Jump to ContentJump to Main Navigation
Show Summary Details
More options …

Cellular and Molecular Biology Letters

Editor-in-Chief: /

IMPACT FACTOR 2016: 1.260
5-year IMPACT FACTOR: 1.506

CiteScore 2016: 1.56

SCImago Journal Rank (SJR) 2016: 0.615
Source Normalized Impact per Paper (SNIP) 2016: 0.470

See all formats and pricing
More options …
Volume 19, Issue 4 (Dec 2014)

Dysregulation of gene expression in ABCC6 knockdown HepG2 cells

Rocchina Miglionico / Maria Armentano / Monica Carmosino / Antonella Salvia / Flavia Cuviello / Faustino Bisaccia / Angela Ostuni
Published Online: 2014-12-21 | DOI: https://doi.org/10.2478/s11658-014-0208-2


ABCC6 protein is an ATP-dependent transporter that is mainly found in the basolateral plasma membrane of hepatocytes. ABCC6 deficiency is the primary cause of several forms of ectopic mineralization syndrome. Mutations in the human ABCC6 gene cause pseudoxanthoma elasticum (PXE), an autosomal recessive disease characterized by ectopic calcification of the elastic fibers in dermal, ocular and vascular tissues. Mutations in the mouse ABCC6 gene were also associated with dystrophic cardiac calcification. Reduced levels of ABCC6 protein were found in a β-thalassemic mouse model. Moreover, some cases of generalized arterial calcification in infancy are due to ABCC6 mutations. In order to study the role of ABCC6 in the pathogenesis of ectopic mineralization, the expressions of genes involved in this process were evaluated in HepG2 cells upon stable knockdown of ABCC6 by small hairpin RNA (shRNA) technology. ABCC6 knockdown in HepG2 cells causes a significant upregulation of the genes promoting mineralization, such as TNAP, and a parallel downregulation of genes with anti-mineralization activity, such as NT5E, Fetuin A and Osteopontin. Although the absence of ABCC6 has been already associated with ectopic mineralization syndromes, this study is the first to show a direct relationship between reduced ABCC6 levels and the expression of pro-mineralization genes in hepatocytes.

Keywords: ABCC6; Knockdown; HepG2 cells; Gene expression; Mineralization; TNAP; NT5E; OPN; Fetuin A

  • [1] Boskey, A.L. Biomineralization: conflicts, challenges and opportunities. J. Cell. Biochem. Suppl. 30–31 (1998) 83–91. http://dx.doi.org/10.1002/(SICI)1097-4644(1998)72:30/31+<83::AID-JCB12>3.0.CO;2-FCrossrefGoogle Scholar

  • [2] Addison, W.N., Azari, F., Sørensen, E.S., Kaartinen, M.T. and McKee, M.D. Pyrophosphate inhibits mineralization of osteoblast cultures by binding to mineral, upregulating osteopontin, and inhibiting alkaline phosphatase activity. J. Biol. Chem. 282 (2007) 15872–15883. http://dx.doi.org/10.1074/jbc.M701116200Web of ScienceCrossrefGoogle Scholar

  • [3] Jiang, Q., Li, Q. and Uitto, J. Aberrant mineralization of connective tissues in a mouse model of pseudoxanthoma elasticum: systemic and local regulatory factors. J. Invest. Dermatol. 127 (2007) 1392–1402. http://dx.doi.org/10.1038/sj.jid.5700729CrossrefGoogle Scholar

  • [4] Steitz, S.A, Speer, M.Y., McKee, M.D., Liaw, L., Almeida, M., Yang, H. and Giachelli, C.M. Osteopontin inhibits mineral deposition and promotes regression of ectopic calcification. Am. J. Pathol. 161 (2002) 2035–2046. http://dx.doi.org/10.1016/S0002-9440(10)64482-3CrossrefGoogle Scholar

  • [5] Le Saux, O., Martin, L., Aherrahrou, Z., Leftheriotis, G., Váradi, A. and Brampton, C.N. The molecular and physiological roles of ABCC6: more than meets the eye. Front. Genet. 3 (2012) 289. Google Scholar

