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Sox9/Sox6 and Sp1 are involved in the insulin-like growth factor-I-mediated upregulation of human type II collagen gene expression in articular chondrocytes

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Abstract

Type II collagen is a marker of articular cartilage encoded by the COL2A1 gene. The nature of the trans factors involved in the upregulation of this gene by insulin-like growth factor-I (IGF-I) remains unclear. We found that IGF-I increased type II collagen synthesis by a transcriptional control mechanism involving a 715-bp region within the COL2A1 first-intron specific enhancer. The overproduction of L-Sox5/Sox6/Sox9 and Sp1 and decoy experiments targeting these factors demonstrated their action in concert in IGF-I trans-activation. These results were supported by the data obtained in knockdown experiments in which siRNA against Sox9/Sox6 and Sp1 prevented the IGF-I-induced increase in collagen II production. Indeed, each of these trans-activators increased the expression of others. IGF-I increased the binding of Sox9 and Sp1/Sp3 to their cis elements in the enhancer, and we provide the first evidence of Sox9 interaction with the promoter by chromatin immunoprecipitation. Interactions with COL2A1 were also observed for Sp1, p300/CBP, and Tip60. Finally, a physical interaction between Sox9, p300, Sp3, and Sp1 was detected. These data demonstrate the role of Sox9, Sp1/Sp3, and euchromatin-associated factors (p300, Tip60) in the IGF-I-induced upregulation of COL2A1, indicating possible use of this growth factor in articular cartilage engineering applications to promote repair in patients with degenerative diseases, such as osteoarthritis.

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References

  1. Lefebvre V, Zhou G, Mukhopadhyay K, Smith CN, Zhang Z, Eberspaecher H, Zhou X, Sinha S, Maity SN, De Crombrugghe B (1996) A 18-base pair sequence in the mouse proα(II) collagen gene is sufficient for expression in cartilage and binds nuclear proteins that are selectively expressed in chondrocytes. Mol Cell Biol 16:4512–4523

    PubMed  CAS  Google Scholar 

  2. Krebsbach PH, Nakata K, Bernier SM, Hatano O, Miyashita T, Rhodes CS, Yamada Y (1996) Identification of a minimum enhancer sequence for the type II collagen gene reveals several core sequence motifs in common with the link protein gene. J Biol Chem 271:4298–4303

    Article  PubMed  CAS  Google Scholar 

  3. Bell DM, Leung KK, Wheatley SC, Ng LJ, Zhou S, Ling KW, Sham MH, Koopman P, Tam PPL, Cheah KSE (1997) SOX9 directly regulates the type-II collagen gene. Nat Genet 16:174–178

    Article  PubMed  CAS  Google Scholar 

  4. Leung KK, Ng LJ, Ho KK, Tam PPL, Cheah KSE (1998) Different cis-regulatory DNA elements mediate developmental stage- and tissue-specific expression of the human COL2A1 gene in transgenic mice. J Cell Biol 141:1291–1300

    Article  PubMed  CAS  Google Scholar 

  5. Ghayor C, Herrouin JF, Chadjichristos C, Ala-Kokko L, Takigawa M, Pujol JP, Galéra P (2000) Regulation of human COL2A1 gene expression in chondrocytes: identification of C-Krox responsive elements and modulation by phenotype alteration. J Biol Chem 275:27421–27438

    PubMed  CAS  Google Scholar 

  6. Zhou G, Lefebvre V, Zhang ZP, Eberspaecher H, de Crombrugghe B (1998) Three high mobility group-like sequences within a 48-base pair enhancer of the Col2a1 gene are required for cartilage-specific expression in vivo. J Biol Chem 273:14989–14997

    Article  PubMed  CAS  Google Scholar 

  7. Lefebvre V, Li P, de Crombrugghe B (1998) A new long form of Sox5 (L-Sox5), Sox6 and Sox9 are coexpressed in chondrogenesis and cooperatively activate the type II collagen gene. EMBO J 17:5718–5733

