Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Acute myeloid leukemia

NPM1c impedes CTCF functions through cytoplasmic mislocalization in acute myeloid leukemia

Leukemia volume 34, pages 1278–1290 (2020)Cite this article

Subjects

Abstract

Normal cytogenetic acute myeloid leukemia (AML) frequently harbor a TCTG insertion in exon 12 of Nucleophosmin 1 (NPM1); the resulting frameshift creates a nuclear export signal (NES) and cytoplasmic localization of NPM1c. However, how NPM1c causes AML is not completely understood. NPM1 participates in multiple protein–protein interactions one of which involves the CCCTC-binding factor (CTCF). Through binding of CTCF binding sites (CBS), CTCF mediates nuclear functions including DNA looping, regulation of gene expression, and RNA splicing. We hypothesized that mislocalization of CTCF into the cytoplasm by NPM1c reduces the functional level of nuclear CTCF and so alters gene expression. We verified the interaction of CTCF with NPM1 and showed that CTCF interacts with NPM1c, with redistribution of CTCF into the cytoplasm. The interaction of CTCF and NPM1c involves the amino terminus of CTCF and the last 50 amino acids of NPM1. By interfering with the interaction of CTCF and NPM1c, CTCF becomes relocalized into the nucleus.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: CTCF interacts with NPM1 and NPM1c and is mislocalized with NPM1c into the cytoplasm.
Fig. 2: Altered CTCF-regulatory functions due to NPM1c cytoplasmic mislocalization of CTCF.
Fig. 3: CD45 exon 5 exclusion is a characteristic of NPM1c.
Fig. 4: Increased DNA methylation of CBSs in NPM1c AML cases.
Fig. 5: The interaction of CTCF with NPM1 is dependent on the N-terminus of CTCF.
Fig. 6: The CTCF:NPM1 interaction is dependent on the C-terminus of NPM1wt and NPM1c.
Fig. 7: NPM1c-mislocalized CTCF can be relocalized back into the nucleus by disrupting the CTCF:NPM1c interaction.

Similar content being viewed by others

References

  1. Falini B, Nicoletti I, Martelli MF, Mecucci C, Harrison C, Harrison G, et al. Acute myeloid leukemia carrying cytoplasmic/mutated nucleophosmin (NPMc+ AML): biologic and clinical features. Blood. 2007;109:874–85.

    CAS  PubMed  Google Scholar 

  2. Ley TJ, Miller C, Ding L, Raphael BJ, Mungall AJ, Robertson G, et al. Genomic and epigenomic landscapes of adult de novo acute myeloid leukemia. N. Engl J Med. 2013;368:2059–74.

    PubMed  Google Scholar 

  3. Falini B, Bolli N, Liso A, Martelli MP, Mannucci R, Pileri S, et al. Altered nucleophosmin transport in acute myeloid leukaemia with mutated NPM1: molecular basis and clinical implications. Leukemia. 2009;23:1731–43.

    CAS  PubMed  Google Scholar 

  4. Wanzel M, Russ AC, Kleine-Kohlbrecher D, Colombo E, Pelicci P-G, Eilers M. A ribosomal protein L23-nucleophosmin circuit coordinates Miz1 function with cell growth. Nat Cell Biol. 2008;10:1051–61.

    CAS  PubMed  Google Scholar 

  5. Bonetti P, Davoli T, Sironi C, Amati B, Pelicci PG, Colombo E. Nucleophosmin and its AML-associated mutant regulate c-Myc turnover through Fbw7? J Cell Biol. 2008;182:19–26.

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Gurumurthy M, Tan CH, Ng R, Zeiger L, Lau J, Lee J, et al. Nucleophosmin Interacts with HEXIM1 and regulates RNA polymerase II transcription. J Mol Biol. 2008;378:302–17.

    CAS  PubMed  Google Scholar 

  7. Gu X, Ebrahem Q, Mahfouz RZ, Hasipek M, Enane F, Radivoyevitch T, et al. Leukemogenic nucleophosmin mutation disrupts the transcription factor hub that regulates granulomonocytic fates. J Clin Investig. 2018;128:4260–79.

    PubMed  Google Scholar 

  8. Yusufzai TM, Tagami H, Nakatani Y, Felsenfeld G. CTCF tethers an insulator to subnuclear sites, suggesting shared insulator mechanisms across species. Mol Cell. 2004;13:291–8.

    CAS  PubMed  Google Scholar 

  9. Phillips JE, Corces VG. CTCF: master weaver of the genome. Cell. 2009;137:1194–211.

    PubMed  PubMed Central  Google Scholar 

  10. Ohlsson R, Renkawitz R, Lobanenkov V. CTCF is a uniquely versatile transcription regulator linked to epigenetics and disease. Trends Genet. 2001;17:520–7.

