Studies of the functions of TGFBI in bone homeostasis and cancer development
Final Report Abstract
Goals of the funded work were to investigate the Role of Fos and TGFBI in i) bone in respect to bone homeostasis and tumorigenesis with a focus on OS and in ii) RMS development. Here the main focus was put onto part i) as discussed above. Using different mouse models (loss of function and gain of function) we were able to show that the putative Fos target TGFBI has an impact on age mediated bone loss in mice while at earlier time points we did not find a significant influence of TGFBI on bone homeostasis confirming the published results of others. We are currently analyzing TGFBI∆/∆; POSTN∆/∆ mice and preliminary results suggest an interaction of these ECM proteins in regulating bone homeostasis which is an exciting finding and will be a major focus of future work. Furthermore we were able to show that TGFBI has a significant influence on tumorigenesis in a Fos-dependent model of OS since deletion of TGFBI resulted in a significant increase of OS in H2K-Fos-LTR mice. Here future work will address the potential mechanisms of how TGFBI impacts OS. We also obtained preliminary results that suggest a role of TGFBI in cancer cell metastasis to bone. Here, it will be worthwhile to extend the analysis to TGFBI∆/∆; POSTN∆/∆ mice to account for potential compensation of these family members. Since deletion of TGFBI has been linked to genomic instability we addressed the question whether altered expression of Fos influences genomic instability in osteoblasts. Here we made the unexpected observation that Fos overexpression does not cause replication stress/ DNA damage which is a known cause of genomic instability despite increased cell cycle progression. Furthermore, increased Fos expression is able to protect cells against agents like UCN-01 that lead to replication stress and DNA damage. Using RNAseq we were able to show that Fos regulates Chk1 levels and thus the DNA damage response. Whether this regulation is direct is still under investigation. These findings are especially intriguing since increased Chk1 levels are known to play a pro-tumorigenic role in Myc-associated malignancies balancing replication stress and preventing cell death. Using Ras/E1a transduced cells, we could show that Fos synergizes with these oncogenes to promote transformation protecting cells from DNA damage likely through increased Chk1 levels. Future work will address if inhibiting Chk1 in H2K-Fos-LTR mice can reduce OS in a therapeutic setting. Also Fos deficient cells will be used to investigate the role of endogenous Fos in response to genotoxic insults. This is especially important since we found that UCN-01 treatment leads to the specifc upregulation of Fos expression in human OS cells. It will be of hallmark importance to investigate whether inhibiting both AP-1/ Fos and Chk1 can synergize to efficiently kill tumor cells since several Chk1 inhibitors including UCN-01 are currently under investigation in clinical trials. Inhibiting Fos/ AP-1 could potentially increase the efficacy of Chk1 inhibitors and thus lead to improved therapy outcomes. Additionally it will be interesting to investigate a potential role of TGFBI downstream of Fos in regulating replication stress in osteoblasts. In summary these results indentify Fos as a previously unknown regulator of the DNA damage response that protects cells against genotoxic insults and promotes transformation suggesting a role in tumor progression with a potential relevance in a therapeutic setting.
