracellular signals and by integrating them into defined signaling pathways provides an appropriate cellular adaptive response. Germline mutations in the HRAS gene are the underlying cause of the Costello syndrome (CS), a monogenic rare disease with premature aging-like features. Among multiple pathologies, osteoporosis in a major pathophenotype in adult patients. Most CS mutations are G12 substitutions that lock Hras in a constitutively active state. To understand the underlying mechanisms of altered bone metabolism CS patients we investigated a CS Hras G12V knock­in mause model. We identified that a constitutive activation of Hras was accompanied with bone lass due to an increased number of osteoclasts. Focusing further an in vitro analyses of osteoclasts, we observed that Hras G12V mutations (i) did not trigger senescence in osteoclasts, irrespective to the age of mause used as donor for bone marrow cells, as cells were able to proliferate and differentiate to higher rates than wild type controls, (ii) did not impair osteoclast actin cytoskeleton and acidification, but (iii) increased energy metabolism. Furthermore, in vitro rescue experiments of CS osteoclastogenesis indicated that MAPK inhibition using MEK inhibitor reduced number and size of osteoclasts, while Pl3K inhibitor rescued only osteoclast size. We also studied the impact of CS Hras G12V (strong CS pathophenotype) and G12S (most frequent and milder pathophenotype) activated mutations an osteoblast differentiation in MC3T3-E1 murine preosteoblasts expressing these mutant Hras proteins in an inducible manner. In this in vitro set up, Hras mutations inhibited the expression of osteoblast differentiation master regulator Runx2, osteoblast differentiation and mineralization, with G12V showing strenger effects than G12S mutant. Rescue experiments indicated that only farnesyltransferase inhibitor Tipifarnib, that blocks Hras membrane localization, was able to rescue differentiation and mineralization in G12S-expressing cells, while in G12V-expressing cells was able to rescue differentiation and not mineralization. An intriguing finding is that Hras impaired differentiation and mineralization correlated with an increased the expression of Opn and this was rescued by Tipifarnib. Additionally, in two spinoff projects we revealed two novel mechanisms that regulate (H)RAS and EGFR-(H)RAS-RAF signaling module, where the GR (NR3C1) and ACE2, respectively, acted as negative regulators of (H)RAS activation and its downstream signaling. These mechanisms that are relevant to RAS cancers and SARS-CoV-2 infection, respectively, will be further studied in HRAS­controlled mechanisms in bone homeostasis, as both GR and ACE2 are also associated with bone remodeling and osteoporosis. Altogether, in this project we were able to identify cellular bases of osteoporosis in CS, we provided evidences that Hras is a critical regulator of bone remodeling though its implications in osteoclastogenesis and osteoblast differentiation and mineralization, and identified two novel inhibitory mechanisms an RAS activation driven by GR and ACE2, respectively, both mechanisms being extremely relevant for various human diseases.

Publications

 
 

Additional Information

© 2026 DFG Disclaimer / Copyright / Privacy Policy

Textvergrößerung und Kontrastanpassung