    Article  PubMed  CAS  Google Scholar 

  8. Ghayor C, Chadjichristos C, Herrouin JF, Ala-Kokko L, Suske G, Pujol JP, Galéra P (2001) Sp3 represses the Sp1-mediated transactivation of the human COL2A1 gene in primary and de-differentiated chondrocytes. J Biol Chem 276:36881–36895

    Article  PubMed  CAS  Google Scholar 

  9. Sakiya I, Tsuji K, Koopman P, Watanabe H, Yamada Y, Shinomiya K, Nifuji A, Noda M (2000) SOX9 enhances aggrecan gene promoter/enhancer activity and is up-regulated by retinoic acid in a cartilage-derived cell line, TC6. J Biol Chem 275:10738–10744

    Article  Google Scholar 

  10. Bi W, Deng JM, Zhang Z, Beringher RR, de Crombrugghe B (1999) Sox9 is required for cartilage formation. Nat Genet 22:85–89

    Article  PubMed  CAS  Google Scholar 

  11. Lefebvre V, Huang W, Harley VR, Goodfellow PN, de Crombrugghe B (1997) SOX9 is a potent activator of the chondrocyte-specific enhancer of the pro alpha1(II) collagen gene. Mol Cell Biol 17:2336–2346

    PubMed  CAS  Google Scholar 

  12. Lefebvre V, de Crombrugghe B (1998) Toward understanding Sox9 function in chondrocyte differentiation. Matrix Biol 16:529–540

    Article  PubMed  CAS  Google Scholar 

  13. Ng LJ, Wheatley S, Muscat GE, Conway-Campbell J, Bowles J, Wright E, Bell DM, Tam PP, Cheah KS, Koopman P (1997) SOX9 binds DNA, activates transcription, and coexpresses with type II collagen during chondrogenesis in the mouse. Dev Biol 183:108–121

    Article  PubMed  CAS  Google Scholar 

  14. Zhao Q, Eberspaecher H, Lefebvre V, de Crombrugghe B (1997) Parallel expression of Sox9 and Col2α1 in cells undergoing chondrogenesis. Dev Dyn 209:377–386

    Article  PubMed  CAS  Google Scholar 

  15. Matsuno A, Sasaki T, Nagashima T, Matsuura R, Tanaka H, Hirakawa M, Murakami M, Kirino T (1997) Immunohistochemical examination of proliferative potentials and the expression of cell cycle-related proteins of intracranial chordomas. Hum Pathol 28:714–719

    Article  PubMed  CAS  Google Scholar 

  16. Kypriotou M, Fossard-Demoor M, Chadjichristos C, Ghayor C, de Crombrugghe B, Pujol JP, Galéra P (2003) Sox9 exerts a bifunctional effect on type II collagen gene (COL2A1) expression in chondrocytes depending on the differentiation state. DNA Cell Biol 22:119–129

    Article  PubMed  CAS  Google Scholar 

  17. Kolettas E, Muir HI, Barrett JC, Hardingham TE (2001) Chondrocyte phenotype and cell survival are regulated by culture conditions and by specific cytokines through the expression of Sox-9 transcription factor. Rheumatology 40:1146–1156

    Article  PubMed  CAS  Google Scholar 

  18. Martel-Pelletier J, Di Battista JA, Lajeunesse D, Pelletier JP (1998) IGF/IGFBP axis in cartilage and bone in osteoarthritis pathogenesis. Inflamm Res 47:90–100

    Article  PubMed  CAS  Google Scholar 

  19. Wang Y, Nishida S, Sakata T, Elalieh HZ, Chang W, Halloran BP, Doty SB, Bikle DD (2006) Insulin-like growth factor is essential for embryonic bone development. Endocrinology 147:4753–4761

    Article  PubMed  CAS  Google Scholar 

  20. Liu JP, Baker J, Perkins AS, Robertson E, Efstratiadis A (1993) Mice carrying null mutations of the genes encoding insulin-like growth factor I (Igf-1) and type 1 IGF receptor (Igf1r). Cell 75:59–72

