Abstract

Phenotypic plasticity and subsequent generation of intratumoral heterogeneity underly key traits in malignant melanoma such as drug resistance and metastasis. Melanoma plasticity promotes a switch between proliferative and invasive phenotypes characterized by different transcriptional programs of which MITF is a critical regulator. Here, we show that the acid ceramidase ASAH1, which controls sphingolipid metabolism, acted as a rheostat of the phenotypic switch in melanoma cells. Low ASAH1 expression was associated with an invasive behavior mediated by activation of the integrin alphavbeta5-FAK signaling cascade. In line with that, human melanoma biopsies revealed heterogeneous staining of ASAH1 and low ASAH1 expression at the melanoma invasive front. We also identified ASAH1 as a new target of MITF, thereby involving MITF in the regulation of sphingolipid metabolism. Together, our findings provide new cues to the mechanisms underlying the phenotypic plasticity of melanoma cells and identify new anti-metastatic targets.

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References

  1. 1.

    Colombino M, Sini M, Lissia A, De Giorgi V, Stanganelli I, Ayala F, et al. Discrepant alterations in main candidate genes among multiple primary melanomas. J Transl Med. 2014;12:117.

  2. 2.

    Kemper K, Krijgsman O, Cornelissen-Steijger P, Shahrabi A, Weeber F, Song JY, et al. Intra- and inter-tumor heterogeneity in a vemurafenib-resistant melanoma patient and derived xenografts. EMBO Mol Med. 2015;7:1104–18.

  3. 3.

    Hoek KS, Eichhoff OM, Schlegel NC, Dobbeling U, Kobert N, Schaerer L, et al. In vivo switching of human melanoma cells between proliferative and invasive states. Cancer Res. 2008;68:650–6.

  4. 4.

    Johannessen CM, Johnson LA, Piccioni F, Townes A, Frederick DT, Donahue MK, et al. A melanocyte lineage program confers resistance to MAP kinase pathway inhibition. Nature. 2013;504:138–42.

  5. 5.

    Konieczkowski DJ, Johannessen CM, Abudayyeh O, Kim JW, Cooper ZA, Piris V, et al. A melanoma cell state distinction influences sensitivity to MAPK pathway inhibitors. Cancer Discov. 2014;4:816–27.

  6. 6.

    Muller J, Krijgsman O, Tsoi J, Robert L, Hugo W, Song C, et al. Low MITF/AXL ratio predicts early resistance to multiple targeted drugs in melanoma. Nat Commun. 2014;5:5712.

  7. 7.

    Van Allen EM, Foye A, Wagle N, Kim W, Carter SL, McKenna A, et al. Successful whole-exome sequencing from a prostate cancer bone metastasis biopsy. Prostate Cancer Prostatic Dis. 2014;17:23–27.

  8. 8.

    Falletta P, Sanchez-Del-Campo L, Chauhan J, Effern M, Kenyon A, Kershaw CJ, et al. Translation reprogramming is an evolutionarily conserved driver of phenotypic plasticity and therapeutic resistance in melanoma. Genes Dev. 2017;31:18–33.

  9. 9.

    Riesenberg S, Groetchen A, Siddaway R, Bald T, Reinhardt J, Smorra D, et al. MITF and c-Jun antagonism interconnects melanoma dedifferentiation with pro-inflammatory cytokine responsiveness and myeloid cell recruitment. Nat Commun. 2015;6:8755.

  10. 10.

    Bertolotto C, Lesueur F, Giuliano S, Strub T, de Lichy M, Bille K, et al. A SUMOylation-defective MITF germline mutation predisposes to melanoma and renal carcinoma. Nature. 2011;480:94–98.

  11. 11.

    Paillerets BB, Lesueur F, Bertolotto C. A germline oncogenic MITF mutation and tumor susceptibility. Eur J Cell Biol. 2014;93:71–5.

  12. 12.