    CAS  PubMed  Google Scholar 

  11. Bell AC, West AG, Felsenfeld G. The protein CTCF is required for the enhancer blocking activity of vertebrate insulators. Cell. 1999;98:387–96.

    CAS  PubMed  Google Scholar 

  12. Xu N, Donohoe ME, Silva SS, Lee JT. Evidence that homologous X-chromosome pairing requires transcription and Ctcf protein. Nat Genet. 2007;39:1390–6.

    CAS  PubMed  Google Scholar 

  13. Szabó PE, Tang SHE, Rentsendorj A, Pfeifer GP, Mann JR. Maternal-specific footprints at putative CTCF sites in the H19 imprinting control region give evidence for insulator function. Curr Biol. 2000;10:607–10.

    PubMed  Google Scholar 

  14. Bell AC, Felsenfeld G. Methylation of a CTCF-dependent boundary controls imprinted expression of the Igf2 gene. Nature. 2000;405:482–5.

    CAS  PubMed  Google Scholar 

  15. Klenova EM, Nicolas RH, Paterson HF, Carne AF, Heath CM, Goodwin GH, et al. CTCF, a conserved nuclear factor required for optimal transcriptional activity of the chicken c-myc gene, is an 11-Zn-finger protein differentially expressed in multiple forms. Mol Cell Biol. 1993;13:7612–24.

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Burcin M, Arnold R, Lutz M, Kaiser B, Runge D, Lottspeich F, et al. Negative protein 1, which is required for function of the chicken lysozyme gene silencer in conjunction with hormone receptors, is identical to the multivalent zinc finger repressor CTCF. Mol Cell Biol. 1997;17:1281–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Vostrov AA, Quitschke WW. The zinc finger protein CTCF binds to the APBbeta domain of the amyloid beta-protein precursor promoter. Evidence for a role in transcriptional activation. J Biol Chem. 1997;272:33353–9.

    CAS  PubMed  Google Scholar 

  18. Farrar D, Rai S, Chernukhin I, Jagodic M, Ito Y, Yammine S, et al. Mutational analysis of the poly(ADP-Ribosyl)ation sites of the transcription factor CTCF provides an insight into the mechanism of its regulation by poly(ADP-Ribosyl)ation. Mol Cell Biol. 2010;30:1199–216.

    CAS  PubMed  Google Scholar 

  19. Zuin J, Dixon JR, van der Reijden MIJA, Ye Z, Kolovos P, RWW Brouwer, et al. Cohesin and CTCF differentially affect chromatin architecture and gene expression in human cells. Proc Natl Acad Sci USA. 2014;111:996–1001.

    CAS  PubMed  Google Scholar 

  20. Ruiz-Velasco M, Zaugg JB. Structure meets function: how chromatin organisation conveys functionality. Curr Opin Syst Biol. 2017;1:129–36.

    Google Scholar 

  21. Botta M, Haider S, Leung IXY, Lio P, Mozziconacci J. Intra- and inter-chromosomal interactions correlate with CTCF binding genome wide. Mol Syst Biol. 2010;6:426.

    PubMed  PubMed Central  Google Scholar 

  22. Shukla S, Kavak E, Gregory M, Imashimizu M, Shutinoski B, Kashlev M, et al. CTCF-promoted RNA polymerase II pausing links DNA methylation to splicing. Nature. 2011;479:74–79.

    CAS  PubMed  Google Scholar 

  23. Zampieri M, Guastafierro T, Calabrese R, Ciccarone F, Bacalini MG, Reale A, et al. ADP-ribose polymers localized on Ctcf–Parp1–Dnmt1 complex prevent methylation of Ctcf target sites. Biochem J. 2012;441:645–52.

    CAS  PubMed  Google Scholar 

  24. Pant V, Mariano P, Kanduri C, Mattsson A, Lobanenkov V, Heuchel R, et al. The nucleotides responsible for the direct physical contact between the chromatin insulator protein CTCF and the H19 imprinting control region manifest parent of origin-specific long-distance insulation and methylation-free domains. Genes Dev. 2003;17:586–90.

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Kemp CJ, Moore JM, Moser R, Bernard B, Teater M, Smith LE, et al. CTCF haploinsufficiency destabilizes DNA methylation and predisposes to cancer. Cell Rep. 2014;7:1020–9.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Verhaak RGW, Goudswaard CS, Van Putten W, Bijl MA, Sanders MA, Hugens W, et al. Mutations in nucleophosmin (NPM1) in acute myeloid leukemia (AML): association with other gene abnormalities and previously established gene expression signatures and their favorable prognostic significance. Blood. 2005;106:3747–54.

