Abstract
Natural human skin colour is determined both by environmental exposure to ultraviolet light and through inherited genetic variation in a very limited number of genes. Variation of a non-synonymous single-nucleotide polymorphism (nsSNP; rs1426654) in the gene (SLC24A5) encoding the NCKX5 protein is associated with differences in constitutive skin colour in South Asians. The nsSNP encodes the substitution of alanine for threonine at residue 111 (A111T) near a transmembrane region required for exchanger activity, a region which is highly conserved across different species and between NCKX family members. We have shown that NCKX5 is located at the trans-Golgi network of melanocytes and functions as a potassium-dependent sodium-calcium exchanger. When heterologously expressed, the 111T variant of NCKX5 shows significantly lower exchanger activity than the A111 variant. We have postulated that lower exchanger activity causes the reduced melanogenesis and lighter skin in Thr111-positive individuals. We used gene expression microarrays with qPCR replication and validation to assess the impact of siRNA-mediated knockdown of SLC24A5 on the transcriptome of cultured normal human melanocytes (NHM). Very few genes associated with melanogenesis were altered at the transcript level except for MC1R, suggesting that SLC24A5 interacts with at least one well-characterized melanogenic signalling pathway. More surprisingly, the expression of a number of cholesterol homeostatic genes was altered after SLC24A5 knockdown, and the total cholesterol content of NHM was increased. Cholesterol has previously been identified as a potential melanogenic regulator, and our data imply that NCKX5 exchanger function influences natural variation in skin pigmentation via a novel, unknown mechanism affecting cellular sterol levels.
Keywords
These authors contributed equally to the work.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Z.A. Abdel-Malek, M.C. Scott, M. Furumura, M.L. Lamoreux, M. Ollmann, G.S. Barsh, V.J. Hearing, The melanocortin 1 receptor is the principal mediator of the effects of agouti signaling protein on mammalian melanocytes. J. Cell Sci. 114(Pt 5), 1019–1024 (2001)
H.F. Altimimi, P.P. Schnetkamp, Na+-dependent inactivation of the retinal cone/brain Na+/Ca2+-K+ exchanger NCKX2. J. Biol. Chem. 282, 3720–3729 (2007a)
H.F. Altimimi, P.P.M. Schnetkamp, Na+/Ca2+-K+ exchangers (NCKX): functional properties and physiological roles. Channels 1(2), 62–69 (2007b)
K.A. Beaumont, S.N. Shekar, A.L. Cook, D.L. Duffy, R.A. Sturm, Red hair is the null phenotype of MC1R. Hum. Mutat. 29(8), E88–E94 (2008)
P.V. Burgos, C. Klattenhoff, E. de la Fuente, A. Rigotti, A. González, Cholesterol depletion induces PKA-mediated basolateral-to-apical transcytosis of the scavenger receptor class B type I in MDCK cells. Proc. Natl. Acad. Sci. U. S. A. 101(11), 3845–3850 (2004)
Y. Cheli, F. Luciani, M. Khaled, L. Beuret, K. Bille, P. Gounon, J.P. Ortonne, C. Bertolotto, R. Ballotti, αMSH and Cyclic AMP elevating agents control melanosome pH through a protein kinase A-independent mechanism. J. Biol. Chem. 284(28), 18699–18706 (2009)
H. Chen, J. Yang, P.S. Low, J.X. Cheng, Cholesterol level regulates endosome motility via Rab proteins. Biophys. J. 94(4), 1508–1520 (2008)
X. Chen, M.D. Resh, Cholesterol depletion from the plasma membrane triggers ligand-independent activation of the epidermal growth factor receptor. J. Biol. Chem. 277(51), 49631–49637 (2002)
