L-Sox5 and Sox6 drive expression of the aggrecan gene in cartilage by securing binding of Sox9 to a far-upstream enhancer.
The Sry-related high-mobility-group box transcription factor Sox9 recruits the redundant L-Sox5 and Sox6 proteins to effect chondrogenesis, but the mode of action of the trio remains unclear. We identify here a highly conserved 359-bp sequence 10 kb upstream of the Agc1 gene for aggrecan, a most essential cartilage proteoglycan and key marker of chondrocyte differentiation. This sequence directs expression of a minimal promoter in both embryonic and adult cartilage in transgenic mice, in a manner that matches Agc1 expression. The chondrogenic trio is required and sufficient to mediate the activity of this enhancer. It acts directly, Sox9 binding to a critical cis-acting element and L-Sox5/Sox6 binding to three additional elements, which are cooperatively needed. Upon binding to their specific sites, L-Sox5/Sox6 increases the efficiency of Sox9 binding to its own recognition site and thereby robustly potentiates the ability of Sox9 to activate the enhancer. L-Sox5/Sox6 similarly secures Sox9 binding to Col2a1 (encoding collagen-2) and other cartilage-specific enhancers. This study thus uncovers critical cis-acting elements and transcription factors driving Agc1 expression in cartilage and increases understanding of the mode of action of the chondrogenic Sox trio.
Han
Yu
Y
Department of Cell Biology and Orthopaedic Research Center, Cleveland Clinic, Cleveland, OH 44195, USA.
Lefebvre
Véronique
V
D053674:Aggrecans / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002356:Cartilage / Q000378:metabolism*; D002460:Cell Line; D019902:Chondrocytes / Q000378:metabolism; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005091:Exons / Q000235:genetics; D005786:Gene Expression Regulation; D006609:High Mobility Group Proteins / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011485:Protein Binding; D051381:Rats; D055758:SOX9 Transcription Factor; D055755:SOXD Transcription Factors; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection
D053674:Aggrecans; D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; D009687:Nuclear Proteins; D055758:SOX9 Transcription Factor; C527932:SOX9 protein, human; D055755:SOXD Transcription Factors; C089186:Sox5 protein, mouse; C095939:Sox6 protein, mouse; C527933:Sox9 protein, mouse; D014157:Transcription Factors