title: Transcriptional regulation of Vascular Endothelial Growth Factor (VEGF) by osteoblast-specific transcription factor Osterix (Osx) in osteoblasts.
abstract: Osterix (Osx) is an osteoblast-specific transcription factor required for bone formation and osteoblast differentiation. The critical step in bone formation is to replace the avascular cartilage template with vascularized bone. Osteogenesis and angiogenesis are associated with each other, sharing some essential regulators. Vascular endothelial growth factor (VEGF) is involved in both angiogenesis and osteogenesis. Transcriptional regulation of VEGF expression is not well known in osteoblasts. In this study, quantitative real-time RT-PCR results revealed that VEGF expression was down-regulated in Osx-null calvarial cells and that osteoblast marker osteocalcin expression was absent. Overexpression of Osx in stable C2C12 mesenchymal cells using a Tet-off system resulted in up-regulation of both osteocalcin and VEGF expression. The inhibition of Osx by siRNA led to repression of VEGF expression in osteoblasts. These results suggest that Osx controls VEGF expression. Transfection assays demonstrated that Osx activated VEGF promoter activity. A series of VEGF promoter deletion mutants were examined and the minimal Osx-responsive region was defined to the proximal 140-bp region of the VEGF promoter. Additional point mutants were used to identify two GC-rich regions that were responsible for VEGF promoter activation by Osx. Gel shift assay showed that Osx bound to the VEGF promoter sequence directly. Chromatin immunoprecipitation assays indicated that endogenous Osx associated with the native VEGF promoter in primary osteoblasts. Moreover, immunohistochemistry staining showed decreased VEGF protein levels in the tibiae of Osx conditional knock-out mice. We provide the first evidence that Osx controlled VEGF expression, suggesting a potential role of Osx in coordinating osteogenesis and angiogenesis.
authors: Bone Research Laboratory, Texas Scottish Rite Hospital for Children, Dallas, Texas 75219, USA.
authors: Wanjin
authors: W
authors: Tang
authors: Fan
authors: F
authors: Yang
authors: Yang
authors: Y
authors: Li
authors: Benoit
authors: B
authors: de Crombrugghe
authors: Hongli
authors: H
authors: Jiao
authors: Guozhi
authors: G
authors: Xiao
authors: Chi
authors: C
authors: Zhang
mesh_terms: D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D005786:Gene Expression Regulation / Q000502:physiology*; D057809:HEK293 Cells; D006801:Humans; D051379:Mice; D008817:Mice, Mutant Strains; D018919:Neovascularization, Physiologic / Q000235:genetics; D010006:Osteoblasts / Q000166:cytology / Q000378:metabolism*; D015675:Osteocalcin / Q000096:biosynthesis / Q000235:genetics; D010012:Osteogenesis / Q000502:physiology; D017354:Point Mutation; D020218:Response Elements / Q000502:physiology*; D017384:Sequence Deletion; D000076182:Sp7 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000502:physiology*; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000076182:Sp7 Transcription Factor; C479466:Sp7 protein, mouse; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; C467480:vascular endothelial growth factor A, mouse; D015675:Osteocalcin