    PubMed  CAS  Google Scholar 

  21. Baker J, Liu JP, Perkins AS, Robertson E, Efstratiadis A (1993) Role of insulin-like growth factors in embryonic and postnatal growth. Cell 75:73–82

    PubMed  CAS  Google Scholar 

  22. Liu JL, Yakar S, LeRoith D (2000) Conditional knockout of mouse insulin-like growth factor-1 gene using the Cre/lox P system. Proc Soc Exp Biol Med 223:344–351

    Article  PubMed  CAS  Google Scholar 

  23. Middleton J, Manthey A, Tyler J (1996) Insulin-like growth factor (IGF) receptor, IGF-I, interleukin-1 beta (IL-1 beta), and IL-6 mRNA expression in osteoarthritic and normal human cartilage. J Histochem Cytochem 44:133–141

    Article  PubMed  CAS  Google Scholar 

  24. Morales TI (1992) Articular cartilage and osteoarthritis, 1st edn. Raven, New York

    Google Scholar 

  25. Van der Kraan P, Buma P, van Kuppevelt T, van der Berg WB (2002) Interaction of chondrocytes, extracellular matrix and growth factors: relevance for articular cartilage tissue engineering. Osteoarthritis Cartilage 10:631–637

    Article  PubMed  Google Scholar 

  26. Garcia AM, Szasz N, Trippel SB, Morales TI, Grodzinsky AJ, Frank EH (2003) Transport and binding of insulin-like growth factor I through articular cartilage. Arch Biochem Biophys 415:69–79

    Article  PubMed  CAS  Google Scholar 

  27. Mierisch CM, Anderson PC, Balian G, Diduch DR (2002) Treatment with insulin-like growth factor-1 increases chondrogenesis by periosteum in vitro. Connect Tissue Res 43:559–568

    PubMed  CAS  Google Scholar 

  28. Luyten FP, Hascall VC, Nissley SP, Morales TI, Reddi AH (1988) Insulin-like growth factors maintain steady-state metabolism of proteoglycans in bovine articular cartilage explants. Arch Biochem Biophys 267:416–425

    Article  PubMed  CAS  Google Scholar 

  29. Smith P, Schuler FD, Georgescu HI, Ghivizzani SC, Johnstone B, Niyibizi C, Robbins PD, Evans CH (2000) Genetic enhancement of matrix synthesis by articular chondrocytes: comparison of different growth factor genes in the presence and absence of interleukin-1. Arthritis Rheum 43:1156–1164

    Article  PubMed  CAS  Google Scholar 

  30. Madry H, Pareda R, Seidel J, Langer R, Freed LE, Trippel SB, Vunjak-Novakovic G (2002) Gene transfer of a human insulin-like growth factor I cDNA enhances tissue engineering of cartilage. Human Genet Ther 8:1443–1449

    Google Scholar 

  31. Mathieu P (2002) A new mechanism of action of chondroitin sulfates ACS4-ACS6 in osteoarthritic cartilage. Presse Med 31:1383–1385

    PubMed  Google Scholar 

  32. Porée B, Kypriotou M, Chadjichristos C, Beauchef G, Renard E, Legendre F, Melin M, Gueret S, Hartmann DJ, Mallein-Guerin F et al (2008) Interleukin-6 (IL-6) and/or soluble IL-6 receptor down-regulation of human type II collagen gene expression in articular chondrocytes requires a decrease of Sp1.Sp3 ratio and of the binding activity of both factors to the COL2A1 promoter. J Biol Chem 283:4850–4865

    Article  PubMed  Google Scholar 

  33. Chadjichristos C, Ghayor C, Herrouin JF, Ala-Kokko L, Suske G, Pujol JP, Galéra P (2002) Down-regulation of human type II collagen gene expression by transforming growth factor-β1 (TGF-β1) in articular chondrocytes involves Sp1/Sp3 ratio. J Biol Chem 277:43903–43917