  • [6] Bergen, A.A., Plomp, A.S., Schuurman, E.J., Terry, S., Breuning, M., Dauwerse, H., Swart, J., Kool, M., van Soest, S., Baas, F., ten Brink, J.B. and de Jong, P.T. Mutations in ABCC6 cause pseudoxanthoma elasticum. Nat. Genet. 25 (2000) 228–231. http://dx.doi.org/10.1038/76109CrossrefGoogle Scholar

  • [7] Fabbri, E., Forni, GL., Guerrini, G. and Borgna-Pignatti, C. Pseudoxanthoma-elasticum-like syndrome and thalassemia: an update. Dermatol. Online J. 15 (2009) 7. Google Scholar

  • [8] Martin, L., Douet, V., VanWart, C.M., Heller, M.B. and Le Saux, O. A mouse model of β-thalassemia shows a liver-specific down-regulation of Abcc6 expression. Am. J. Pathol. 178 (2011) 774–783. http://dx.doi.org/10.1016/j.ajpath.2010.10.004CrossrefGoogle Scholar

  • [9] Ruf, N., Uhlenberg, B., Terkeltaub, R., Nürnberg, P. and Rutsch, F. The mutational spectrum of ENPP1 as arising after the analysis of 23 unrelated patients with generalized arterial calcification of infancy (GACI). Hum. Mutat. 25 (2005) 98. http://dx.doi.org/10.1002/humu.9297CrossrefGoogle Scholar

  • [10] Li, Q., Baker, J., Kowalczyk, J., Jiang, Q., Uitto, J. and Schachner, L. Paediatric pseudoxanthoma elasticum with cardiovascular involvement. Br. J. Dermatol. 169 (2013) 1148–1151. http://dx.doi.org/10.1111/bjd.12462CrossrefGoogle Scholar

  • [11] Nitschke, Y., Baujat, G., Botschen, U., Wittkampf, T., du Moulin, M., Stella, J., Le Merrer, M., Guest, G., Lambot, K., Tazarourte-Pinturier, M.F., Chassaing, N., Roche O., Feenstra, I., Loechner, K., Deshpande., C, Garber, S.J., Chikarmane, R., Steinmann, B., Shahinyan, T., Martorell, L., Davies, J., Smith, W.E., Kahler, S.G., McCulloch, M., Wraige, E., Loidi, L., Höhne, W., Martin, L., Hadj-Rabia, S., Terkeltaub, R. and Rutsch, F. Generalized arterial calcification of infancy and pseudoxanthoma elasticum can be caused by mutations in either ENPP1 or ABCC6. Am. J. Hum. Genet. 90 (2012) 25–39. http://dx.doi.org/10.1016/j.ajhg.2011.11.020Web of ScienceCrossrefGoogle Scholar

  • [12] Rutsch, F., Nitschke, Y. and Terkeltaub, R. Genetics in arterial calcification: pieces of a puzzle and cogs in a wheel. Circ. Res. 109 (2011) 578–592. http://dx.doi.org/10.1161/CIRCRESAHA.111.247965CrossrefWeb of ScienceGoogle Scholar

  • [13] Meng, H., Vera, I., Che, N., Wang, X., Wang, S.S., Ingram-Drake, L., Schadt, E.E., Drake, T.A. and Lusis, A.J. Identification of Abcc6 as the major causal gene for dystrophic cardiac calcification in mice through integrative genomics. Proc. Natl. Acad. Sci. USA 104 (2007) 4530–4535. http://dx.doi.org/10.1073/pnas.0607620104CrossrefGoogle Scholar

  • [14] Aherrahrou, Z., Doehring, L.C., Ehlers, E.M., Liptau, H., Depping, R., Linsel-Nitschke, P., Kaczmarek, P.M., Erdmann, J. and Schunkert, H. An alternative splice variant in Abcc6, the gene causing dystrophic calcification, leads to protein deficiency in C3H/He mice. J. Biol. Chem. 283 (2008) 7608–7615. http://dx.doi.org/10.1074/jbc.M708290200CrossrefWeb of ScienceGoogle Scholar

  • [15] Beck, K., Hayashi, K., Nishiguchi, B., Le Saux, O., Hayashi, M. and Boyd, C.D. The distribution of Abcc6 in normal mouse tissues suggests multiple functions for this ABC transporter. J. Histochem. Cytochem. 51 (2003) 887–902. http://dx.doi.org/10.1177/002215540305100704CrossrefGoogle Scholar

