Matrix Gla Protein Gene Research Articles

Basic fibroblast growth factor as a selective inducer of matrix Gla protein gene expression in proliferative chondrocytes.

Matrix Gla protein (MGP) is a member of the vitamin K-dependent gamma carboxylase protein family expressed in cartilage. Insulin-like growth factor I (IGF1) stimulates chondrocyte differentiation, whereas basic fibroblast growth factor (FGF2) acts in an opposite manner. We explored the differential expression and regulation by IGF1 and FGF2 of the MGP gene during chondrocyte differentiation. We used a primary culture system of rabbit epiphyseal chondrocytes to show that MGP mRNA is mainly expressed during serum-induced proliferation. Much lower MGP mRNA content is observed in post-mitotic chondrocytes, which newly express alpha 1X procollagen mRNA, a marker of late-differentiated cells. From studies of a series of growth factors, it was shown that IGF1 decreased chondrocyte MGP transcripts, whereas FGF2 had the opposite effect. FGF2 stimulated chondrocyte MGP production in a dose- and time-dependent manner at the mRNA and protein levels. FGF2 acted in a dose- and time-dependent manner, reaching a maximum at 10 ng/ml at 20 h. The protein synthesis inhibitor cycloheximide did not modify FGF2 action, in agreement with a direct effect. Actinomycin D abolished FGF2-induced stimulation, strongly suggesting that FGF2 modulated MGP gene transcription. We transiently transfected chondrocytes with a construct containing the mouse MGP promoter from -5000 to -168 base pairs, relative to the transcription start site of the gene linked to the luciferase gene (MGP-Luc). In transfected cells, FGF2 stimulated luciferase activity up to sevenfold while IGF1 had no effect. Hence, FGF2 induces transcription of the MGP gene via the 5'-flanking region of the gene. Using a series of deleted MGP-Luc constructs, we identified a sequence of 748 base pairs which was sufficient for transcriptional activation by FGF2. These results led us to postulate that the inhibitory chondrogenic action of FGF2 involves a mechanism whereby MGP gene transcription and protein are induced.

Molecular cloning of the Matrix Gla Protein gene from Xenopus laevis. Functional analysis of the promoter identifies a calcium sensitive region required for basal activity

To analyze the regulation of Matrix Gla Protein (MGP) gene expression in Xenopus laevis, we cloned the xMGP gene and its 5′ region, determined their molecular organization, and characterized the transcriptional properties of the core promoter. The Xenopus MGP (xMGP) gene is organized into five exons, one more as its mammalian counterparts. The first two exons in the Xenopus gene encode the DNA sequence that corresponds to the first exon in mammals whereas the last three exons show homologous organization in the Xenopus MGP gene and in the mammalian orthologs. We characterized the transcriptional regulation of the xMGP gene in transient transfections using Xenopus A6 cells. In our assay system the identified promoter was shown to be transcriptionally active, resulting in a 12-fold induction of reporter gene expression. Deletional analysis of the 5′ end of the xMGP promoter revealed a minimal activating element in the sequence from −70 to −36 bp. Synthetic reporter constructs containing three copies of the defined regulatory element delivered 400-fold superactivation, demonstrating its potential for the recruitment of transcriptional activators. In gel mobility shift assays we demonstrate binding of X. laevis nuclear factors to an extended regulatory element from −180 to −36, the specificity of the interaction was proven in competition experiments using different fragments of the xMGP promoter. By this approach the major site of factor binding was demonstrated to be included in the minimal activating promoter fragment from −70 to −36 bp. In addition, in transient transfection experiments we could show that this element mediates calcium dependent transcription and increasing concentrations of extracellular calcium lead to a significant dose dependent activation of reporter gene expression.

Molecular cloning of the Matrix Gla Protein gene from Xenopus laevis. Functional analysis of the promoter identifies a calcium sensitive region required for basal activity.