    Article  PubMed  CAS  Google Scholar 

  34. Chadjichristos C, Ghayor C, Kypriotou M, Martin G, Renard E, Ala-Kokko L, Suske G, de Crombrugghe B, Pujol JP, Galéra P (2003) Sp1 and Sp3 transcription factors mediate interleukin-1β down-regulation of human type II collagen gene expression in articular chondrocytes. J Biol Chem 278:39762–39772

    Article  PubMed  CAS  Google Scholar 

  35. Shakibaei M, Seifarth C, John T, Rahmanzadeh M, Mobasheri A (2006) IGF-I extends the chondrogenic potential of human articular chondrocytes in vitro: molecular association between Sox9 and Erk1/2. Biochem Pharmacol 72:1382–1395

    Article  PubMed  CAS  Google Scholar 

  36. Magee C, Nurminskaya M, Faverman L, Galera P, Linsenmayer T (2005) Sp3/Sp1 transcription activity regulates specific expression of collagen type X in hypertrophic chondrocytes. J Biol Chem 280:25331–25338

    Article  PubMed  CAS  Google Scholar 

  37. Savagner P, Krebsbach PH, Hatano O, Miyashita T, Liebman J, Yamada Y (1995) Collagen II promoter and enhancer interact synergistically through Sp1 and distinct nuclear factors. DNA Cell Biol 14:501–510

    Article  PubMed  CAS  Google Scholar 

  38. Hattori T, Coustry F, Stephens S, Eberspaecher H, Takigawa M, Yasuda H, de Crombrugghe B (2008) Transcriptional regulation of chondrogenesis by coactivator Tip60 via chromatin association with Sox9 and Sox5. Nucleic Acids Res 36:3011–3024

    Article  PubMed  CAS  Google Scholar 

  39. Furumatsu T, Tsuda M, Yoshida K, Taniguchi N, Ito T, Hashimoto M, Ito T, Asahara H (2005) Sox9 and p300 cooperatively regulate chromatin-mediated transcription. J Biol Chem 280:35203–35208

    Article  PubMed  CAS  Google Scholar 

  40. Tsuda M, Takahashi S, Takahashi Y, Asahara H (2003) Transcriptional co-activators CREB-binding protein and p300 regulate chondrocyte-specific gene expression via association with Sox9. J Biol Chem 278:27224–27229

    Article  PubMed  CAS  Google Scholar 

  41. Kawakami Y, Tsuda M, Takahashi S, Taniguchi N, Esteban CR, Zemmyo M, Furumatsu T, Lotz M, Belmonte JC, Asahara H (2005) Transcriptional coactivator PGC-1α regulates chondrogenesis via association with Sox9. Proc Natl Acad Sci USA 102:2414–2419

    Article  PubMed  CAS  Google Scholar 

  42. Piera-Velazquez S, Hawkins DF, Whitecavage MK, Colter DC, Stokes DG, Jimenez SA (2007) Regulation of the human SOX9 promoter by Sp1 and CREB. Exp Cell Res 313:1069–1079

    Article  PubMed  CAS  Google Scholar 

  43. Breitner-Johnson D, Werner H, Roberts CT Jr, LeRoith D (1995) Regulation of insulin-like growth factor receptor gene expression by Sp1: physical and functional interactions of Sp1 at GC boxes and at a CT element. Mol Endo 9:1147–1156

    Article  Google Scholar 

  44. Kramps C, Strieder V, Sapetschnig A, Suske G, Lutz W (2004) E2F and Sp1/Sp3 synergize but are not sufficient to activate the MYCN gene in neuroblastomas. J Biol Chem 279:5110–5117

    Article  PubMed  CAS  Google Scholar 

  45. Brittberg M, Lindahl A, Nilsson A, Ohlsson C, Isaksson O, Peterson L (1994) Treatment of deep cartilage defects in the knee with autologous chondrocyte transplantation. N Engl J Med 331:889–895