    Bonet C, Luciani F, Ottavi JF, Leclerc J, Jouenne FM, Boncompagni M, et al. Deciphering the role of oncogenic MITFE318K in senescence delay and melanoma progression. J Natl Cancer Inst 2017; 109. djw340

  13. 13.

    Yokoyama S, Woods SL, Boyle GM, Aoude LG, MacGregor S, Zismann V, et al. A novel recurrent mutation in MITF predisposes to familial and sporadic melanoma. Nature. 2011;480:99–103.

  14. 14.

    Carreira S, Goodall J, Denat L, Rodriguez M, Nuciforo P, Hoek KS, et al. Mitf regulation of Dia1 controls melanoma proliferation and invasiveness. Genes Dev. 2006;20:3426–39.

  15. 15.

    Cheli Y, Ohanna M, Ballotti R, Bertolotto C. Fifteen-year quest for microphthalmia-associated transcription factor target genes. Pigment Cell Melanoma Res. 2010;23:27–40.

  16. 16.

    Cheli Y, Giuliano S, Botton T, Rocchi S, Hofman V, Hofman P, et al. Mitf is the key molecular switch between mouse or human melanoma initiating cells and their differentiated progeny. Oncogene. 2011;30:2307–18.

  17. 17.

    Cheli Y, Giuliano S, Fenouille N, Allegra M, Hofman V, Hofman P, et al. Hypoxia and MITF control metastatic behaviour in mouse and human melanoma cells. Oncogene. 2012;31:2461–70.

  18. 18.

    Cairns RA, Harris IS, Mak TW. Regulation of cancer cell metabolism. Nat Rev Cancer. 2011;11:85–95.

  19. 19.

    Warburg O. On respiratory impairment in cancer cells. Science. 1956;124:269–70.

  20. 20.

    Ganapathy-Kanniappan S, Geschwind JF. Tumor glycolysis as a target for cancer therapy: progress and prospects. Mol Cancer. 2013;12:152.

  21. 21.

    Haq R, Shoag J, Andreu-Perez P, Yokoyama S, Edelman H, Rowe GC, et al. Oncogenic BRAF regulates oxidative metabolism via PGC1alpha and MITF. Cancer Cell. 2013;23:302–15.

  22. 22.

    Vazquez F, Lim JH, Chim H, Bhalla K, Girnun G, Pierce K, et al. PGC1alpha expression defines a subset of human melanoma tumors with increased mitochondrial capacity and resistance to oxidative stress. Cancer Cell. 2013;23:287–301.

  23. 23.

    Currie E, Schulze A, Zechner R, Walther TC, Farese RV Jr. Cellular fatty acid metabolism and cancer. Cell Metab. 2013;18:153–61.

  24. 24.

    Morad SA, Cabot MC. Ceramide-orchestrated signalling in cancer cells. Nat Rev Cancer. 2013;13:51–65.

  25. 25.

    Ferlinz K, Kopal G, Bernardo K, Linke T, Bar J, Breiden B, et al. Human acid ceramidase: processing, glycosylation, and lysosomal targeting. J Biol Chem. 2001;276:35352–60.

  26. 26.

    Rambow F, Job B, Petit V, Gesbert F, Delmas V, Seberg H, et al. New functional signatures for understanding melanoma biology from tumor cell lineage-specific analysis. Cell Rep. 2015;13:840–53.

  27. 27.

    Shtraizent N, Eliyahu E, Park JH, He X, Shalgi R, Schuchman EH. Autoproteolytic cleavage and activation of human acid ceramidase. J Biol Chem. 2008;283:11253–9.

  28. 28.

    Zhou J, Tawk M, Tiziano FD, Veillet J, Bayes M, Nolent F, et al. Spinal muscular atrophy associated with progressive myoclonic epilepsy is caused by mutations in ASAH1. Am J Hum Genet. 2012;91:5–14.

  29. 29.

    Bedia C, Casas J, Garcia V, Levade T, Fabrias G. Synthesis of a novel ceramide analogue and its use in a high-throughput fluorogenic assay for ceramidases. Chembiochem. 2007;8:642–8.

