Sox9, a key transcription factor of bone morphogenetic protein‐2‐induced chondrogenesis, is activated through BMP pathway and a CCAAT box in the proximal promoter

Abstract

Mouse embryonic fibroblasts (MEFs) can be differentiated into fully functional chondrocytes in response to bone morphogenetic protein-2 (BMP-2). The expression of Sox9, a critical transcription factor for the multiple steps of chondrogenesis, has been reported to be upregulated during this process. But the molecular mechanisms by which BMP-2 promotes chondrogenesis still remain largely unknown. The aim of the present study was therefore to investigate the underlying mechanism. In the MEFs, BMP-2 efficiently induced Sox9 expression along with chondrogenic differentiation in a time- and dose-dependent manner. SB203580, a specific inhibitor for p38 pathway, blocked BMP-2-induced chondrogenic differentiation as well as Sox9 expression and its transactivation of downstream genes. Forced expression of Smad6, a natural antagonist for BMP/Smad pathway, only inhibited Sox9 protein function without rendering any effects on its mRNA expression. A CCAAT box was identified in Sox9 promoter as the cis-elements responsible for BMP-2 stimulation. This study provides insight into the mechanisms underlying BMP-2-regulated Sox9 expression and activity in MEFs, and suggests differential roles of BMP-2/p38 and BMP-2/Smad pathways in modulating the function of Sox9 during chondrogenesis.

miR-199a*, a Bone Morphogenic Protein 2-responsive MicroRNA, Regulates Chondrogenesis via Direct Targeting to Smad1

“…5E), a result that is consistent with previous findings, which identify Smad1 as an intracellular transducer of BMP-2 signaling (32,48,49). Smad1 is also an established mediator of osteoblast differentiation through the activation of target genes, including p204, a factor that binds Cbfa1, pRb, and Ids (40 -42).…”

“…5E), a known downstream signaling molecule of BMP-2 (32,48,49). Thus, we propose that miR199a* mediates early chondrogenesis by following the model proposed by Li et al (50).…”

“…Although we did not find a statistically significant reduction in Sox9 expression levels in C3H10T1/2 cells, this may be due to insufficient sampling, because Sox9 is known to be an essential transcription factor of chondrogenic differentiation. Alternatively, a recent study has found that the BMP-2/Smads pathway regulates Sox9 activity, but does not alter Sox9 mRNA expression levels (48). This may explain why we were unable to observe decreased Sox9 mRNA expression in C3H10T1/2 cells that were overexpressing miR-199a*.…”

TGF-β Family Signaling in Connective Tissue and Skeletal Diseases

“…BMP signaling, in contrast, is dispensable for the initial step of mesenchymal condensation but required for commitment to the chondrogenic lineage, proliferation, differentiation, and maturation of chondrocytes (Li et al 2003;Horiki et al 2004;Yoon et al 2005;Retting et al 2009;Culbert et al 2014). BMP-mediated chondrogenic induction is achieved through the direct transcriptional activation of the transcription factor Sex determining region Y-box 9 (Sox9), which is the master regulator of chondrogenesis (Zehentner et al 1999;Akiyama 2008;Pan et al 2008). Genetic disruption of Sox9 results in perturbed skeletogenesis throughout the body (Akiyama et al 2002;Mori-Akiyama et al 2003).…”

Mechanisms of digit formation: Human malformation syndromes tell the story

“…BMP/Gdf signaling is fine-tuned at several levels, for example by secreted antagonists that interfere with ligand-receptor binding, such as Noggin (NOG; reviewed e.g., in Hoffmann and Gross, 2001;Hartung et al, 2006). Sox9 is a major target of BMP signaling in chondrogenesis (Healy et al, 1999;Zehentner et al, 1999;Pan et al, 2008Pan et al, , 2009). Sox9 in turn positively regulates expression of NOG, providing negative feedback (Zehentner et al, 2002).…”