To analyze the regulation of Matrix Gla Protein (MGP) gene expression in Xenopus laevis, we cloned the xMGP gene and its 5' region, determined their molecular organization, and characterized the transcriptional properties of the core promoter. The Xenopus MGP (xMGP) gene is organized into five exons, one more as its mammalian counterparts. The first two exons in the Xenopus gene encode the DNA sequence that corresponds to the first exon in mammals whereas the last three exons show homologous organization in the Xenopus MGP gene and in the mammalian orthologs. We characterized the transcriptional regulation of the xMGP gene in transient transfections using Xenopus A6 cells. In our assay system the identified promoter was shown to be transcriptionally active, resulting in a 12-fold induction of reporter gene expression. Deletional analysis of the 5' end of the xMGP promoter revealed a minimal activating element in the sequence from -70 to -36 bp. Synthetic reporter constructs containing three copies of the defined regulatory element delivered 400-fold superactivation, demonstrating its potential for the recruitment of transcriptional activators. In gel mobility shift assays we demonstrate binding of X. laevis nuclear factors to an extended regulatory element from -180 to -36, the specificity of the interaction was proven in competition experiments using different fragments of the xMGP promoter. By this approach the major site of factor binding was demonstrated to be included in the minimal activating promoter fragment from -70 to -36 bp. In addition, in transient transfection experiments we could show that this element mediates calcium dependent transcription and increasing concentrations of extracellular calcium lead to a significant dose dependent activation of reporter gene expression.

Down-regulation of matrix Gla protein messenger RNA in human colorectal adenocarcinomas

Matrix Gla protein (MGP) is a vitamin K-dependent extracellular matrix protein commonly found in a variety of tissues. In this study, we describe the potential use of MGP gene expression as the tumor marker of colorectal cancer. A decrease in expression of the MGP gene was also discovered in colorectal cancer using differential screening of cDNA libraries. The MGP expression in 80 human colorectal adenocarcinomas was quantified by a Northern blot analysis to better define the expression pattern of MGP in colorectal cancer. The expression of MGP mRNA was reduced in 63 of 80 (79%) colorectal adenocarcinomas ( P<0.001) as compared to the mRNA in adjacent normal tissue, implying that a decrease in MGP expression is associated with colorectal cancer development. The proportion of tumors with downregulated expression of MGP was lower in Duke's A/B than Duke's C/D (34 of 47 versus 26 of 33, respectively) tumors and was lower in moderate differentiation than poor differentiation (44 of 64 versus 16 of 16, respectively). However, χ 2 analysis does not reveal any correlation between a loss of MGP expression and tumor progression or differentiation state. In conclusion, the downregulation of MGP mRNA generally occurs in colorectal adenocarcinomas. Although the role of MGP in cancer development is unknown, the reduced expression of MGP may be used to distinguish the normal colorectal cells from malignant cells.

Polymorphisms of the human matrix gla protein (MGP) gene, vascular calcification, and myocardial infarction.

The matrix Gla protein (MGP) is an important inhibitor of vessel and cartilage calcification that is strongly expressed in human calcified, atherosclerotic plaques and could modulate plaque calcification and coronary heart disease risk. Using a genetic approach, we explored this possibility by identifying polymorphisms of the MGP gene and testing their possible association with myocardial infarction (MI) and plaque calcification. Eight polymorphisms were identified in the coding and 5'-flanking sequences of the MGP gene. All polymorphisms were investigated in 607 patients with MI and 667 control subjects recruited into the ECTIM Study (Etude Cas-Témoins de l'Infarctus du Myocarde) and in 717 healthy individuals with echographically assessed arterial calcification and atherosclerosis who were participating in the AXA Study. In the ECTIM Study, alleles and genotypes were distributed similarly in patients and controls in the whole study group; in only 1 subgroup of subjects defined as being at low risk for MI were the concordant A-7 and Ala 83 alleles more frequent in patients with MI than in controls (P<0.003). In the AXA Study among subjects with femoral atherosclerosis, the same alleles were more common in the presence than the absence of plaque calcification (P<0.025). The other MGP polymorphisms were not associated with any investigated clinical phenotype. Transient transfection experiments with allelic promoter-reporter gene constructs and DNA-protein interaction assays were carried out to assess possible in vitro functionality of the promoter variants detected at positions -814, -138, and -7 relative to the start of transcription. When compared with the -138 T allele, the minor -138 C: allele consistently conferred a reduced promoter activity of -20% (P<0.0001) in rat vascular smooth muscle cells and of -50% (P<0.004) in a human fibroblast cell line, whereas the other polymorphisms, including -7, displayed no evidence of in vitro functionality. We conclude that the A-7 or Ala 83 alleles of the MGP gene may confer an increased risk of plaque calcification and MI; however, the observed relationships are weak or limited to subgroups of patients and therefore need confirmation.

