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Proc Natl Acad Sci U S A. Jun 25, 1996; 93(13): 6819–6824.
PMCID: PMC39111

Cloning and characterization of cDNAs for matrix metalloproteinases of regenerating newt limbs.

Abstract

Matrix metalloproteinases (MMPs) of regenerating urodele limbs have been suggested to play crucial roles in the process of the dedifferentiation of cells in the damaged tissues and the ensuing blastema formation because the activation of MMPs is an early and conspicuous event occurring in the amputated limb. MMP cDNAs were cloned as products of the reverse transcription-PCR from cDNA libraries of newt limbs, and their structures were characterized. Three cDNAs encoding newt MMPs (2D-1, 2D-19, and 2D-24) have been cloned from second day postamputation regenerating limbs, and a cDNA (EB-1) was cloned from early bud-stage regenerating limbs. These cDNAs included the full-length coding regions. The deduced amino acid sequences of 2D-1, 2D-19, 2D-24, and EB-1 had a homology with mammalian MMP9, MMP3/10, MMP3/10, and MMP13, respectively. The basic motif of these newt MMP genes was similar to mammalian counterparts and contained regions encoding a putative signal sequence, a propeptide, an active site with three zinc-binding histidine residues, a calcium-binding domain, a hemopexin region, and three key cysteine residues. However, some unique molecular evolutionary features were also found in the newt MMPs. cDNAs of 2D-19 and 2D-24 contained a specific insertion and deletion, respectively. The insertion of 2D-19 is threonine-rich, similar to the threonine cluster found in the collagenase-like sea urchin hatching enzyme. Northern blot analysis showed that the expression levels of the newt MMPs were dramatically increased after amputation, suggesting that they play an important role(s) in tissue remodeling of the regenerating limb.

