BMP2-activated Erk/MAP Kinase Stabilizes Runx2 by Increasing p300 Levels and Histone Acetyltransferase Activity

Abstract

Runx2 is a critical transcription factor for osteoblast differentiation. Regulation of Runx2 expression levels and transcriptional activity is important for bone morphogenetic protein (BMP)-induced osteoblast differentiation. Previous studies have shown that extracellular signal-regulated kinase (Erk) activation enhances the transcriptional activity of Runx2 and that BMP-induced Runx2 acetylation increases Runx2 stability and transcriptional activity. Because BMP signaling induces Erk activation in osteoblasts, we sought to investigate whether BMP-induced Erk signaling regulates Runx2 acetylation and stability. Erk activation by overexpression of constitutively active MEK1 increased Runx2 transcriptional activity, whereas U0126, an inhibitor of MEK1/2, suppressed basal Runx2 transcriptional activity and BMP-induced Runx2 acetylation and stabilization. Overexpression of constitutively active MEK1 stabilized Runx2 protein via up-regulation of acetylation and down-regulation of ubiquitination. Erk activation increased p300 protein levels and histone acetyltransferase activity. Knockdown of p300 using siRNA diminished Erk-induced Runx2 stabilization. Overexpression of Smad5 increased Runx2 acetylation and stabilization. Erk activation further increased Smad-induced Runx2 acetylation and stabilization, whereas U0126 suppressed these functions. On the other hand, knockdown of Smad1 and Smad5 by siRNA suppressed both basal and Erk-induced Runx2 protein levels. Erk activation enhanced the association of Runx2 with p300 and Smad1. Taken together these results indicate that Erk signaling increases Runx2 stability and transcriptional activity, partly via increasing p300 protein levels and histone acetyltransferase activity and subsequently increasing Runx2 acetylation by p300. In addition to the canonical Smad pathway, a BMP-induced non-Smad Erk signaling pathway cooperatively regulates osteoblast differentiation partly via increasing the stability and transcriptional activity of Runx2.The bone morphogenetic proteins (BMPs) 3 are members of the transforming growth factor-␤ superfamily and are primary growth factors that induce formation of both cartilage and bone. Receptors for BMP are serine/threonine kinase receptors and consist of type I (BMPR-I) and II (BMPR-II) receptors. After ligand binding, BMPR-I kinases are activated by BMPR-II kinase-induced phosphorylation. R-Smad proteins are then recruited to activated receptors and play a role in transmitting the BMP signal from the receptor to target genes such as alkaline phosphatase (ALP), bone sialoprotein, osteocalcin (OC), Runx2, and Dlx5 (1, 2). In addition to the Smad pathway, diverse intracellular signaling molecules also participate in BMP-induced osteoblast differentiation. These are collectively called the non-Smad pathway of BMP signaling and include extracellular signal-regulated protein kinase (Erk), p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, phosphatidylinositol 3-kinase, and protein kinases C and D (3-7). These cooperate with and/or reg...

Lung Tumor-associated Osteoblast-derived Bone Morphogenetic Protein-2 Increased Epithelial-to-Mesenchymal Transition of Cancer by Runx2/Snail Signaling Pathway

“…p300, a histone acetyltransferase, has been reported to interact with Runx2 and to subsequently regulate the expression of osteocalcin and MMP-13 (37,38). In addition, p300 also acts as a stabilizer of Runx2 via acetylation (39). Our data are the first to demonstrate that BMP-2 increases the expression of Runx2, which interacts with p300 protein and in turn enhances the acetylation of histone H3 and H4, resulting in the up-regulation of Snail.…”

Inhibition of CK2 binding to BMPRIa induces C2C12 differentiation into osteoblasts and adipocytes

“…BMP2-induced osteoblast differentiation markers, alkaline phosphatase (ALP) staining, Osterix, and the expression of collagen type Iα and ALP are decreased with the incubation with the MEK inhibitor, U0126, in C2C12 cells. Additionally BMP2 activated ERK increases mRNA and protein levels of RUNX2 and its stability suggesting a possible mechanism (Gallea et al 2001;Jun et al 2010). The MEK inhibitor, PD98059, decreases osteocalcin (a marker for osteoblasts) expression in C2C12 cells although its direct effect on ALP is unclear (Gallea et al 2001 Although the direct activation of the MEK/ERK pathway due to MCK2.2 or MCK2.3 was not shown here, the previous studies did demonstrate that BMP2 activates the MEK/ERK pathway in C2C12 cells and the pathway directs osteogenesis (Gallea et al 2001;Jun et al 2010), supporting the results with the MEK inhibitor and MCK2.3.…”

“…Additionally BMP2 activated ERK increases mRNA and protein levels of RUNX2 and its stability suggesting a possible mechanism (Gallea et al 2001;Jun et al 2010). The MEK inhibitor, PD98059, decreases osteocalcin (a marker for osteoblasts) expression in C2C12 cells although its direct effect on ALP is unclear (Gallea et al 2001 Although the direct activation of the MEK/ERK pathway due to MCK2.2 or MCK2.3 was not shown here, the previous studies did demonstrate that BMP2 activates the MEK/ERK pathway in C2C12 cells and the pathway directs osteogenesis (Gallea et al 2001;Jun et al 2010), supporting the results with the MEK inhibitor and MCK2.3. It does not explain the results demonstrating MEK/ERK pathway is involved with adipogenesis via MCK2.2, although in primary bone marrow stromal stem cells ERK activation is needed for adipogenesis (Liao et al 2008).…”

c-Jun N-terminal kinase 1 negatively regulates osteoblastic differentiation induced by BMP2 via phosphorylation of Runx2 at Ser104

“…(8,14,16) For example, both phosphorylation and transcriptional activity of Runx2 can be stimulated by the ERK and p38 pathways. (17)(18)(19)(20)(21)(22) Although the involvements of ERK and p38 pathways in osteoblastic differentiation have been well studied, the phosphorylation and activity of Runx2 modified by JNK during osteoblastic differentiation remains to be clarified.…”