Association of bone mineral density with polymorphism of the human matrix Gla protein locus in elderly women

The contribution of genetic factors has been implicated in the determination of bone mass in twin and family studies. Some of the genes involved would regulate bone metabolism, bone formation, and resorption, all processes that determine bone mass. One candidate gene matrix Gla protein gene (MGP), has been implicated in the pathogenesis of bone loss through a repression of bone formation. To analyze the genetic background for osteoporosis in elderly women, we have investigated a possible association between the CA repeat polymorphism at the human MGP gene locus and bone mineral density (BMD) of radial bone in 460 elderly Japanese women. Genotypes were classified into six groups according to the number of CA repeats present, from 13 to 18 (alleles A1 through A6). BMD was expressed as the adjusted BMD (ADJBMD), which was the body mass index (BMI)- and age-adjusted average BMD. The 214 women who lacked an A2 allele (212 bp, containing 17 repeats of CA) had significantly lower adjusted BMD than the participants (n = 246) who possessed an allele of that size (mean +/- SD; 0.303 +/- 0.062 vs 0.315 +/- 0.062 g/cm2; P = 0.0382). This result suggests that genetic variation at the MGP locus is associated with some determinants for BMD in elderly women. Therefore, this locus should serve as one of the genetic markers for osteoporosis.

Matrix GLA protein is a developmental regulator of chondrocyte mineralization and, when constitutively expressed, blocks endochondral and intramembranous ossification in the limb.

Matrix GLA protein (MGP), a gamma-carboxyglutamic acid (GLA)-rich, vitamin K-dependent and apatite-binding protein, is a regulator of hypertrophic cartilage mineralization during development. However, MGP is produced by both hypertrophic and immature chondrocytes, suggesting that MGP's role in mineralization is cell stage-dependent, and that MGP may have other roles in immature cells. It is also unclear whether MGP regulates the quantity of mineral or mineral nature and quality as well. To address these issues, we determined the effects of manipulations of MGP synthesis and expression in (a) immature and hypertrophic chondrocyte cultures and (b) the chick limb bud in vivo. The two chondrocyte cultures displayed comparable levels of MGP gene expression. Yet, treatment with warfarin, a gamma-carboxylase inhibitor and vitamin K antagonist, triggered mineralization in hypertrophic but not immature cultures. Warfarin effects on mineralization were highly selective, were accompanied by no appreciable changes in MGP expression, alkaline phosphatase activity, or cell number, and were counteracted by vitamin K cotreatment. Scanning electron microscopy, x-ray microanalysis, and Fourier-transform infrared spectroscopy revealed that mineral forming in control and warfarin-treated hypertrophic cell cultures was similar and represented stoichiometric apatite. Virally driven MGP overexpression in cultured chondrocytes greatly decreased mineralization. Surprisingly, MGP overexpression in the developing limb not only inhibited cartilage mineralization, but also delayed chondrocyte maturation and blocked endochondral ossification and formation of a diaphyseal intramembranous bone collar. The results show that MGP is a powerful but developmentally regulated inhibitor of cartilage mineralization, controls mineral quantity but not type, and appears to have a previously unsuspected role in regulating chondrocyte maturation and ossification processes.