Y. Chen, X.Z. Ruan, Q. Li, A. Huang, J.F. Moorhead, S.H. Powis, Z. Varghese, Inflammatory cytokines disrupt LDL-receptor feedback regulation and cause statin resistance: a comparative study in human hepatic cells and mesangial cells. Am. J. Physiol. Renal Physiol. 293(3), F680–F687 (2007)
A. Chi, J.C. Valencia, Z.Z. Hu, H. Watabe, H. Yamaguchi, N.J. Mangini, H. Huang, V.A. Canfield, K.C. Cheng, F. Yang, R. Abe, S. Yamagishi, J. Shabanowitz, V.J. Hearing, C. Wu, E. Appella, D.F. Hunt, Proteomic and bioinformatic characterization of the biogenesis and function of melanosomes. J. Proteome Res. 5(11), 3135–3144 (2006)
C. Chiaverini, L. Beuret, E. Flori, R. Busca, P. Abbe, K. Bille, P. Bahadoran, J.P. Ortonne, C. Bertolotto, R. Ballotti, Microphthalmia-associated transcription factor regulates RAB27A gene expression and controls melanosome transport. J. Biol. Chem. 283(18), 12635–12642 (2008)
J.A. D’Orazio, T. Nobuhisa, R. Cui, M. Arya, M. Spry, K. Wakamatsu, V. Igras, T. Kunisada, S.R. Granter, E.K. Nishimura, S. Ito, D.E. Fisher, Topical drug rescue strategy and skin protection based on the role of Mc1r in UV-induced tanning. Nature 443(7109), 340–344 (2006)
N.J. Dolman, A.V. Tepikin, Calcium gradients and the Golgi. Cell Calcium 40(5–6), 505–512 (2006)
S. Emr, B.S. Glick, A.D. Linstedt, J. Lippincott-Schwartz, A. Luini, V. Malhotra, B.J. Marsh, A. Nakano, S.R. Pfeffer, C. Rabouille, J.E. Rothman, G. Warren, F.T. Wieland, Journeys through the Golgi – taking stock in a new era. J. Cell Biol. 187(4), 449–453 (2009)
Z. Freyberg, S. Bourgoin, D. Shields, Phospholipase D2 is localized to the rims of the Golgi apparatus in mammalian cells. Mol. Biol. Cell 13(11), 3930–3942 (2002)
R.S. Ginger, S.E. Askew, R.M. Ogborne, S. Wilson, D. Ferdinando, T. Dadd, A.M. Smith, S. Kazi, R.T. Szerencsei, R.J. Winkfein, P.P. Schnetkamp, M.R. Green, SLC24A5 encodes a trans-Golgi network protein with potassium-dependent sodium-calcium exchange activity that regulates human epidermal melanogenesis. J. Biol. Chem. 283(9), 5486–5495 (2008)
A.M. Hall, L. Krishnamoorthy, S.J. Orlow, 25-hydroxycholesterol acts in the Golgi compartment to induce degradation of tyrosinase. Pigment Cell Res. 17(4), 396–406 (2004)
T. Hida, K. Wakamatsu, E.V. Sviderskaya, A.J. Donkin, L. Montoliu, M. Lynn Lamoreux, B. Yu, G.L. Millhauser, S. Ito, G.S. Barsh, K. Jimbow, D.C. Bennett, Agouti protein, mahogunin, and attractin in pheomelanogenesis and melanoblast-like alteration of melanocytes: a cAMP-independent pathway. Pigment Cell Melanoma Res. 22(5), 623–634 (2009)
K.S. Hoek, N.C. Schlegel, O.M. Eichhoff, D.S. Widmer, C. Praetorius, S.O. Einarsson, S. Valgeirsdottir, K. Bergsteinsdottir, A. Schepsky, R. Dummer, E. Steingrimsson, Novel MITF targets identified using a two-step DNA microarray strategy. Pigment Cell Melanoma Res. 21, 665–676 (2008)
M. Hozoji, Y. Munehira, Y. Ikeda, M. Makishima, M. Matsuo, N. Kioka, K. Ueda, Direct interaction of nuclear liver X receptor-beta with ABCA1 modulates cholesterol efflux. J. Biol. Chem. 283(44), 30057–30063 (2008)
International HapMap Consortium, A haplotype map of the human genome. Nature 437(7063), 1299–1320 (2005)
N. Iwamoto, S. Abe-Dohmae, R. Lu, S. Yokoyama, Involvement of protein kinase D in phosphorylation and increase of DNA binding of activator protein 2 alpha to downregulate ATP-binding cassette transporter A1. Arterioscler. Thromb. Vasc. Biol. 28(12), 2282–2287 (2008)