  30. 30.

    Strub T, Giuliano S, Ye T, Bonet C, Keime C, Kobi D, et al. Essential role of microphthalmia transcription factor for DNA replication, mitosis and genomic stability in melanoma. Oncogene. 2011;30:2319–32.

  31. 31.

    Ohanna M, Cerezo M, Nottet N, Bille K, Didier R, Beranger G, et al. Pivotal role of NAMPT in the switch of melanoma cells toward an invasive and drug-resistant phenotype. Genes Dev. 2018;32:448–61.

  32. 32.

    Walkley SU, Vanier MT. Secondary lipid accumulation in lysosomal disease. Biochim Biophys Acta. 2009;1793:726–36.

  33. 33.

    Verfaillie A, Imrichova H, Atak ZK, Dewaele M, Rambow F, Hulselmans G, et al. Decoding the regulatory landscape of melanoma reveals TEADS as regulators of the invasive cell state. Nat Commun. 2015;6:6683.

  34. 34.

    Caramel J, Papadogeorgakis E, Hill L, Browne GJ, Richard G, Wierinckx A, et al. A switch in the expression of embryonic EMT-inducers drives the development of malignant melanoma. Cancer Cell. 2013;24:466–80.

  35. 35.

    Realini N, Palese F, Pizzirani D, Pontis S, Basit A, Bach A, et al. Acid ceramidase in melanoma: expression, localization, and effects of pharmacological inhibition. J Biol Chem. 2016;291:2422–34.

  36. 36.

    Lucki N, Sewer MB. The cAMP-responsive element binding protein (CREB) regulates the expression of acid ceramidase (ASAH1) in H295R human adrenocortical cells. Biochim Biophys Acta. 2009;1791:706–13.

  37. 37.

    Lucki NC, Sewer MB. Genistein stimulates MCF-7 breast cancer cell growth by inducing acid ceramidase (ASAH1) gene expression. J Biol Chem. 2011;286:19399–409.

  38. 38.

    Don AS, Lim XY, Couttas TA. Re-configuration of sphingolipid metabolism by oncogenic transformation. Biomolecules. 2014;4:315–53.

  39. 39.

    Truman JP, Garcia-Barros M, Obeid LM, Hannun YA. Evolving concepts in cancer therapy through targeting sphingolipid metabolism. Biochim Biophys Acta. 2014;1841:1174–88.

  40. 40.

    Cheng JC, Bai A, Beckham TH, Marrison ST, Yount CL, Young K, et al. Radiation-induced acid ceramidase confers prostate cancer resistance and tumor relapse. J Clin Invest. 2013;123:4344–58.

  41. 41.

    Elojeimy S, Liu X, McKillop JC, El-Zawahry AM, Holman DH, Cheng JY, et al. Role of acid ceramidase in resistance to FasL: therapeutic approaches based on acid ceramidase inhibitors and FasL gene therapy. Mol Ther. 2007;15:1259–63.

  42. 42.

    Mahdy AE, Cheng JC, Li J, Elojeimy S, Meacham WD, Turner LS, et al. Acid ceramidase upregulation in prostate cancer cells confers resistance to radiation: AC inhibition, a potential radiosensitizer. Mol Ther. 2009;17:430–8.

  43. 43.

    Ruckhaberle E, Holtrich U, Engels K, Hanker L, Gatje R, Metzler D, et al. Acid ceramidase 1 expression correlates with a better prognosis in ER-positive breast cancer. Climacteric. 2009;12:502–13.

  44. 44.

    Sanger N, Ruckhaberle E, Gyorffy B, Engels K, Heinrich T, Fehm T, et al. Acid ceramidase is associated with an improved prognosis in both DCIS and invasive breast cancer. Mol Oncol. 2015;9:58–67.

  45. 45.

    Park JH, Schuchman EH. Acid ceramidase and human disease. Biochim Biophys Acta. 2006;1758:2133–8.