Different Effects of Bone Morphogenetic Proteins 2, 4, 12, and 13 on the Expression of Cartilage and Bone Markers in the MC615 Chondrocyte Cell Line

In order to study the lineage leading to chondrocyte and osteoblast phenotype in vertebrate development, we examined the effect of recombinant human bone morphogenetic protein (BMP)-2, BMP-4, BMP-12 [or growth and differentiation factor (GDF)-7], and BMP-13 (or GDF-6) on the phenotypic expression of the mouse chondrocyte cell line MC615, grown for 1 or 2 weeks in monolayer. Protein synthesis rates were monitored after incubation with [14C]proline. BMP-2 and BMP-4 increased protein synthesis, in agreement with our observation by phase-contrast microscopy of a highly refractile matrix around MC615 cells treated with BMP-2 and -4. Markers of the chondrocytic and osteoblastic differentiation were analyzed at mRNA level. Expression of the type II collagen gene, a marker of the cartilage phenotype, was up-regulated in the presence of low concentration of BMP-2 or -4 (50 ng/ml) and down-regulated at higher concentrations (100–400 ng/ml). In parallel, this expression was stable in the presence of BMP-12 or -13 in the dose range tested (50–400 ng/ml). Expression of the matrix Gla protein (MGP) gene, another marker of cartilage, was also reduced in the presence of 100 ng/ml BMP-2 or -4, while it remained stable in the presence of BMP-12 or -13 at the same concentration. In contrast, expression of the bone Gla protein (BGP) gene, or osteocalcin, a marker of the bone phenotype, was induced when the cells were treated with BMP-2 or -4 but was not detected when the cells were treated with BMP-12 or -13. At the same time, BMP-2 or -4 markedly up-regulated expression of type X collagen mRNA, indicating that MC615 cells possess the ability to express traits associated with endochondral ossification, when exposed to specific BMPs. Furthermore, detailed analysis of type II collagen expression showed that the alternatively spliced transcript collagen IIB, specific for cartilage, is expressed concomitantly with BGP. Therefore, MC615 chondrocytes can simultaneously express chondrocytic and osteoblastic markers, in response to BMP-2 or -4, but show minimal response to BMP-12 (or GDF-7) or to BMP-13 (or GDF-6). These results raise the possibility that chondrocytes in vivo can express osteoblastic properties, provided they are induced by BMP-2 or -4.

Expression of matrix Gla protein (MGP) in an in vitro model of vascular calcification

To investigate the role of matrix Gla protein (MGP), which can bind mineral ions through γ-carboxylated glutamic acid residues, in vascular calcification, we examined the expression of MGP in an in vitro calcification model by using bovine vascular smooth muscle cells (BVSMC). The expression of MGP mRNA was decreased during BVSMC calcification and its levels were inversely correlated with the quantities of BVSMC calcification. MGP mRNA expression was restored to the level of uncalcified control by inhibiting BVSMC calcification with bisphosphonates. These data suggest that the expression of MGP gene is modulated in the development of vascular calcification.

Gene expression of noncollagenous bone matrix proteins in the limb joints and intervertebral disks of the twy mouse.

The twy (tiptoe walking Yoshimura) mouse is an autosomal recessive mutant manifesting multiple osteochondral lesions characterized by pathologic calcium deposition. To elucidate the pathophysiology of the limb joint lesions and the intervertebral disk lesions of the twy mouse, we assessed the mRNA expression of noncollagenous bone matrix proteins such as osteocalcin, osteonectin, osteopontin, and matrix Gla protein (MGP) by in situ hybridization, but only expression of MGP was observed in association with the pathologic calcium deposits in twy mice. Mild degeneration and abnormal growth of the cartilage in contact with the joint capsule was observed at 5 weeks in the articular cartilage of the ankle joint of the twy mouse, and MGP gene expression was observed at the same time. Simultaneous growth of synovial membrane cells and relatively undifferentiated articular cartilage cells in the knee joint, and of cartilage-like cells near the insertion of the cruciate ligament was observed in the twy mouse, and MGP gene expression was found to be present at the same time. Hypertrophy of abnormally proliferated chondrocyte-like cells, which are different from fibrocartilaginous cells of the annulus fibrosus, was observed in the intervertebral disks of the twy mouse at 3 weeks of age, and MGP gene expression was noted at the same time. These findings suggest that abnormal expression of MGP plays a major role in the pathologic calcification of the twy mouse.