  • [16] Pomozi, V., Le Saux, O., Brampton, C., Apana, A., Iliás, A., Szeri, F., Martin, L., Monostory, K., Paku, S., Sarkadi, B., Szakács, G. and Váradi, A. ABCC6 is a basolateral plasma membrane protein. Circ. Res. 112 (2013) 148–151. http://dx.doi.org/10.1161/CIRCRESAHA.111.300194CrossrefGoogle Scholar

  • [17] Uitto, J., Pulkkinen, L., and Ringpfeil, F. Molecular genetics of pseudoxanthoma elasticum: a metabolic disorder at the environment-genome interface?. Trends Mol. Med. 7 (2001) 13–17. http://dx.doi.org/10.1016/S1471-4914(00)01869-4CrossrefGoogle Scholar

  • [18] de Boussac, H., Ratajewski, M., Sachrajda, I., Köblös, G., Tordai, A., Pulaski, L., Buday, L., Váradi, A. and Arányi, T. The ERK1/2-hepatocyte nuclear factor 4alpha axis regulates human ABCC6 gene expression in hepatocytes. J. Biol. Chem. 285 (2010) 22800–22808. http://dx.doi.org/10.1074/jbc.M110.105593CrossrefGoogle Scholar

  • [19] R Development Core Team. 2008. R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. ISBN 3-900051-07-0 URL http://www.R-project.org Google Scholar

  • [20] Klement, J.F., Matsuzaki, Y., Jiang, Q.J., Terlizzi, J., Choi, H.Y., Fujimoto, N., Li, K., Pulkkinen, L., Birk, D.E., Sundberg, J.P. and Uitto, J. Targeted ablation of the abcc6 gene results in ectopic mineralization of connective tissues. Mol. Cell. Biol. 18 (2005) 8299–8310. http://dx.doi.org/10.1128/MCB.25.18.8299-8310.2005CrossrefGoogle Scholar

  • [21] Gorgels, T.G., Hu, X., Scheffer, G.L., van der Wal, A.C., Toonstra, J., de Jong, P.T., van Kuppevelt, T.H., Levelt, C.N., de Wolf, A., Loves, W.J., Scheper, R.J., Peek, R., and Bergen, A.A. Disruption of Abcc6 in the mouse: novel insight in the pathogenesis of pseudoxanthoma elasticum. Hum. Mol. Genet. 14 (2005) 1763–1773. http://dx.doi.org/10.1093/hmg/ddi183CrossrefGoogle Scholar

  • [22] Ronchetti, I., Boraldi, F., Annovi, G., Cianciulli, P. and Quaglino, D. Fibroblast involvement in soft connective tissue calcification. Front. Genet. 4 (2013) 22. http://dx.doi.org/10.3389/fgene.2013.00022CrossrefGoogle Scholar

  • [23] Orimo, H. The mechanism of mineralization and the role of alkaline phosphatase in health and disease. J. Nippon Med. Sch. 77 (2010) 4–12. http://dx.doi.org/10.1272/jnms.77.4CrossrefGoogle Scholar

  • [24] Yegutkin, G.G. Nucleotide- and nucleoside-converting ectoenzymes: Important modulators of purinergic signaling cascade. Biochem. Biophys. Acta 1783 (2008) 673–694. http://dx.doi.org/10.1016/j.bbamcr.2008.01.024CrossrefGoogle Scholar

  • [25] St Hilaire, C., Ziegler, S.G., Markello, T.C., Brusco, A., Groden, C., Gill, F., Carlson-Donohoe, H., Lederman, R.J., Chen, M.Y., Yang, D., Siegenthaler, M.P., Arduino, C., Mancini, C., Freudenthal, B., Stanescu, H.C., Zdebik, A.A., Chaganti, R.K., Nussbaum, R.L., Kleta, R., Gahl, W.A. and Boehm, M. NT5E mutations and arterial calcifications. N. Engl. J. Med. 364 (2011) 432–442. http://dx.doi.org/10.1056/NEJMoa0912923Google Scholar