    Article  PubMed  CAS  Google Scholar 

  46. Steinert AF, Nöth U, Tuan RS (2008) Concepts in gene therapy for cartilage repair. Injury 39S1:S97–S113

    Article  Google Scholar 

  47. Zhai G, Rivadeneira F, Houwing-Duistermaat JJ, Meulenbelt I, Bijkerk C, Hofman A, van Meurs JBJ, Uitterlinden AG, Pols HAP, Slagboom PE et al (2003) Insulin-like growth factor I gene promoter polymorphism, collagen type II α1 (COL2A1) gene, and the prevalence of radiographic osteoarthritis: the Rotterdam Study. Ann Rheum Dis 63:544–548

    Article  Google Scholar 

  48. Tew SR, Li Y, Pothacharoen P, Tweats LM, Hawkins RE, Hardingham TE (2005) Retroviral transduction with SOX9 enhances re-expression of the chondrocyte phenotype in passaged osteoarthritic human articular chondrocytes. Osteoarthritis Cartilage 13:80–89

    Article  PubMed  Google Scholar 

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Acknowledgments

This work was supported by the Regional Council of Lower Normandy, the French Ministry of Regional Development, and the ANR (Agence Nationale de la Recherche) TecSan (Technologies de la Santé) program (PROMOCART 2006).

Emmanuelle Renard, Fellowship from the Regional Council of Lower Normandy and Pierre Fabre Laboratories (Vigoulet-Auzil, BP74, 31322 Castanet-Tolosan, France).

Benoît Porée, Laure Maneix and David Ollitrault, Fellowship from the French Ministry of Research and Technology.

Magdalini Kypriotou, Fellowship from the Regional Council of Lower Normandy and the French Ministry of Regional Development.

Nicolas Bigot, Fellowship from the Regional Council of Lower Normandy and Johnson & Johnson Laboratories (Johnson & Johnson Consumer France. R&D Europe. Campus de Maigremont, 27100 Val de Reuil, France).

Florence Legendre, Post-doctoral fellow supported by the PROMOCART grant (Health Technology) from the French National Agency of Research (ANR).

We would like to thank G. Suske (Institut für Molekularbiologie und Tumorforschung, Philipps-Universität, Marburg, Germany) for generously providing the Sp1 and Sp3 expression vectors.

Conflict of interest

The authors declare that they have no conflict of interests.

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Authors and Affiliations

  1. Laboratoire Matrice Extracellulaire et Pathologie, IFR ICORE 146, Université de Caen/Basse-Normandie, UFR de Médecine, CHU niveau 3, Avenue de la Côte de Nacre, 14032, Caen Cedex, France

    Emmanuelle Renard, Benoît Porée, Christos Chadjichristos, Magdalini Kypriotou, Laure Maneix, Nicolas Bigot, Florence Legendre, David Ollitrault, Safa Moslemi, Magali Demoor, Karim Boumediene & Philippe Galéra

  2. Department of Molecular Genetics, University of Texas, M.D. Anderson Cancer Center, Holcombe Boulevard, Houston, TX, 77030, USA

    Benoît De Crombrugghe

  3. Laboratoire de Biologie et Ingénierie du Cartilage, Institut de Biologie et Chimie des Protéines, UMR 5086 CNRS/Université Claude Bernard Lyon 1-IFR 128 Biosciences Lyon-Gerland, 7 Passage du Vercors, 69367, Lyon Cedex 07, France

    Frédéric Malléin-Gérin

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  1. Emmanuelle Renard
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Correspondence to Philippe Galéra.

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Renard, E., Porée, B., Chadjichristos, C. et al. Sox9/Sox6 and Sp1 are involved in the insulin-like growth factor-I-mediated upregulation of human type II collagen gene expression in articular chondrocytes. J Mol Med 90, 649–666 (2012). https://doi.org/10.1007/s00109-011-0842-3

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