Isolation and radiation hybrid mapping of dinucleotide repeat polymorphism at the human matrix Gla protein (MGP) locus.

Matrix Gla protein (MGP) is an 84-residue, vitamin K-dependent protein expressed by chondrocytes and vascular smooth muscle cells, and is a potent regulator of calcium deposition in cartilage and arterial wall. We isolated a polymorphic dinucleotide CA repeat marker from a genomic clone containing the human MGP gene. This polymorphism will be useful in genetic studies of arteriosclerosis and osteoporosis.

Set of optimized luciferase reporter gene plasmids compatible with widely used CAT vectors.

BioTechniquesVol. 23, No. 3 BenchmarksOpen AccessSet of Optimized Luciferase Reporter Gene Plasmids Compatible with Widely Used CAT VectorsJoachim Altschmied & Jutta DuschlJoachim Altschmied*Address correspondence to Joachim Altschmied, Department of Physiological Chemistry I, Biocenter (Theodor Boveri Institute), University of Würzburg, Am Hubland, D-97074 Würzburg, Germany. Internet: E-mail Address: yogi@biozentrum.uni-wuerzburg.deUniversity of Würzburg, Würzburg, GermanySearch for more papers by this author & Jutta DuschlUniversity of Würzburg, Würzburg, GermanySearch for more papers by this authorPublished Online:7 Aug 2018https://doi.org/10.2144/97233bm19AboutSectionsPDF/EPUB ToolsAdd to favoritesDownload CitationsTrack Citations ShareShare onFacebookTwitterLinkedInRedditEmail FiguresReferencesRelatedDetailsCited ByThe transcriptional coactivator Bob1 promotes the development of follicular T helper cells via Bcl68 March 2016 | The EMBO Journal, Vol. 35, No. 8Transcriptional control analyses of the Xiphophorus melanoma oncogeneComparative Biochemistry and Physiology Part C: Toxicology & Pharmacology, Vol. 178Characterization of sea bass FSHβ 5′ flanking region: transcriptional control by 17β-estradiol23 November 2013 | Fish Physiology and Biochemistry, Vol. 40, No. 3Somatic gene transfer in European sea bass (Dicentrarchus labrax): Optimal fish conditions and exogenous control of foreign genesAquaculture, Vol. 388-391Mitf is a transcriptional activator of medaka germ genes in cultureBiochimie, Vol. 94, No. 3Fusion Gene Vectors Allowing for Simultaneous Drug Selection, Cell Labeling, and Reporter Assay in Vitro and in Vivo23 December 2011 | Analytical Chemistry, Vol. 84, No. 2The transcription factor Grainyhead like 3 (GRHL3) affects endothelial cell apoptosis and migration in a NO-dependent mannerBiochemical and Biophysical Research Communications, Vol. 412, No. 4Transcriptional Rewiring of the Sex Determining dmrt1 Gene Duplicate by Transposable Elements12 February 2010 | PLoS Genetics, Vol. 6, No. 2Reporter Gene27 November 2009Wild-type p53 and p73 negatively regulate expression of proliferation related genes5 November 2007 | Oncogene, Vol. 27, No. 18Transcriptional Activities of the Sleeping Beauty Transposon and Shielding Its Genetic Cargo With InsulatorsMolecular Therapy, Vol. 16, No. 2Identification of a distal tandem STAT6 element within the CCL17 locusHuman Immunology, Vol. 68, No. 12The Ancient mariner Sails Again: Transposition of the Human Hsmar1 Element by a Reconstructed Transposase and Activities of the SETMAR Protein on Transposon EndsMolecular and Cellular Biology, Vol. 27, No. 12Bacteria Displaying Interleukin-4 Mutants Stimulate Mammalian Cells and Reflect the Biological Activities of Variant Soluble Cytokines26 May 2004 | ChemBioChem, Vol. 5, No. 6Sesquiterpene lactones inhibit luciferase but not β-galactosidase activity in vitro and ex vivoAnalytical Biochemistry, Vol. 328, No. 2MITF-M plays an essential role in transcriptional activation and signal transduction