RUNX2 Regulates Osteoblast Differentiation via the BMP4 Signaling Pathway

Abstract

RUNX2 is a master osteogenic transcription factor, and mutations in RUNX2 cause the inherited skeletal disorder cleidocranial dysplasia (CCD). Studies have revealed that RUNX2 is not only a downstream target of the bone morphogenetic protein (BMP) pathway but can also regulate the expression of BMPs. However, the underlying mechanism of the regulation of BMPs by RUNX2 remains unknown. In this project, we diagnosed a CCD patient with a 7.86-Mb heterozygous deletion on chromosome 6 containing all exons of RUNX2 by multiplex ligation-dependent probe amplification (MLPA) and whole-genome sequencing (WGS). Bone marrow mesenchymal stem cells (BMSCs) were further extracted from patient alveolar bone fragments (CCD-BMSCs), an excellent natural model to explore the possible mechanism. The osteogenic differentiation ability of CCD-BMSCs was severely affected by RUNX2 heterozygous deletion. Also, BMP4 decreased most in BMP ligands, and CHRDL1, a BMP antagonist, was abnormally elevated in CCD-BMSCs. Furthermore, BMP4 treatment essentially rescued the osteogenic capacity of CCD-BMSCs, and RUNX2 overexpression reversed the abnormal expression of BMP4 and CHRDL1. Notably, we constructed CRISPR/Cas9 Runx2+/m MC3T3-E1 cells, which simulated a variant in CCD-BMSCs, to exclude the interference of other gene deletions and the heterogeneity of the genetic background of primary cells, and verified all findings from the CCD-BMSCs. Moreover, the luciferase reporter experiment showed that RUNX2 could inhibit the transcription of CHRDL1. Through immunofluorescence, the inhibitory effect of CHRDL1 on BMP4/Smad signaling was confirmed in MC3T3-E1 cells. These results revealed that RUNX2 regulated the BMP4 pathway by inhibiting CHRDL1 transcription. We collectively identified a novel RUNX2/CHRDL1/BMP4 axis to regulate osteogenic differentiation and noted that BMP4 might be a valuable therapeutic option for treating bone diseases.

Oligomeric proanthocyanidins ameliorates osteoclastogenesis through reducing OPG/RANKL ratio in chicken's embryos

“…Previous studies have shown that loss of BMD can be improved by enhancing the osteogenesis of BMSCs in age-related rat model bone ( Yu et al, 2020 ). Runx2 is a major transcription factor for osteogenesis and plays a key regulatory role during the differentiation of BMSCs into osteoblasts ( Fisher and Franz-Odendaal, 2012 ; Liu et al, 2022 ). Deletion of Runx2 in BMSCs impairs the prolongation and the thickness of bone ( Shu et al, 2021 ).…”

TRIM21 silencing inhibits the apoptosis and expedites the osteogenic differentiation of dexamethasone‑induced MC3T3‑E1 cells by activating the Keap1/Nrf2 pathway

“…Belonging to the BMP family, BMP2, which plays a major role in bone development and remodeling, is a well-established promoter of osteoblastic differentiation and bone formation ( 39 ). Runx2 is the main downstream regulator of the BMP signaling pathway that regulates the expression of several osteogenic genes ( 40 ). Osx is indispensable for osteogenic differentiation and its overexpression is related to the enhancement of osteogenic differentiation ( 41 ).…”

KLF transcription factors in bone diseases

“…156,157 RUNX2 is a key central regulator of osteoblast differentiation, matrix synthesis and mineralization. [157][158][159][160] By interacting with RUNX2 in osteoblasts, KLF2…”

“…New research demonstrated that overexpression of KLF2 enhanced the formation of bone nodules and drastically up‐regulated the expression of the genes that indicated osteoblast differentiation (RUNX2, ALP, OSX, OCN and BSP) 156,157 . RUNX2 is a key central regulator of osteoblast differentiation, matrix synthesis and mineralization 157–160 . By interacting with RUNX2 in osteoblasts, KLF2 increases the activity of genes unique to osteoblasts, hence boosting osteogenic differentiation 157 .…”