  46. 46.

    Bonet C, Giuliano S, Ohanna M, Bille K, Allegra M, Lacour JP, et al. Aurora B is regulated by the mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) signaling pathway and is a valuable potential target in melanoma cells. J Biol Chem. 2012;287:29887–98.

  47. 47.

    Giuliano S, Cheli Y, Ohanna M, Bonet C, Beuret L, Bille K, et al. Microphthalmia-associated transcription factor controls the DNA damage response and a lineage-specific senescence program in melanomas. Cancer Res. 2010;70:3813–22.

  48. 48.

    Lai M, Realini N, La Ferla M, Passalacqua I, Matteoli G, Ganesan A, et al. Complete acid ceramidase ablation prevents cancer-initiating cell formation in melanoma cells. Sci Rep. 2017;7:7411.

  49. 49.

    Ohanna M, Giuliano S, Bonet C, Imbert V, Hofman V, Zangari J, et al. Senescent cells develop a PARP-1 and nuclear factor-{kappa}B-associated secretome (PNAS). Genes Dev. 2011;25:1245–61.

  50. 50.

    Ohanna M, Cheli Y, Bonet C, Bonazzi VF, Allegra M, Giuliano S, et al. Secretome from senescent melanoma engages the STAT3 pathway to favor reprogramming of naive melanoma towards a tumor-initiating cell phenotype. Oncotarget. 2013;4:2212–24.

  51. 51.

    Gupta PB, Kuperwasser C, Brunet JP, Ramaswamy S, Kuo WL, Gray JW, et al. The melanocyte differentiation program predisposes to metastasis after neoplastic transformation. Nat Genet. 2005;37:1047–54.

  52. 52.

    Pinner S, Jordan P, Sharrock K, Bazley L, Collinson L, Marais R, et al. Intravital imaging reveals transient changes in pigment production and Brn2 expression during metastatic melanoma dissemination. Cancer Res. 2009;69:7969–77.

  53. 53.

    Villanueva J, Vultur A, Lee JT, Somasundaram R, Fukunaga-Kalabis M, Cipolla AK, et al. Acquired resistance to BRAF inhibitors mediated by a RAF kinase switch in melanoma can be overcome by cotargeting MEK and IGF-1R/PI3K. Cancer Cell. 2010;18:683–95.

  54. 54.

    Widmer DS, Cheng PF, Eichhoff OM, Belloni BC, Zipser MC, Schlegel NC, et al. Systematic classification of melanoma cells by phenotype-specific gene expression mapping. Pigment Cell Melanoma Res. 2012;25:343–53.

  55. 55.

    Reinhardt J, Landsberg J, Schmid-Burgk JL, Ramis BB, Bald T, Glodde N, et al. MAPK signaling and inflammation link melanoma phenotype switching to induction of CD73 during immunotherapy. Cancer Res. 2017;77:4697–709.

  56. 56.

    Webster MR, Xu M, Kinzler KA, Kaur A, Appleton J, O’Connell MP, et al. Wnt5A promotes an adaptive, senescent-like stress response, while continuing to drive invasion in melanoma cells. Pigment Cell Melanoma Res. 2015;28:184–95.

  57. 57.

    Canel M, Serrels A, Frame MC, Brunton VG. E-cadherin-integrin crosstalk in cancer invasion and metastasis. J Cell Sci. 2013;126:393–401.

  58. 58.

    Vesuna F, van Diest P, Chen JH, Raman V. Twist is a transcriptional repressor of E-cadherin gene expression in breast cancer. Biochem Biophys Res Commun. 2008;367:235–41.

  59. 59.

    Wang F, Van Brocklyn JR, Edsall L, Nava VE, Spiegel S. Sphingosine-1-phosphate inhibits motility of human breast cancer cells independently of cell surface receptors. Cancer Res. 1999;59:6185–91.

  60. 60.

