IADR Abstract Archives
Epigenetic Remodeling and Modification to Maintain Bone Mass in Vivo
Objectives: Current studies do not have any insight on how epigenetic remodeling and modification of bone-specific chromatin are maintaining bone mass in vivo. Understanding of this gap is pivotal for further studies to identify specific targets for future therapies which protect bone loss. In this study, we are testing epigenetic mechanisms where a miRNA cluster controls bone synthesis and homeostasis by regulating Histone H3K27 modification activity.
Methods: To address the above queries in vivo, we created an inducible anti-miR-23a cluster (miR-23aClZIP) knockdown mouse model, an activating chromatin remodeler, BAF45A conditional mouse line (Baf45afl/fl,) and an Ezh2fl/fl mouse line for Polycomb Repressive 2 Complex (PRC2) mouse line. RNA sequencing, microCT, Co-IP, in-vitro overexpression and knockdown, and ChIP assay were utilized to decipher the regulatory mechanism.
Results: MiR-23a cluster knockdown mice developed high cortical and trabecular bone. RNA sequencing from these mice displayed increased expression of Runx2 and Phf10 essential for skeletogenesis and decreased expression of Ezh2, a chromatin repressor also indispensable for skeletogenesis. Osteoblast-specific PHf10 deletion (Phf10fl/fl, OCN-Cre) significantly reduced bone formation. ChIP assays with bone essential promoters using miR-23a cluster knockdown and Phf10fl/fl calvarial cells revealed an epigenetic regulation that promotes bone formation. Furthermore, we observed that Runx2 and Phf10 decreased the suppressive activity of Ezh2 to promoters of bone essential genes, for osteogenesis.
Conclusions: Together, our findings strongly suggest that miR-23a cluster connection with tissue-specific RUNX2-Phf10–EZH2 function, linked to bone homeostasis, is a very unique regulatory mechanism.
Methods: To address the above queries in vivo, we created an inducible anti-miR-23a cluster (miR-23aClZIP) knockdown mouse model, an activating chromatin remodeler, BAF45A conditional mouse line (Baf45afl/fl,) and an Ezh2fl/fl mouse line for Polycomb Repressive 2 Complex (PRC2) mouse line. RNA sequencing, microCT, Co-IP, in-vitro overexpression and knockdown, and ChIP assay were utilized to decipher the regulatory mechanism.
Results: MiR-23a cluster knockdown mice developed high cortical and trabecular bone. RNA sequencing from these mice displayed increased expression of Runx2 and Phf10 essential for skeletogenesis and decreased expression of Ezh2, a chromatin repressor also indispensable for skeletogenesis. Osteoblast-specific PHf10 deletion (Phf10fl/fl, OCN-Cre) significantly reduced bone formation. ChIP assays with bone essential promoters using miR-23a cluster knockdown and Phf10fl/fl calvarial cells revealed an epigenetic regulation that promotes bone formation. Furthermore, we observed that Runx2 and Phf10 decreased the suppressive activity of Ezh2 to promoters of bone essential genes, for osteogenesis.
Conclusions: Together, our findings strongly suggest that miR-23a cluster connection with tissue-specific RUNX2-Phf10–EZH2 function, linked to bone homeostasis, is a very unique regulatory mechanism.