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Cloning, molecular characterization, and tissue differential expression of connective tissue growth factor (ctgf) of grass carp

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Abstract

Connective tissue growth factor (ctgf) is involved in the proliferation, migration, adhesion of cell, and the constituent of extracellular matrix, which plays an important role in embryogenesis, angiogenesis, wound repair, and fibrosis diseases. In this study, the cDNA sequence of grass carp ctgf gene was cloned by rapid amplification of cDNA ends (RACE) method; then, the characteristics of this gene and the predicted protein sequence were analyzed by bioinformatics methods, and the tissue differential expression pattern was detected by the quantitative real-time PCR. The results showed that the grass carp ctgf gene has a full-length of 2223 bp, encoding 343 amino acids. The deduced CTGF protein is a hydrophilic and secretary protein with a molecular mass of 37,978.2 Da and an isoelectric point of 8.22. The signal peptide locates between residue positions 1 and 22 of the polypeptide chain. The protein contains α-helix, β-strand, and loops. The CTGF protein of grass carp shows a homology of 98%, 96%, 91%, and 91% with Wuchang bream (Megalobrama amblycephala), zebrafish (Danio rerio), common carp (Cyprinus carpio), and Mexican tetra (Astyanax mexicanus). The grass carp ctgf gene expressed significantly higher in blood and spleen than that in other tissues (P < 0.05). The low expression tissues included the heart, gill, skin, muscle, kidney, brain, and intestinal, and the lowest expression tissue was the liver. The results are consistent with the function of this gene.

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References

  • Abreu JG, Ketpura NI, Reversade B, De Robertis EM (2002) Connective-tissue growth factor (CTGF) modulates cell signalling by BMP and TGF-beta. Nat Cell Biol 4:599–604

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Babic AM, Chen CC, Lau LF (1999) Fisp12/mouse connective tissue growth factor mediates endothelial cell adhesion and migration through integrin v3, promotes endothelial cell survival, and induces angiogenesis in vivo. Mol Cell Biol 19:2958–2966

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ball DK, Surveyor GA, Diehl JR, Steffen CL, Uzumcu M, Mirando MA, Brigstock DR (1998) Characterization of 16- to 20-kilodalton (kDa) connective tissue growth factors (CTGFs) and demonstration of proteolytic activity for 38-kDa CTGF in pig uterine luminal flushings. Biol Reprod 59:828–835

    Article  CAS  PubMed  Google Scholar 

  • Beddy D, Mulsow J, Watson RW, Fitzpatrick JM, O’Connell PR (2006) Expression and regulation of connective tissue growth factor by transforming growth factor beta and tumour necrosis factor alpha in fibroblasts isolated from strictures in patients with Crohn’s disease. Br J Surg 93:1290–1296

    Article  CAS  PubMed  Google Scholar 

  • Blom IE, van Dijk AJ, Wieten L, Duran K, Ito Y, Kleij L, deNichilo M, Rabelink TJ, Weening JJ, Aten J, Goldschmeding R (2001) In vitro evidence for differential involvement of CTGF, TGFbeta, and PDGF-BB in mesangial response to injury. Nephrol Dial Transplant 16:1139–1148

    Article  CAS  PubMed  Google Scholar 

  • Bonniaud P, Margetts PJ, Kolb M, Haberberger T, Kelly M, Robertson J, Gauldie J (2003) Adenoviral gene transfer of connective tissue growth factor in the lung induces transient fibrosis. Am J Respir Crit Care Med 168:770–778

    Article  PubMed  Google Scholar 

  • Brigstock DR (1999) The connective tissue growth factor/cysteine-rich 61/nephroblastoma overexpressed (CCN) family. Endocr Rev 20:189–206

    CAS  PubMed  Google Scholar 

  • Brigstock DR, Goldschmeding R, Katsube KI, Lam SC, Lau LF, Lyons K, Naus C, Perbal B, Riser B, Takigawa M, Yeger H (2003) Proposal for a unified CCN nomenclature. Mol Pathol 56:127–128