K. Jimbow, J.S. Park, F. Kato, K. Hirosaki, K. Toyofuku, C. Hua, T. Yamashita, Assembly, target-signaling and intracellular transport of tyrosinase gene family proteins in the initial stage of melanosome biogenesis. Pigment Cell Res. 13(4), 222–229 (2000)
S.H. Jin, Y.Y. Lee, H.Y. Kang, Methyl-beta-cyclodextrin, a specific cholesterol-binding agent, inhibits melanogenesis in human melanocytes through activation of ERK. Arch. Dermatol. Res. 300(8), 451–454 (2008)
R.W. Klemm, C.S. Ejsing, M.A. Surma, H.J. Kaiser, M.J. Gerl, J.L. Sampaio, Q. de Robillard, C. Ferguson, T.J. Proszynski, A. Shevchenko, K. Simons, Segregation of sphingolipids and sterols during formation of secretory vesicles at the trans-Golgi network. J. Cell Biol. 185(4), 601–612 (2009)
H.M. Koo, M. VanBrocklin, M.J. McWilliams, S.H. Leppla, N.S. Duesbery, G.F. Woude, Apoptosis and melanogenesis in human melanoma cells induced by anthrax lethal factor inactivation of mitogen-activated protein kinase kinase. Proc. Natl. Acad. Sci. U. S. A. 99(5), 3052–3057 (2002)
R.L. Lamason, M.A. Mohideen, J.R. Mest, A.C. Wong, H.L. Norton, M.C. Aros, M.J. Jurynec, X. Mao, V.R. Humphreville, J.E. Humbert, S. Sinha, J.L. Moore, P. Jagadeeswaran, W. Zhao, G. Ning, I. Makalowska, P.M. McKeigue, D. O’donnell, R. Kittles, E.J. Parra, N.J. Mangini, D.J. Grunwald, M.D. Shriver, V.A. Canfield, K.C. Cheng, SLC24A5, a putative cation exchanger, affects pigmentation in zebrafish and humans. Science 310(5755), 1782–17866 (2005)
E. Le Pape, T. Passeron, A. Giubellino, J.C. Valencia, R. Wolber, V.J. Hearing, Microarray analysis sheds light on the dedifferentiating role of agouti signal protein in murine melanocytes via the Mc1r. Proc. Natl. Acad. Sci. U. S. A. 106(6), 1802–1807 (2009)
L. Liscum, J.R. Faust, The intracellular transport of low density lipoprotein-derived cholesterol is inhibited in Chinese hamster ovary cells cultured with 3-beta-[2-(diethylamino)ethoxy]androst-5-en-17-one. J. Biol. Chem. 264(20), 11796–11806 (1989)
A. Luini, A.A. Mironov, E.V. Polishchuk, R.S. Polishchuk, Morphogenesis of post-Golgi transport carriers. Histochem. Cell Biol. 129(2), 153–161 (2008)
N.J. Nadeau, F. Minvielle, S. Ito, M. Inoue-Murayama, D. Gourichon, S.A. Follett, T. Burke, N.I. Mundy, Characterization of Japanese quail yellow as a genomic deletion upstream of the avian homolog of the mammalian ASIP (agouti) gene. Genetics 178(2), 777–786 (2008)
R.A. Newton, A.L. Cook, D.W. Roberts, J.H. Leonard, R.A. Sturm, Post-transcriptional regulation of melanin biosynthetic enzymes by cAMP and resveratrol in human melanocytes. J. Invest. Dermatol. 127(9), 2216–2227 (2007)
C. Olivares, F. Solano, J.C. García-Borrón, Conformation-dependent post-translational glycosylation of tyrosinase. Requirement of a specific interaction involving the CuB metal binding site. J. Biol. Chem. 278(18), 15735–15743 (2003)
I. Palmisano, P. Bagnato, A. Palmigiano, G. Innamorati, G. Rotondo, D. Altimare, C. Venturi, E.V. Sviderskaya, R. Piccirillo, M. Coppola, V. Marigo, B. Incerti, A. Ballabio, E.M. Surace, C. Tacchetti, D.C. Bennett, M.V. Schiaffino, The ocular albinism type 1 protein, an intracellular G protein-coupled receptor, regulates melanosome transport in pigment cells. Hum. Mol. Genet. 17(22), 3487–3501 (2008)
H.Y. Park, C. Wu, L. Yonemoto, M. Murphy-Smith, H. Wu, C.M. Stachur, B.A. Gilchrest, MITF mediates cAMP-induced protein kinase C-beta expression in human melanocytes. Biochem. J. 395(3), 571–578 (2006)