    CAS  PubMed  Google Scholar 

  27. Brunetti L, Gundry MC, Sorcini D, Guzman AG, Huang YH, Ramabadran R, et al. Mutant NPM1 maintains the leukemic state through HOX expression. Cancer Cell. 2018. https://doi.org/10.1016/j.ccell.2018.08.005.

  28. Vassiliou GS, Cooper JL, Rad R, Li J, Rice S, Uren A, et al. Mutant nucleophosmin and cooperating pathways drive leukemia initiation and progression in mice. Nat Genet. 2011;43:470–5.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Kim TH, Abdullaev ZK, Smith AD, Ching KA, Loukinov DI, Green RDD, et al. Analysis of the vertebrate insulator protein CTCF-binding sites in the human genome. Cell. 2007;128:1231–45.

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Luo H, Wang F, Zha J, Li H, Yan B, Du Q, et al. CTCF boundary remodels chromatin domain and drives aberrant HOX gene transcription in acute myeloid leukemia. Blood. 2018. https://doi.org/10.1182/blood-2017-11-814319.

  31. Gruszka AM, Lavorgna S, Consalvo MI, Ottone T, Martinelli C, Cinquanta M, et al. A monoclonal antibody against mutated nucleophosmin 1 for the molecular diagnosis of acute myeloid leukemias. Blood. 2010;116:2096–102.

    CAS  PubMed  Google Scholar 

  32. Schmidt D, Wilson MD, Spyrou C, Brown GD, Hadfield J, Odom DT. ChIP-seq: using high-throughput sequencing to discover protein-DNA interactions. Methods. 2009;48:240–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Aryee MJ, Jaffe AE, Corrada-Bravo H, Ladd-Acosta C, Feinberg AP, Hansen KD, et al. Minfi: A flexible and comprehensive Bioconductor package for the analysis of Infinium DNA methylation microarrays. Bioinformatics. 2014;30:1363–9.

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Ziebarth JD, Bhattacharya A, Cui Y CTCFBSDB 2.0: A database for CTCF-binding sites and genome organization. Nucleic Acids Res. 2013; 41. https://doi.org/10.1093/nar/gks1165.

  35. Mancini E, Iserte J, Yanovsky M, Chernomoretz A. ASpli: analysis of alternative splicing using RNA-Seq. 2017. https://doi.org/10.18129/B9.bioc.ASpli.

  36. Kim YJ, Cecchini KR, Kim TH. Conserved, developmentally regulated mechanism couples chromosomal looping and heterochromatin barrier activity at the homeobox gene A locus. Proc Natl Acad Sci USA. 2011;108:7391–6.

    CAS  PubMed  Google Scholar 

  37. Bailey SD, Zhang X, Desai K, Aid M, Corradin O, Cowper-Sallari R, et al. ZNF143 provides sequence specificity to secure chromatin interactions at gene promoters. Nat Commun. 2015;2. https://doi.org/10.1038/ncomms7186.

  38. De La Rosa-Velazquez IA, Rincon-Arano H, Benitez-Bribiesca L, Recillas-Targa F. Epigenetic regulation of the human retinoblastoma tumor suppressor gene promoter by CTCF. Cancer Res. 2007;67:2577–85.

    Google Scholar 

  39. Schmidt D, Schwalie PC, Wilson MD, Ballester B, Gonalves ngela, Kutter C, et al. Waves of retrotransposon expansion remodel genome organization and CTCF binding in multiple mammalian lineages. Cell. 2012;148:335–48.

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Stoilov P, Lin C-H, Damoiseaux R, Nikolic J, Black DL. A high-throughput screening strategy identifies cardiotonic steroids as alternative splicing modulators. Proc Natl Acad Sci USA. 2008;105:11218–23.

    CAS  PubMed  Google Scholar 

  41. Winteringham LN, Endersby R, Kobelke S, McCulloch RK, Williams JH, Stillitano J, et al. Myeloid leukemia factor 1 associates with a novel heterogeneous nuclear ribonucleoprotein U-like molecule. J Biol Chem. 2006;281:38791–38800.

    CAS  PubMed  Google Scholar 

  42. Falini B, Bigerna B, Pucciarini A, Tiacci E, Mecucci C, Morris SW, et al. Aberrant subcellular expression of nucleophosmin and NPM-MLF1 fusion protein in acute myeloid leukaemia carrying t(3;5): A comparison with NPMc+ AML. Leukemia. 2006;20:368–71.

    CAS  PubMed  Google Scholar 

  43. Spencer DH, Young MA, Lamprecht TL, Helton NM, Fulton R, O’Laughlin M, et al. Epigenomic analysis of the HOX gene loci reveals mechanisms that may control canonical expression patterns in AML and normal hematopoietic cells. Leukemia. 2015. https://doi.org/10.1038/leu.2015.6.

