title: A role for fibroblast growth factor receptor-2 in the altered osteoblast phenotype induced by Twist haploinsufficiency in the Saethre-Chotzen syndrome.
abstract: Genetic mutations of Twist, a bHLH transcription factor, induce premature fusion of cranial sutures (craniosynostosis) in the Saethre-Chotzen syndrome (SCS). The mechanisms by which Twist haploinsufficiency may alter osteoblast differentiation are poorly understood. In this study, we investigated the role of fibroblast growth factor receptor-2 (Fgfr2) in the abnormal osteoblast differentiation in SCS. Cranial osteoblasts from an SCS patient with a Y103X mutation inducing deletion of the Twist bHLH domain showed decreased Fgfr2 mRNA levels associated with decreased expression of Runx2, bone sialoprotein (BSP) and osteocalcin (OC), markers of differentiated osteoblasts, compared with wild-type osteoblasts. Transfection with Twist or Runx2 expression vectors, but not with Runx2 mutant which impairs DNA binding, restored Fgfr2, Runx2, BSP and OC expression in Twist mutant osteoblasts. EMSA analysis of mutant osteoblast nuclear extracts showed reduced Runx2 binding to a target OSE2 site in the Fgfr2 promoter. ChIP analyses showed that both Twist and Runx2 in mutant osteoblast nuclear extracts bind to a specific region in the Fgfr2 promoter. Significantly, forced expression of Fgfr2 restored Runx2 and osteoblast marker genes, whereas a dominant-negative Fgfr2 further decreased Runx2 and downstream genes in Twist mutant osteoblasts, indicating that alteration of Fgfr2 results in downregulation of osteoblast genes in Twist mutant osteoblasts. We conclude that Twist haploinsufficiency downregulates Fgfr2 mRNA expression, which in turn reduces Runx2 and downstream osteoblast-specific genes in human calvarial osteoblasts. This provides genetic and biochemical evidence for a role of Fgfr2 in the altered osteoblast phenotype induced by Twist haploinsufficiency in the SCS.
authors.affiliation: Laboratory of Osteoblast Biology and Pathology, INSERM U606, Paris, University Paris 7, Hôpital Lariboisière, Paris, France.
authors.forename: Hind
authors.initials: H
authors.lastname: Guenou
authors.forename: Karim
authors.initials: K
authors.lastname: Kaabeche
authors.forename: Sandrine Le
authors.initials: SL
authors.lastname: Mée
authors.forename: Pierre J
authors.initials: PJ
authors.lastname: Marie
mesh_terms: D000168:Acrocephalosyndactylia / Q000235:genetics*; D001483:Base Sequence; D050659:Core Binding Factor Alpha 1 Subunit / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers; D006801:Humans; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D010006:Osteoblasts / Q000378:metabolism*; D010641:Phenotype; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics; D051497:Receptor, Fibroblast Growth Factor, Type 2 / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014162:Transfection; D051879:Twist-Related Protein 1 / Q000235:genetics*
chemical_list: D050659:Core Binding Factor Alpha 1 Subunit; D017931:DNA Primers; D009687:Nuclear Proteins; D012333:RNA, Messenger; C493709:RUNX2 protein, human; C103173:TWIST1 protein, human; D051879:Twist-Related Protein 1; D051497:Receptor, Fibroblast Growth Factor, Type 2