Runx2- and Histone Deacetylase 3-mediated Repression Is Relieved in Differentiating Human Osteoblast Cells to Allow High Bone Sialoprotein Expression

Abstract

Bone sialoprotein (BSP) is a bone matrix glycoprotein whose expression coincides with terminal osteoblastic differentiation and the onset of mineralization. In this study we show that BSP expression is considerably increased in confluent Saos-2 human osteosarcoma cells and in differentiating normal human osteoblasts, concomitantly with the decrease of Runx2, a key transcription factor controlling bone formation. Therefore, we investigated the role of Runx2 in the regulation of BSP expression in Saos-2 cells. Using a mobility shift assay, we demonstrated that Runx2 binds to the BSP promoter only in preconfluent cells. Histone deacetylase 3 (HDAC3) has been recently shown to act as a Runx2 co-repressor. Chromatin immunoprecipitation assays demonstrated that both Runx2 and HDAC3 are detectable at the BSP promoter in preconfluent Saos-2 cells but not when they are confluent and overexpress BSP. Consistently, nuclear Runx2 protein level is down-regulated, whereas Saos-2 cells became increasingly confluent. Finally, the suppression of HDAC3, Runx2, or both by RNA interference induced the expression of BSP at both mRNA and protein levels in Saos-2 cells. Our data demonstrate that Runx2 and HDAC3 repress BSP gene expression and that this repression is suspended upon osteoblastic cell differentiation. Both the nuclear disappearance of Runx2 and the non-recruitment of HDAC3 represent new means to relieve Runx2-mediated suppression of BSP expression, thus allowing the acquisition of a fully differentiated and mineralization-competent phenotype by osteoblast cells.

The role for runt related transcription factor 2 (RUNX2) as a transcriptional repressor in luteinizing granulosa cells

“…HDACs were found to act as co-repressors of RUNX2 in various cell types (Hug, 2004; Jensen et al, 2007; Jensen et al, 2008; Lamour et al, 2007; Shimizu et al, 2010; Sun et al, 2009; Westendorf, 2006). Therefore, the periovulatory expression of Hdac1, Hdac3 and rSinc3a were assessed in whole ovaries collected at different times after hCG administration.…”

Osteosarcoma Development and Stem Cell Differentiation

“…Runx2 levels and function are biologically linked to a cell growth-related G(1) transition in osteoblastic cells [52]. Runx2 and histone deacetylase 3-mediated repression is believed to allow high expression of bone sialoprotein-a bone matrix glycoprotein whose expression coincides with terminal osteoblastic differentiation and the onset of mineralization-in differentiating human osteoblast cells [122]. Runx2 is hyperphosphorylated by CDK1/ cyclin B during mitosis, and dynamically converted into a hypophosphorylated form by PP1/PP2A-dependent dephosphorylation after mitosis to support the postmitotic regulation of Runx2 target genes [201].…”

Histone deacetylases in skeletal development and bone mass maintenance

“…Hdac3 binds Runx2, NFATc1, Zfp521 and TCF to suppress osteoblast-specific gene expression (e.g., osteocalcin and bone sialoprotein) (Schroeder et al, 2004; Lamour et al, 2007; Jensen et al, 2008; Choo et al, 2009; Wu et al, 2009; Hesse et al, 2010). Zfp521 may recruit Hdac3 to Runx2 complexes to promote repression of Runx2’s transcriptional activity (Wu et al, 2009; Hesse et al, 2010).…”