  • [26] Szabó, Z., Váradi, A., Li, Q. and Uitto, J. ABCC6 does not transport adenosine-relevance to pathomechanism of pseudoxanthoma elasticum. Mol. Genet. Metab. 104(3) (2011) 421. http://dx.doi.org/10.1016/j.ymgme.2011.07.013CrossrefWeb of ScienceGoogle Scholar

  • [27] Hendig, D., Schulz, V., Arndt, M., Szliska, C., Kleesiek, K. and Götting, C. Role of serum fetuin-A, a major inhibitor of systemic calcification, in pseudoxanthoma elasticum. Clin. Chem. 52 (2006) 227–234. http://dx.doi.org/10.1373/clinchem.2005.059253CrossrefGoogle Scholar

  • [28] Meng, H., Vera, I., Che, N., Wang, X., Wang, S.S., Ingram-Drake, L., Schadt, E.E., Drake, T.A. and Lusis, A.J. Identification of Abcc6 as the major causal gene for dystrophic cardiac calcification in mice through integrative genomics. Proc. Natl. Acad. Sci. USA 104 (2007) 4530–4535. http://dx.doi.org/10.1073/pnas.0607620104CrossrefGoogle Scholar

  • [29] Jansen, R.S., Küçükosmanoglu, A., de Haas, M., Sapthu, S., Andoni Otero, J., Hegman, J.E.M., Bergen, A.A.B., Gorgels, T.G.M.F., Borst, P. and van de Wetering, K. ABCC6 prevents ectopic mineralization seen in pseudoxanthoma elasticum by inducing cellular nucleotide release. Proc. Natl. Acad. Sci. USA 110 (2013) 20206–20211. http://dx.doi.org/10.1073/pnas.1319582110CrossrefGoogle Scholar

About the article

Published Online: 2014-12-21

Published in Print: 2014-12-01

Citation Information: Cellular and Molecular Biology Letters, ISSN (Online) 1689-1392, ISSN (Print) 1425-8153, DOI: https://doi.org/10.2478/s11658-014-0208-2.

Export Citation

© 2014 Versita Warsaw. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License. BY-NC-ND 3.0

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

Toni Schumacher and Ralf A. Benndorf
Molecules, 2017, Volume 22, Number 4, Page 589
Viola Pomozi, Christopher Brampton, Koen van de Wetering, Janna Zoll, Bianca Calio, Kevin Pham, Jesse B. Owens, Joel Marh, Stefan Moisyadi, András Váradi, Ludovic Martin, Carolin Bauer, Jeanette Erdmann, Zouhair Aherrahrou, and Olivier Le Saux
The American Journal of Pathology, 2017, Volume 187, Number 6, Page 1258
Rocchina Miglionico, Angela Ostuni, Maria Francesca Armentano, Luigi Milella, Elvira Crescenzi, Monica Carmosino, and Faustino Bisaccia
Cellular & Molecular Biology Letters, 2017, Volume 22, Number 1
Mie R. Rasmussen, Kirstine L. Nielsen, Mia R. Laursen, Camilla B. Nielsen, Pia Svendsen, Henrik Dimke, Erik I. Christensen, Mogens Johannsen, and Søren K. Moestrup
Journal of Proteome Research, 2016, Volume 15, Number 12, Page 4591
Rocchina Miglionico, Andrea Gerbino, Angela Ostuni, Maria Francesca Armentano, Magnus Monné, Monica Carmosino, and Faustino Bisaccia
Journal of Bioenergetics and Biomembranes, 2016, Volume 48, Number 3, Page 259
Flavia Cuviello, Åsa Tellgren-Roth, Patricia Lara, Frida Ruud Selin, Magnus Monné, Faustino Bisaccia, IngMarie Nilsson, and Angela Ostuni
FEBS Letters, 2015, Volume 589, Number 24PartB, Page 3921
Eva Y. G. De Vilder, Mohammad Jakir Hosen, and Olivier M. Vanakker
BioMed Research International, 2015, Volume 2015, Page 1
Katharina Sies and Thomas Ruzicka
hautnah dermatologie, 2015, Volume 31, Number 4, Page 8
Katharina Sies and Thomas Ruzicka
MMW - Fortschritte der Medizin, 2015, Volume 157, Number 7, Page 60

Comments (0)

Please log in or register to comment.
Log in