in Xiphophorus melanomaGene, Vol. 320The diesel exhaust component pyrene induces expression of IL-8 but not of eotaxinInternational Immunopharmacology, Vol. 3, No. 10-11Protein Kinase A Regulates GATA-3-Dependent Activation of IL-5 Gene Expression in Th2 Cells15 March 2003 | The Journal of Immunology, Vol. 170, No. 6The CD23b promoter is a target for NF-AT transcription factors in B-CLL cellsBiochimica et Biophysica Acta (BBA) - Molecular Basis of Disease, Vol. 1588, No. 1The LIM-only coactivator FHL2 modulates WT1 transcriptional activity during gonadal differentiationBiochimica et Biophysica Acta (BBA) - Gene Structure and Expression, Vol. 1577, No. 1β-Naphthoflavone, an Inducer of Xenobiotic Metabolizing Enzymes, Inhibits Firefly Luciferase ActivityAnalytical Biochemistry, Vol. 304, No. 1Molecular cloning of the Matrix Gla Protein gene from Xenopus laevis3 October 2003 | European Journal of Biochemistry, Vol. 269, No. 7Development of an Androgen Reporter Gene Assay (AR-LUX) Utilizing a Human Cell Line with an Endogenously Regulated Androgen ReceptorAnalytical Biochemistry, Vol. 298, No. 1IL-9 and IL-13 Production by Activated Mast Cells Is Strongly Enhanced in the Presence of Lipopolysaccharide: NF-κB Is Decisively Involved in the Expression of IL-91 April 2001 | The Journal of Immunology, Vol. 166, No. 7Cellular Response to Oncogenic Ras Involves Induction of the Cdk4 and Cdk6 Inhibitor p15 INK4bMolecular and Cellular Biology, Vol. 20, No. 8The environmental pollutant pyrene induces the production of IL-4Journal of Allergy and Clinical Immunology, Vol. 105, No. 4Long chain arginine esters: A new class of cationic detergents for preparation of hydrophobic ion-paired complexesBiochemistry and Cell Biology, Vol. 78, No. 1Evaluation of eukaryotic promoters for the construction of DNA vaccines for aquacultureGenetic Analysis: Biomolecular Engineering, Vol. 15, No. 3-5Hypermethylation of the cell cycle inhibitor p15INK4b 3′-untranslated region interferes with its transcriptional regulation in primary lymphomas20 January 1999 | Oncogene, Vol. 18, No. 2 Vol. 23, No. 3 Follow us on social media for the latest updates Metrics Downloaded 127 times History Published online 7 August 2018 Published in print September 1997 Information© 2018 Author(s)PDF download

Matrix Gla protein gene expression is induced by transforming growth factor-beta in embryonic lung culture.

Matrix Gla protein (MGP) is a vitamin K-dependent extracellular matrix protein with a wide tissue distribution. The current study was designed to investigate the possible regulation of MGP by exogenous transforming growth factor-beta (TGF-beta) during lung development. Using reverse transcription coupled competitive polymerase chain reaction methodology, we determined that exogenous TGF-beta 1 increases MGP mRNA levels in embryonic mouse lung culture in a concentration-dependent manner. MGP mRNA levels were elevated by 5.0-fold at 50 ng/ml TGF-beta 1 in E11 embryonic mouse lungs explanted for 4 days in serumless culture. MGP mRNA stimulation by TGF-beta 1 was a time-dependent event: MGP message increment was initially detected after 1 day in culture, and MGP mRNA levels continued to increase up to 4 days in the presence of TGF-beta 1. In addition, embryonic lungs in serumless medium without exogenously added TGF-beta 1 showed an increase, although to a lesser extent, in pulmonary MGP mRNA level during culture, indicating that MGP expression is also developmentally regulated. The present results indicate that MGP gene expression can be stimulated by exogenous TGF-beta 1 during early embryonic mouse lung branching morphogenesis in culture.