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Brigstock DR, Steffen CL, Kim GY, Vegunta RK, Diehl JR, Harding PA (1997) Purification and characterization of novel heparin-binding growth factors in uterine secretory fluids: identification as heparin-regulated Mr 10,000 forms of connective tissue growth factor. J Biol Chem 272:20275–20282

    Article  CAS  PubMed  Google Scholar 

  • Brunner A, Chinn J, Neubauer M, Purchio AF (1991) Identification of a gene family regulated by transforming growth factor-β. DNA Cell Biol 10:293–300

    Article  CAS  PubMed  Google Scholar 

  • Chen Y, Abraham DJ, Shi WX, Pearson JD, Black CM, Lyons KM, Leask A (2004) CCN2 (connective tissue growth factor) promotes fibroblast adhesion to fibronectin. Mol Biol Cell 15:5635–5646

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dammeier J, Brauchle M, Falk W, Grotendorst GR, Werner S (1998) Connective tissue growth factor: a novel regulator of mucosal repair and fibrosis in inflammatory bowel disease? Int J Biochem Cell Biol 30:909–922

    Article  CAS  PubMed  Google Scholar 

  • Dean RA, Butler GS, Yamina HK, Delbé J, Brigstock DR, Courty J, Overal CM (2007) Identification of candidate angiogenic inhibitors processed by matrix metalloproteinase 2 (MMP-2) in cell-based proteomic screens: disruption of vascular endothelial growth factor (VEGF)/heparin affin regulatory peptide (pleiotrophin) and VEGF/connective tissue growth factor angiogenic inhibitory complexes by MMP-2 proteolysis. Mol Cell Biol 27:8454–8465

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fernando CA, Conrad PA, Bartels CF, Marques T, To M, Balow SA, Nakamura Y, Warman ML (2010) Temporal and spatial expression of CCN genes in zebrafish. J Developmental Dynamics 239(6):1755–1767

  • Gao R, Brigstock DR (2004) Connective tissue growth factor (CCN2) induces adhesion of rat activated hepatic stellate cells by binding of its C-terminal domain to integrin alpha (v) beta (3) and heparan sulfate proteoglycan. J Biol Chem 279:8848–8855

    Article  CAS  PubMed  Google Scholar 

  • Grotendorst GR, Okochi H, Hayashi N (1996) A novel transforming growth factor β response element controls the expression of the connective tissue growth factor gene. Cell Growth Differ 7:469–480

    CAS  PubMed  Google Scholar 

  • Hashimoto G, Inoki I, Fujii Y, Aoki T, Ikeda E, Okada Y (2002) Matrix metalloproteinases cleave connective tissue growth factor and reactivate angiogenic activity of vascular endothelial growth factor 165. J Biol Chem 277:36288–36295

    Article  CAS  PubMed  Google Scholar 

  • Igarashi A, Nashiro K, Kikuchi K, Sato S, Ihn H, Grotendorst GR, Takehara K (1995) Significant correlation between connective tissue growth factor gene expression and skin sclerosis in tissue sections from patients with systemic sclerosis. J Invest Dermatol 105:280–284

    Article  CAS  PubMed  Google Scholar 

  • Igarashi A, Okochi H, Bradham DM, Grotendorst GR (1993) Regulation of connective tissue growth factor gene expression in human skin fibroblasts and during wound repair. Mol Biol Cell 4:637–645

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ihn H (2002) Pathogenesis of fibrosis: role of TGF-beta and CTGF. Curr Opin Rheumatol 14:681–685

    Article  CAS  PubMed  Google Scholar 

  • Ivkovic S, Yoon BS, Popoff SN, Safadi FF, Libuda de Stephenson RC, Daluiski A, Lyons KM (2003) Connective tissue growth factor coordinates chondrogenesis and angiogenesis during skeletal development. Development 130:2779–2791

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kong XH, Wang ZX, Gan XN, Li JB, He SP (2008) Molecular evolution of connective tissue growth factor of Cyprinidae (Teleostei: Cypriniformes). Prog Nat Sci 18:155–160