A.E. Radulescu, A. Siddhanta, D. Shields, A role for clathrin in reassembly of the Golgi apparatus. Mol. Biol. Cell 18(1), 94–105 (2007)
D. Rodriguez-Agudo, S. Ren, E. Wong, D. Marques, K. Redford, G. Gil, P. Hylemon, W.M. Pandak, Intracellular cholesterol transporter StarD4 binds free cholesterol and increases cholesteryl ester formation. J. Lipid Res. 49(7), 1409–1419 (2008)
K.U. Schallreuter, S. Hasse, H. Rokos, B. Chavan, M. Shalbaf, J.D. Spencer, J.M. Wood, Cholesterol regulates melanogenesis in human epidermal melanocytes and melanoma cells. Exp. Dermatol. 18(8), 680–688 (2009)
A. Schepsky, K. Bruser, G.J. Gunnarsson, J. Goodall, J.H. Hallsson, C.R. Goding, E. Steingrimsson, A. Hecht, The microphthalmia-associated transcription factor Mitf interacts with beta-catenin to determine target gene expression. Mol. Cell. Biol. 26(23), 8914–8927 (2006)
P.P. Schnetkamp, The SLC24 Na+/Ca2+-K+ exchanger family: vision and beyond. Pflugers Arch. 447(5), 683–688 (2004)
M.C. Scott, I. Suzuki, Z.A. Abdel-Malek, Regulation of the human melanocortin 1 receptor expression in epidermal melanocytes by paracrine and endocrine factors and by ultraviolet radiation. Pigment Cell Res. 15(6), 433–439 (2002)
N. Sekar, J.D. Veldhuis, Concerted transcriptional activation of the low density lipoprotein receptor gene by insulin and luteinizing hormone in cultured porcine granulosa-luteal cells: possible convergence of protein kinase a, phosphatidylinositol 3-kinase, and mitogen-activated protein kinase signaling pathways. Endocrinology 142(7), 2921–2928 (2001)
S.R. Setty, D. Tenza, E.V. Sviderskaya, D.C. Bennett, G. Raposo, M.S. Marks, Cell-specific ATP7A transport sustains copper-dependent tyrosinase activity in melanosomes. Nature 454(7208), 1142–1146 (2008)
R.P. Stokowski, P.V. Pant, T. Dadd, A. Fereday, D.A. Hinds, C. Jarman, W. Filsell, R.S. Ginger, M.R. Green, F.J. van der Ouderaa, D.R. Cox, A genomewide association study of skin pigmentation in a South Asian population. Am. J. Hum. Genet. 81(6), 1119–1132 (2007)
P. Vogel, R.W. Read, R.B. Vance, K.A. Platt, K. Troughton, D.S. Rice, Ocular albinism and hypopigmentation defects in Slc24a5-/- mice. Vet. Pathol. 45(2), 264–279 (2008)
Y. Yamaguchi, V.J. Hearing, Physiological factors that regulate skin pigmentation. Biofactors 35(2), 193–199 (2009)
L. Zhou, H.Y. Choi, W.P. Li, F. Xu, J. Herz, LRP1 controls cPLA2 phosphorylation, ABCA1 expression and cellular cholesterol export. PLoS One 4(8), e6853 (2009)
Acknowledgements
We thank Carl Jarman, Wendy Filsell, Amelia Jarman, Gordon James and Dawn Mazzatti for their support and helpful discussions.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2013 Springer Science+Business Media New York
About this chapter
Cite this chapter
Wilson, S. et al. (2013). NCKX5, a Natural Regulator of Human Skin Colour Variation, Regulates the Expression of Key Pigment Genes MC1R and Alpha-MSH and Alters Cholesterol Homeostasis in Normal Human Melanocytes. In: Annunziato, L. (eds) Sodium Calcium Exchange: A Growing Spectrum of Pathophysiological Implications. Advances in Experimental Medicine and Biology, vol 961. Springer, Boston, MA. https://doi.org/10.1007/978-1-4614-4756-6_9
Download citation
DOI: https://doi.org/10.1007/978-1-4614-4756-6_9
Published:
Publisher Name: Springer, Boston, MA
Print ISBN: 978-1-4614-4755-9
Online ISBN: 978-1-4614-4756-6
eBook Packages: Biomedical and Life SciencesBiomedical and Life Sciences (R0)