Identification of a novel negative retinoic acid responsive element in the promoter of the human matrix Gla protein gene.

The vitamin K-dependent matrix Gla protein (MGP) is synthesized in a wide variety of tissues such as lung, heart, kidney, cartilage, and bone. Expression of the MGP gene is regulated by various growth factors, steroid hormones, and the vitamin A metabolite retinoic acid (RA). In this report, we present evidence that RA down-regulates MGP gene expression in different rat and human cell lines via endogenous retinoid receptors [RA receptor (RAR) and retinoid X receptor (RXR)]. Repression of the human MGP (hMGP) gene is specifically mediated by ligand-activated RAR and RXR. Deletion analysis led to the identification of a novel negative response element (NRE) within the hMGP promoter. DNA binding studies performed with bacterially expressed RAR/RXR reveal the formation of a specific heterodimer/NRE complex. Furthermore, electrophoretic mobility-shift assays performed with proteins from RA-treated cells show that endogenous RAR/RXR binds to the NRE. We demonstrate that the NRE contains a CCAAT box and that both RAR/RXR and CCAAT-binding proteins such as c/EBP beta recognize this common regulatory sequence in the hMGP promoter. Our results indicate that RA-mediated repression of the hMGP gene is due to binding of liganded RAR/RXR to a novel negative RA response element.

Matrix Gla protein gene expression is elevated during postnatal development.

Matrix Gla protein (MGP) is a vitamin K-dependent extracellular matrix protein with a wide tissue distribution. Developmental expression of the MGP gene is characterized by competitive RT-PCR in kidney and calvaria. High levels of MGP mRNA were observed in kidneys and calvaria from 19-day-old embryos to 1-month-old rats. There was a peak in MGP mRNA at 7 days in both tissues. MGP mRNA expression was very low or undetectable in 3-, 5- and 7- month-old kidneys. Similar observations were seen in lung, heart and spleen. However, in connective tissues like calvaria, tibia and trachea, low levels of MGP mRNA are maintained throughout life. Kidney MGP protein was present from birth to 15 days, with the highest MGP protein level at 7 days. Calvarial MGP protein was present throughout development and maturation but peaked at 7 days. The highest MGP protein levels were coincident with peak levels of MGP mRNA. Thus, MGP protein level correlated with mRNA level during rat development. In situ hybridization revealed that MGP staining was most intense in the straight tubules of the developing kidney medulla at 7 days. Staining was absent in stromal cells and in mature nephrons. Taken together, our finding of high MGP mRNA and its intense in situ staining during the postnatal growth phase prove that increased MGP synthesis occurs at a specific time and place during development and maturation.

The matrix Gla protein gene is a marker of the chondrogenesis cell lineage during mouse development