    Article  CAS  Google Scholar 

  • Lasky JA, Ortiz LA, Tonthat B, Hoyle GW, Corti M, Athas G, Lungarella G, Brody A, Friedman M (1998) Connective tissue growth factor mRNA expression is upregulated in bleomycin-induced lung fibrosis. Am J Phys 275:L365–L371

    CAS  Google Scholar 

  • Lau LF, Lam SC (1999) The CCN family of angiogenic regulators: the integrin connection. Exp Cell Res 248:44–57

    Article  CAS  PubMed  Google Scholar 

  • Leask A, Abraham DJ (2006) All in the CCN family: essential matricellular signaling modulators emerge from the bunker. J Cell Sci 119(Pt23):4803–4810

    Article  CAS  PubMed  Google Scholar 

  • Li G, Xie Q, Shi Y, Li D, Zhang M, Jiang S, Zhou H, Lu H, Jin Y (2006) Inhibition of connective tissue growth factor by siRNA prevents liver fibrosis in rats. J Gene Med 8:889–900

    Article  CAS  PubMed  Google Scholar 

  • Mercurio S, Latinkic B, Itasaki N, Krumlauf R, Smith JC (2004) Connective-tissue growth factor modulates WNT signalling and interacts with the WNT receptor complex. Development 131(9):2137–2147

  • Meyer A, Peer YV (2005) From 2R to 3R: evidence for a fish-specific genome duplication (FSGD). Bioessays 27(9):937–945

  • Meyer A, Schartl M (1999) Gene and genome duplications in vertebrates: the one-to-four (-to-eight in fish) rule and the evolution of novel gene functions. Curr Op Cell Biol 11:699–704

    Article  CAS  PubMed  Google Scholar 

  • Moussad EE, Brigstock DR (2000) Connective tissue growth factor: what’s in a name? Mol Genet Metab 71:276–292

    Article  CAS  PubMed  Google Scholar 

  • Mori Y, Hinchcliff M, Wu M, Warner BM, M Lyons K, Varga J (2008) Connective tissue growth factor/CCN2-null mouse embryonic fibroblasts retain intact transforming growth factor-beta responsiveness. Exp Cell Res 314:1094–1104

    Article  CAS  PubMed  Google Scholar 

  • Mori T, Kawara S, Shinozaki M, Hayashi N, Kakinuma T, Igarashi A, Takigawa M, Nakanishi T, Takehara K (1999) Role and interaction of connective tissue growth factor with transforming growth factorbeta in persistent fibrosis: a mouse fibrosis model. J Cell Physiol 181:153–159

    Article  CAS  PubMed  Google Scholar 

  • Nakanishi T, Nishida T, Shimo T, Kobayashi K, Kubo T, Tamatani T, Tezuka K, Takigawa M (2000) Effects of CTGF/Hcs24, a product of a hypertrophic chondrocyte-specific gene, on the proliferation and differentiation of chondrocytes in culture. Endocrinology 141:264–273

    Article  CAS  PubMed  Google Scholar 

  • Nakanishi T, Yamaai T, Asano M, Nawachi K, Suzuki M, Sugimoto T, Takigawa M (2001) Overexpression of connective tissue growth factor/hypertrophic chondrocyte-specific gene product 24 decreases bone density in adult mice and induces dwarfism. Biochem Biophys Res Commun 281:678–681

    Article  CAS  PubMed  Google Scholar 

  • Nishida T, Nakanishi T, Asano M, Shimo T, Takigawa M (2000) Effects of CTGF/Hcs24, a hypertrophic chondrocyte-specific gene product, on the proliferation and differentiation of osteoblastic cells in vitro. J Cell Physiol 184:197–206

    Article  CAS  PubMed  Google Scholar 

  • Perbal B, Takigawa M (2005) CCN proteins: a new family of cell growth and differentiation regulators. In: Hackensack. Imperial College Press, London