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  • Yoshizato K. Biochemistry and cell biology of amphibian metamorphosis with a special emphasis on the mechanism of removal of larval organs. Int Rev Cytol. 1989;119:97–149. [PubMed]
  • Gailit J, Clark RA. Wound repair in the context of extracellular matrix. Curr Opin Cell Biol. 1994 Oct;6(5):717–725. [PubMed]
  • Hay ED. Extracellular matrix alters epithelial differentiation. Curr Opin Cell Biol. 1993 Dec;5(6):1029–1035. [PubMed]
  • Talhouk RS, Bissell MJ, Werb Z. Coordinated expression of extracellular matrix-degrading proteinases and their inhibitors regulates mammary epithelial function during involution. J Cell Biol. 1992 Sep;118(5):1271–1282. [PMC free article] [PubMed]
  • Grillo HC, Lapière CM, Dresden MH, Gross J. Collagenolytic activity in regenerating forelimbs of the adult newt (Triturus viridescens). Dev Biol. 1968 May;17(5):571–583. [PubMed]
  • Yang EV, Bryant SV. Developmental regulation of a matrix metalloproteinase during regeneration of axolotl appendages. Dev Biol. 1994 Dec;166(2):696–703. [PubMed]
  • Mailman ML, Dresden MH. Collagen metabolism in the regenerating forelimb of Notophthalmus viridescens: synthesis, accumulation, and maturation. Dev Biol. 1976 Jun;50(2):378–394. [PubMed]
  • Gulati AK, Zalewski AA, Reddi AH. An immunofluorescent study of the distribution of fibronectin and laminin during limb regeneration in the adult newt. Dev Biol. 1983 Apr;96(2):355–365. [PubMed]
  • Onda H, Poulin ML, Tassava RA, Chiu IM. Characterization of a newt tenascin cDNA and localization of tenascin mRNA during newt limb regeneration by in situ hybridization. Dev Biol. 1991 Nov;148(1):219–232. [PubMed]
  • Toole BP, Gross J. The extracellular matrix of the regenerating newt limb: synthesis and removal of hyaluronate prior to differentiation. Dev Biol. 1971 May;25(1):57–77. [PubMed]
  • Birkedal-Hansen H. Proteolytic remodeling of extracellular matrix. Curr Opin Cell Biol. 1995 Oct;7(5):728–735. [PubMed]
  • Liu X, Wu H, Byrne M, Jeffrey J, Krane S, Jaenisch R. A targeted mutation at the known collagenase cleavage site in mouse type I collagen impairs tissue remodeling. J Cell Biol. 1995 Jul;130(1):227–237. [PMC free article] [PubMed]
  • Stetler-Stevenson WG, Aznavoorian S, Liotta LA. Tumor cell interactions with the extracellular matrix during invasion and metastasis. Annu Rev Cell Biol. 1993;9:541–573. [PubMed]
  • Oofusa K, Yomori S, Yoshizato K. Regionally and hormonally regulated expression of genes of collagen and collagenase in the anuran larval skin. Int J Dev Biol. 1994 Jun;38(2):345–350. [PubMed]
  • Patterton D, Hayes WP, Shi YB. Transcriptional activation of the matrix metalloproteinase gene stromelysin-3 coincides with thyroid hormone-induced cell death during frog metamorphosis. Dev Biol. 1995 Jan;167(1):252–262. [PubMed]
  • Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. [PubMed]
  • Chirgwin JM, Przybyla AE, MacDonald RJ, Rutter WJ. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. [PubMed]
  • Wilhelm SM, Collier IE, Marmer BL, Eisen AZ, Grant GA, Goldberg GI. SV40-transformed human lung fibroblasts secrete a 92-kDa type IV collagenase which is identical to that secreted by normal human macrophages. J Biol Chem. 1989 Oct 15;264(29):17213–17221. [PubMed]
  • Wilhelm SM, Collier IE, Kronberger A, Eisen AZ, Marmer BL, Grant GA, Bauer EA, Goldberg GI. Human skin fibroblast stromelysin: structure, glycosylation, substrate specificity, and differential expression in normal and tumorigenic cells. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6725–6729. [PMC free article] [PubMed]
  • Muller D, Quantin B, Gesnel MC, Millon-Collard R, Abecassis J, Breathnach R. The collagenase gene family in humans consists of at least four members. Biochem J. 1988 Jul 1;253(1):187–192. [PMC free article] [PubMed]
  • Freije JM, Díez-Itza I, Balbín M, Sánchez LM, Blasco R, Tolivia J, López-Otín C. Molecular cloning and expression of collagenase-3, a novel human matrix metalloproteinase produced by breast carcinomas. J Biol Chem. 1994 Jun 17;269(24):16766–16773. [PubMed]
  • Aimes RT, French DL, Quigley JP. Cloning of a 72 kDa matrix metalloproteinase (gelatinase) from chicken embryo fibroblasts using gene family PCR: expression of the gelatinase increases upon malignant transformation. Biochem J. 1994 Jun 15;300(Pt 3):729–736. [PMC free article] [PubMed]
  • Takino T, Sato H, Yamamoto E, Seiki M. Cloning of a human gene potentially encoding a novel matrix metalloproteinase having a C-terminal transmembrane domain. Gene. 1995 Apr 3;155(2):293–298. [PubMed]
  • Sanchez-Lopez R, Nicholson R, Gesnel MC, Matrisian LM, Breathnach R. Structure-function relationships in the collagenase family member transin. J Biol Chem. 1988 Aug 25;263(24):11892–11899. [PubMed]
  • Van Wart HE, Birkedal-Hansen H. The cysteine switch: a principle of regulation of metalloproteinase activity with potential applicability to the entire matrix metalloproteinase gene family. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5578–5582. [PMC free article] [PubMed]
  • Hirose T, Patterson C, Pourmotabbed T, Mainardi CL, Hasty KA. Structure-function relationship of human neutrophil collagenase: identification of regions responsible for substrate specificity and general proteinase activity. Proc Natl Acad Sci U S A. 1993 Apr 1;90(7):2569–2573. [PMC free article] [PubMed]
  • Lepage T, Gache C. Early expression of a collagenase-like hatching enzyme gene in the sea urchin embryo. EMBO J. 1990 Sep;9(9):3003–3012. [PMC free article] [PubMed]
  • Garfinkel MD, Pruitt RE, Meyerowitz EM. DNA sequences, gene regulation and modular protein evolution in the Drosophila 68C glue gene cluster. J Mol Biol. 1983 Aug 25;168(4):765–789. [PubMed]
  • Whitham SE, Murphy G, Angel P, Rahmsdorf HJ, Smith BJ, Lyons A, Harris TJ, Reynolds JJ, Herrlich P, Docherty AJ. Comparison of human stromelysin and collagenase by cloning and sequence analysis. Biochem J. 1986 Dec 15;240(3):913–916. [PMC free article] [PubMed]
  • Hammani K, Henriet P, Eeckhout Y. Cloning and sequencing of a cDNA encoding mouse stromelysin 1. Gene. 1992 Oct 21;120(2):321–322. [PubMed]
  • Matrisian LM, Glaichenhaus N, Gesnel MC, Breathnach R. Epidermal growth factor and oncogenes induce transcription of the same cellular mRNA in rat fibroblasts. EMBO J. 1985 Jun;4(6):1435–1440. [PMC free article] [PubMed]
  • Breathnach R, Matrisian LM, Gesnel MC, Staub A, Leroy P. Sequences coding for part of oncogene-induced transin are highly conserved in a related rat gene. Nucleic Acids Res. 1987 Feb 11;15(3):1139–1151. [PMC free article] [PubMed]
  • Murphy GJ, Murphy G, Reynolds JJ. The origin of matrix metalloproteinases and their familial relationships. FEBS Lett. 1991 Sep 2;289(1):4–7. [PubMed]
  • Windsor LJ, Grenett H, Birkedal-Hansen B, Bodden MK, Engler JA, Birkedal-Hansen H. Cell type-specific regulation of SL-1 and SL-2 genes. Induction of the SL-2 gene but not the SL-1 gene by human keratinocytes in response to cytokines and phorbolesters. J Biol Chem. 1993 Aug 15;268(23):17341–17347. [PubMed]

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