Matrix Gla protein (MGP) is, along with osteocalcin, a skeletal member of the family of extracellular mineral-binding Gla proteins. Although the precise function of these proteins remains obscure, circumstantial evidence suggests that they play a role in endochondral ossification. As a first step toward understanding MGP function we have performed a preliminary characterization of its promoter element and studied the developmental pattern of expression of this gene. DNA transfection experiments indicate that the mouse MGP promoter functions better in cells expressing the MGP gene than in cells that do not express the gene. During mouse development, MGP gene expression is detectable as early as day 10.5 of embryonic development (E10.5), before any skeletal structures are identifiable. In situ hybridization analysis shows that MGP mRNA is initially present at the mesenchymal epithelial interphase in lung and limb buds. As development proceeds, MGP gene is predominantly expressed in cells of the chondrocytic lineage in areas that will undergo endochondral ossification as well as in areas that will remain cartilaginous, such as the trachea and bronchi. In growth plate cartilage, MGP mRNA is present in resting, proliferative, and late hypertrophic chondrocytes. Surprisingly, MGP mRNA is absent from the early hypertrophic chondrocytes and from the osteoblasts. Finally, the MGP gene is expressed at a lower level in kidney medulla and uterus smooth muscle but not in brain, spleen, or heart during development. This study demonstrates that during development MGP gene expression occurs early and is predominant at the epithelial mesenchymal interfaces, principally of lung and limb buds, and in cells of the chondrocytic lineage. This finding raises the intriguing possibility that MGP may play distinct roles during embryogenesis and in the adult organism.

Differential regulation of matrix Gla protein (MGP) gene expression by retinoic acid and estrogen in human breast carcinoma cells

Expression of matrix Gla protein (MGP) gene and its regulation by retinoic acid (RA) and estrogen was investigated in eight human breast cancer cell lines. The promoter region of the MGP gene contains a consensus retinoic acid response element (RARE) and the MGP gene expression has been shown to be strongly induced by RA in other systems. Our results suggest that RA negatively regulates MGP mRNA expression in human breast cancer cells that have high levels of estrogen receptors (ER), i.e. MCF-7, ZR-75 and BT474 and positively regulates its expression in cells with either no ERs, i.e. MDA-MB-468 or very low levels of ERs, i.e. T47D. This indicates that ER levels may affect RA modulation of MGP gene expression in human breast cancer cells. The inhibitory effect of RA on MGP gene expression was abolished in RRO-I, the RA-resistant MCF-7 subline. We also demonstrate for the first time that estrogen strongly induces MGP gene expression in ER-positive cells and that estrogen-mediated induction of MGP is blocked by RA even in otherwise RA-resistant cells.

Retinoic acid increases matrix gla protein in rat plasma

The administration of retinoic acid to rats at dosages which achieve circulating levels of 1 μM or more causes a 40 to 50 percent increase in plasma levels of matrix Gla protein (MGP) within 24 hr. This increase is consistent with previous studies which demonstrate strong induction of MGP gene expression by 1 μM retinoic acid in several cell lines and with the presence of a putative retinoic acid responsive element in the MGP gene promoter. Retinoic acid does increase MGP levels to a far greater extent in cells than in the plasma of rats, but this could reflect partial induction of MGP by endogenous retinoic acid in the vitamin A-replete control animals. The consistent induction of MGP expression in cultured cells and in animals suggests that this small, vitamin K-dependent protein may mediate some of the known actions of retinoic acid in differentiation and development.

Retinoic acid induces matrix Gla protein gene expression in human cells

Retinoic acid induces matrix Gla protein gene expression in human cells

Retinoic acid induces matrix Gla protein gene expression in human cells.

The objective of this study was to investigate the possible regulation of the vitamin K-dependent matrix Gla (gamma-carboxyglutamic acid) protein (MGP) by retinoic acid, a regulation suggested by the recent observation that the human MGP promoter has a perfect direct repeat which is nearly identical to the retinoic acid-responsive element in the retinoic acid receptor-beta gene. We report that retinoic acid strongly increases MGP mRNA levels in all human cells tested, including osteoblasts, articular cartilage chondrocytes, and fibroblasts. In osteoblastic cells, MGP mRNA levels are increased by 25-fold at 1 microM retinoic acid and achieve half-maximal levels at 0.1 microM hormone. MGP is a small secreted protein of unknown function that is synthesized in a wide variety of vertebrate tissues. The present results suggest that part of the known actions of retinoic acid on skin, bone, cartilage, and other tissues in the human may be mediated by the stimulation of MGP synthesis and the consequent effect of increased MGP secretion on nearby target cells.