    Google Scholar 

  • Shimo T, Kubota S, Yoshioka N, Ibaragi S, Isowa S, Eguchi T, Sasaki A, Takigawa M (2006) Pathogenic role of connective tissue growth factor (CTGF/CCN2) in osteolytic metastasis of breast cancer. J Bone Miner Res 21:1045–1059

    Article  CAS  PubMed  Google Scholar 

  • Shimo T, Nakanishi T, Kimura Y, Nishida T, Ishizeki K, Matsumura T, Takigawa M (1998) Inhibition of endogenous expression of connective tissue growth factor by its antisense oligonucleotide and antisense RNA suppresses proliferation and migration of vascular endothelial cells. J Biochem (Tokyo) 124:130–140

    Article  CAS  Google Scholar 

  • Sönke F, Heike H, Stephanie C (2005) Gene expression of connective tissue growth factor in adult mouse. J Growth Factors 23:43–53

    Article  CAS  Google Scholar 

  • Su BY, Cai WQ (2002) Progress in the study of CCN family. J First Mil Med Univ 22:179–183

    CAS  Google Scholar 

  • Sun T (2012) Coloning, expression and SNP excavating of CCN family in common carp, PhD thesis, Shanghai Ocean University

  • Takehara K (2003) Hypothesis: pathogenesis of systemic sclerosis. J Rheumatol 30:755–759

    PubMed  Google Scholar 

  • Tamatani T, Kobayashi H, Tezuka K, Sakamoto S, Suzuki K, Nakanishi T, Takigawa M, Miyano T (1998) Establishment of the enzyme-linked immunosorbent assay for connective tissue growth factor (CTGF) and its detection in the sera of biliary atresia. Biochem Biophys Res Commun 251:748–752

    Article  CAS  PubMed  Google Scholar 

  • Vorwerk P, Wex H, Hohmann B, Mohnike K, Schmidt U, Mittler U (2002) Expression of components of the IGF signalling system in child-hood acute lymphoblastic leukaemia. Mol Pathol 55:40–45

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang Y (2014) Molecular cloning of CTGF genes and the study on the construction and transgenic efficiency of PB transposon and PB-Tgf2 transposition system in Megalobrama amblycephala, PhD thesis, Shanghai Ocean University

  • Xie D, Nakachi K, Wang H, Elashoff R, Koeffler HP (2001) Elevated levels of ccn1, ccn2 and ccn4 in primary breast cancers associated with more advanced features. Cancer Res 61:8917–8923

    CAS  PubMed  Google Scholar 

  • Ying Z, Ling ML (1996) Isolation and characterization of xnov, a Xenopus laevis ortholog of the chicken nov gene. Gene 171:243–248

    Article  CAS  PubMed  Google Scholar 

  • Zeng ZJ, Yang LY, Ding X, Wang W (2004) Expressions of cysteine-rich 61, connective tissue growth factor and Nov genes in hepatocellular carcinoma and their clinical significance. World J Gastroenterol 10:3414–3418

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhang Q, Lin Y (2006) The relationship between connective tissue growth factor and vascular remodeling. Int J Respiration 26:220–222

    Google Scholar 

Download references

Funding

This work was supported by the Science and Technology Commission of the Shanghai Municipality (No. 13ZR1419600), the Hydrobiological Project of Shanghai Leading Academic Discipline (No. S30701), and the Shanghai Educational Development Foundation (No. 06KZ002).

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WenQian Pan, XiangJun Leng and XiaoQin Li conceived, designed the experiments, analyzed data and wrote the manuscript together. WenQian Pan, JunPeng Wang, Zhihan Tu, Tian Gan, Jing Hu and Jing Wei collected the samples and performed the experiments.

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Correspondence to Xiang-Jun Leng or Xiao-Qin Li.

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Pan, WQ., Wang, JP., Tu, ZH. et al. Cloning, molecular characterization, and tissue differential expression of connective tissue growth factor (ctgf) of grass carp. Fish Physiol Biochem 45, 1431–1443 (2019). https://doi.org/10.1007/s10695-019-00653-2

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