The Promyelotic Leukemia Zinc Finger Promotes Osteoblastic Differentiation of Human Mesenchymal Stem Cells as an Upstream Regulator of CBFA1

Abstract

Ossification of the posterior longitudinal ligament of the spine (OPLL) is the leading cause of myelopathy in Japan and is diagnosed by ectopic bone formation in the paravertebral ligament. OPLL is a systemic high bone mass disease with a strong genetic background. To detect genes relevant to the pathogenesis of OPLL, we performed a cDNA microarray analysis of systematic gene expression profiles during the osteoblastic differentiation of ligament cells from OPLL patients (OPLL cells), patients with a disorder called ossification of yellow ligament (OYL), and non-OPLL controls together with human mesenchymal stem cells (hMSCs) after stimulating them with osteogenic differentiation medium (OS). Twenty-four genes were up-regulated during osteoblastic differentiation in OPLL cells. Zinc finger protein 145 (promyelotic leukemia zinc finger or PLZF) was one of the highly expressed genes during osteoblastic differentiation in all the cells examined. We investigated the roles of PLZF in the regulation of osteoblastic differentiation of hMSCs and C2C12 cells. Small interfering RNA-mediated gene silencing of PLZF resulted in a reduction in the expression of osteoblast-specific genes such as the alkaline phosphatase, collagen 1A1 (Col1a1), Runx2/core-binding factor 1 (Cbfa1), and osteocalcin genes, even in the presence of OS in hMSCs. The expression of PLZF was unaffected by the addition of bone morphogenetic protein 2 (BMP-2), and the expression of BMP-2 was not affected by PLZF in hMSCs. In C2C12 cells, overexpression of PLZF increased the expression of Cbfa1 and Col1a1; on the other hand, the overexpression of CBFA1 did not affect the expression of Plzf. These findings indicate that PLZF plays important roles in early osteoblastic differentiation as an upstream regulator of CBFA1 and thereby might participate in promoting the ossification of spinal ligament cells in OPLL patients.Ossification of the posterior longitudinal ligament of the spine (OPLL) 1 is a common hyperostotic disorder among Japanese and other Asian populations with a prevalence of 2-4% of the general population (1). OPLL patients show ectopic bone formation in the posterior longitudinal ligament, which compresses the spinal cord leading to various neurological symptoms. Despite the late onset (average age of onset is 50 years), OPLL has a strong genetic background as shown in classical epidemiological studies and by an estimate of relative risk to siblings of ϳ10 (2, 3). Because genetic factors appear to play crucial roles in the etiology of OPLL, genetic screenings to identify susceptibility are important. To date, we have performed genetic linkage and linkage disequilibrium studies to identify the causalities of OPLL and have successfully identified collagen 11A2 (COL11A2) and collagen 6A1 (3-6). We also demonstrated that an intron 6 (Ϫ4a) polymorphism of COL11A2 affects the splicing of exon 6 in ligament cells from OPLL patients (5). Although the identification of genetic factors of OPLL has important implications, the overall picture of the molecul...

Identification of Novel EZH2 Targets Regulating Osteogenic Differentiation in Mesenchymal Stem Cells

“…Small interfering RNA knockdown of ZBTB16 in human and mouse MSC reduced osteogenic differentiation and expression of collagen 1A1 (COL1A1), alkaline phosphatase, RUNX2, and OC. Furthermore, overexpression of Zbtb16 in a murine MSC line increased Runx2 and Col1A expression during osteogenic differentiation [39]. Collectively, these studies and this current study have identified ZBTB16 as a regulator of osteogenic differentiation.…”

“…ZBTB16 has previously been implicated in regulating osteogenesis in human MSC by acting upstream of RUNX2 to promote osteogenic differentiation and the expression of key osteogenic differentiation genes [38,39]. However, the direct role of MX1 and FHL1 in regulating MSC osteogenic differentiation is not as well defined.…”

“…This study indicated that ZBTB16 may play a critical role in the development of the skeleton and bone aging. Furthermore, Ikeda et al (2005) found ZBTB16 was one of 24 genes upregulated during osteoblastic differentiation of OPLL cells [39].…”

The Transcription Factor DLX3 Regulates the Osteogenic Differentiation of Human Dental Follicle Precursor Cells

“…The expression of ZBTB16 was previously reported in DFCs induced with dexamethasone but not in BMP2-or IGF2-differentiated cells [20,24]. ZBTB16 was reported as an inducer of osteoblast-like and osteoblast cell differentiation and mineralization acting upstream of RUNX2 independent but very closely related to the BMP-signaling pathway, [47,48]. Interestingly, ZBTB16, RUNX2, and ALP were up-regulated while BSP was downregulated after DLX3 overexpression.…”

The alpha subunit of Go interacts with promyelocytic leukemia zinc finger protein and modulates its functions

“…In particular, previous studies by our group demonstrate that Goα regulates PKA signaling and increases the number of newly formed neurites [3,4], supporting its role as an activator of neuronal differentiation. PLZF induces cell differentiation in megakaryocytes [42], osteoblasts [43], and neurons [44]. Activation of CB also promotes neuronal differentiation [13,45].…”