    Desch A, Strozyk EA, Bauer AT, Huck V, Niemeyer V, Wieland T, et al. Highly invasive melanoma cells activate the vascular endothelium via an MMP-2/integrin alphavbeta5-induced secretion of VEGF-A. Am J Pathol. 2012;181:693–705.

  61. 61.

    Fane ME, Chhabra Y, Hollingsworth DE, Simmons JL, Spoerri L, Oh TG, et al. NFIB mediates BRN2 driven melanoma cell migration and invasion through regulation of EZH2 and MITF. EBioMedicine. 2017;16:63–75.

  62. 62.

    Goodall J, Carreira S, Denat L, Kobi D, Davidson I, Nuciforo P, et al. Brn-2 represses microphthalmia-associated transcription factor expression and marks a distinct subpopulation of microphthalmia-associated transcription factor-negative melanoma cells. Cancer Res. 2008;68:7788–94.

  63. 63.

    Larribere L, Hilmi C, Khaled M, Gaggioli C, Bille K, Auberger P, et al. The cleavage of microphthalmia associated transcription factor, MITF, by caspases plays an essential role in melanocyte and melanoma cell apoptosis. Genes Dev. 2005;19:1980–5.

  64. 64.

    Merrill AH Jr, Sullards MC, Allegood JC, Kelly S, Wang E. Sphingolipidomics: high-throughput, structure-specific, and quantitative analysis of sphingolipids by liquid chromatography tandem mass spectrometry. Methods. 2005;36:207–24.

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Acknowledgements

This work was supported by Inserm, La Société Française de Dermatologie, and by a grant from INCA (INCA_10573). CP is a fellowship from la Ligue Nationale contre le Cancer. The authors thank Dr. M Sewer (San Diego, USA) for providing the ASAH1 promoter vector. WM3912, WM8, WM3928 and WM3918 human melanoma cell lines were a kind gift from H Meenhard and G Zhang (Wistar melanoma Institute, Philadelphia, USA).

Author contributions

CB, NA-A, RB and TL designed the research, analyzed the results and wrote the manuscript. GT, SD, JC and PB performed and analyzed the immunohistochemistry experiments. NN performed the bioinformatics analysis. JL, DG, CP, CG, KB, VG, PC, SP and BM performed all the other experiments.

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Author notes

  1. These authors contributed equally: Justine Leclerc, David Garandeau, Nathalie Andrieu-Abadie, Corine Bertolotto.

Affiliations

  1. Team 1, Biology and Pathologies of Melanocytes, Equipe labellisée ARC 2015, Université Côte d’Azur, Inserm U1065, C3M, Nice, France

    • Justine Leclerc
    • , Charlotte Pandiani
    • , Céline Gaudel
    • , Karine Bille
    • , Philippe Bahadoran
    • , Robert Ballotti
    •  & Corine Bertolotto
  2. Université Côte d’Azur, Inserm U1065, C3M, Nice, France

    • David Garandeau
    •  & Nicolas Nottet
  3. Team 4, Sphingolipids, Metabolism, Cell Death and Tumor Progression, Université Toulouse III, Toulouse, Inserm, UMR1037, CRCT, Toulouse, France

    • Virginie Garcia
    • , Thierry Levade
    •  & Nathalie Andrieu-Abadie
  4. CarMeN Laboratory, INSERM U1060, INRA1397, INSA, Lyon, France

    • Pascal Colosetti
  5. Université Côte d’Azur, Centre Commun de Microscopie Appliquée, Nice, France

    • Sophie Pagnotta
  6. CHU NICE, Département de Dermatologie, Nice, France

    • Philippe Bahadoran
  7. Université de Lyon, Inserm, U1052, CNRS 5286, Equipe Labellisée Ligue contre le Cancer, Lyon, France

    • Garance Tondeur
    • , Stéphane Dalle
    •  & Julie Caramel
  8. INSERM ERI21/EA 4319, Nice, F-06107, France

    • Baharia Mograbi

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https://doi.org/10.1038/s41388-018-0500-0