title: SOX9 governs differentiation stage-specific gene expression in growth plate chondrocytes via direct concomitant transactivation and repression.
abstract: Cartilage and endochondral bone development require SOX9 activity to regulate chondrogenesis, chondrocyte proliferation, and transition to a non-mitotic hypertrophic state. The restricted and reciprocal expression of the collagen X gene, Col10a1, in hypertrophic chondrocytes and Sox9 in immature chondrocytes epitomise the precise spatiotemporal control of gene expression as chondrocytes progress through phases of differentiation, but how this is achieved is not clear. Here, we have identified a regulatory element upstream of Col10a1 that enhances its expression in hypertrophic chondrocytes in vivo. In immature chondrocytes, where Col10a1 is not expressed, SOX9 interacts with a conserved sequence within this element that is analogous to that within the intronic enhancer of the collagen II gene Col2a1, the known transactivation target of SOX9. By analysing a series of Col10a1 reporter genes in transgenic mice, we show that the SOX9 binding consensus in this element is required to repress expression of the transgene in non-hypertrophic chondrocytes. Forced ectopic Sox9 expression in hypertrophic chondrocytes in vitro and in mice resulted in down-regulation of Col10a1. Mutation of a binding consensus motif for GLI transcription factors, which are the effectors of Indian hedgehog signaling, close to the SOX9 site in the Col10a1 regulatory element, also derepressed transgene expression in non-hypertrophic chondrocytes. GLI2 and GLI3 bound to the Col10a1 regulatory element but not to the enhancer of Col2a1. In addition to Col10a1, paired SOX9-GLI binding motifs are present in the conserved non-coding regions of several genes that are preferentially expressed in hypertrophic chondrocytes and the occurrence of pairing is unlikely to be by chance. We propose a regulatory paradigm whereby direct concomitant positive and negative transcriptional control by SOX9 ensures differentiation phase-specific gene expression in chondrocytes. Discrimination between these opposing modes of transcriptional control by SOX9 may be mediated by cooperation with different partners such as GLI factors.
authors.affiliation: Department of Biochemistry, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Hong Kong, China.
authors.forename: Victor Y L
authors.initials: VY
authors.lastname: Leung
authors.forename: Bo
authors.initials: B
authors.lastname: Gao
authors.forename: Keith K H
authors.initials: KK
authors.lastname: Leung
authors.forename: Ian G
authors.initials: IG
authors.lastname: Melhado
authors.forename: Sarah L
authors.initials: SL
authors.lastname: Wynn
authors.forename: Tiffany Y K
authors.initials: TY
authors.lastname: Au
authors.forename: Nelson W F
authors.initials: NW
authors.lastname: Dung
authors.forename: James Y B
authors.initials: JY
authors.lastname: Lau
authors.forename: Angel C Y
authors.initials: AC
authors.lastname: Mak
authors.forename: Danny
authors.initials: D
authors.lastname: Chan
authors.forename: Kathryn S E
authors.initials: KS
authors.lastname: Cheah
mesh_terms: D000818:Animals; D001483:Base Sequence; D001846:Bone Development / Q000235:genetics*; D002356:Cartilage / Q000254:growth & development*; D002454:Cell Differentiation / Q000235:genetics; D019902:Chondrocytes / Q000166:cytology / Q000378:metabolism; D020219:Chondrogenesis / Q000235:genetics*; D024043:Collagen Type II / Q000235:genetics*; D024121:Collagen Type X / Q000235:genetics*; D004742:Enhancer Elements, Genetic / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D006132:Growth Plate / Q000254:growth & development*; D053823:Hedgehog Proteins / Q000235:genetics; D051741:Kruppel-Like Transcription Factors / Q000235:genetics*; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009154:Mutation; D059372:Nucleotide Motifs / Q000235:genetics; D011401:Promoter Regions, Genetic; D055758:SOX9 Transcription Factor / Q000378:metabolism*; D015398:Signal Transduction; D015533:Transcriptional Activation; D000071676:Zinc Finger Protein GLI1
chemical_list: C447413:Col2a1 protein, mouse; D024043:Collagen Type II; D024121:Collagen Type X; C498520:Gli1 protein, mouse; D053823:Hedgehog Proteins; D051741:Kruppel-Like Transcription Factors; D055758:SOX9 Transcription Factor; D000071676:Zinc Finger Protein GLI1
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