  44. Alcalay M, Tiacci E, Bergomas R, Bigerna B, Venturini E, Minardi SP, et al. Acute myeloid leukemia bearing cytoplasmic nucleophosmin (NPMc+AML) shows a distinct gene expression profile characterized by up-regulation of genes involved in stem-cell maintenance. Blood. 2005;106:899–902.

    CAS  PubMed  Google Scholar 

  45. Németh A, Längst G. Chromatin organization and the mammalian nucleolus. In: O’Day D., Catalano A. (eds) Proteins of the nucleolus: regulation, translocation & biomedical functions. Dordrecht: Springer; 2013, p. 119–48.

  46. Torrano V, Navascués J, Docquier F, Zhang R, Burke LJ, Chernukhin I, et al. Targeting of CTCF to the nucleolus inhibits nucleolar transcription through a poly(ADP-ribosyl)ation-dependent mechanism. J Cell Sci. 2006;119:1746–59.

    CAS  PubMed  Google Scholar 

  47. Docquier F, Kita GX, Farrar D, Jat P, O’Hare M, Chernukhin I, et al. Decreased poly(ADP-ribosyl)ation of CTCF, a transcription factor, is associated with breast cancer phenotype and cell proliferation. Clin Cancer Res. 2009;15:5762–71.

    CAS  PubMed  Google Scholar 

  48. Nakahashi H, Kwon KRK, Resch W, Vian L, Dose M, Stavreva D, et al. A genome-wide map of CTCF multivalency redefines the CTCF code. Cell Rep. 2013;3:1678–89.

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Essien K, Vigneau S, Apreleva S, Singh LN, Bartolomei MS, Hannenhalli S. CTCF binding site classes exhibit distinct evolutionary, genomic, epigenomic and transcriptomic features. Genome Biol. 2009;10:R131.

    PubMed  PubMed Central  Google Scholar 

  50. Plasschaert RN, Vigneau S, Tempera I, Gupta R, Maksimoska J, Everett L, et al. CTCF binding site sequence differences are associated with unique regulatory and functional trends during embryonic stem cell differentiation. Nucleic Acids Res. 2014;42:774–89.

    CAS  PubMed  Google Scholar 

  51. Xiao T, Wongtrakoongate P, Trainor C, Felsenfeld G. CTCF recruits centromeric protein CENP-E to the pericentromeric/centromeric regions of chromosomes through unusual CTCF-binding sites. Cell Rep. 2015;12:1704–14.

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Rubio ED, Reiss DJ, Welcsh PL, Disteche CM, Filippova GN, Baliga NS, et al. CTCF physically links cohesin to chromatin. Proc Natl Acad Sci USA. 2008;105:8309–14.

    CAS  PubMed  Google Scholar 

  53. Soto-Reyes E, Recillas-Targa F. Epigenetic regulation of the human p53 gene promoter by the CTCF transcription factor in transformed cell lines. Oncogene. 2010;29:2217–27.

    CAS  PubMed  Google Scholar 

  54. Fiorentino FP, Macaluso M, Miranda F, Montanari M, Russo A, Bagella L, et al. CTCF and BORIS regulate Rb2/p130 gene transcription: a novel mechanism and a new paradigm for understanding the biology of lung cancer. Mol Cancer Res. 2011;9:225–33.

    CAS  PubMed  Google Scholar 

  55. Gombert WM, Krumm A. Targeted deletion of multiple CTCF-binding elements in the human C-MYC gene reveals a requirement for CTCF in C-MYC expression. PLoS ONE 2009;4:1–8.

    Google Scholar 

  56. Khoury H, Suarez-Saiz F, Wu S, Minden MD. An upstream insulator regulates DLK1 imprinting in AML. Blood. 2010;115:2260–3.

    CAS  PubMed  Google Scholar 

  57. Marina RJ, Sturgill D, Bailly MA, Thenoz M, Varma G, Prigge MF, et al. TET-catalyzed oxidation of intragenic 5-methylcytosine regulates CTCF-dependent alternative splicing. EMBO J. 2016;35:335–55.

    CAS  PubMed  Google Scholar 

  58. Kosugi S, Hasebe M, Tomita M, Yanagawa H. Systematic identification of cell cycle-dependent yeast nucleocytoplasmic shuttling proteins by prediction of composite motifs. Proc Natl Acad Sci USA. 2009;106:10171–6.

    CAS  PubMed  Google Scholar 

  59. Nora EP, Goloborodko A, Valton AL, Gibcus JH, Uebersohn A, Abdennur N, et al. Targeted degradation of CTCF decouples local insulation of chromosome domains from genomic compartmentalization. Cell. 2017;169:930–944.e22.

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from Canadian Institute of Health Research, Leukemia Lymphoma Society of Canada. AJW is a PhD candidate at University of Toronto. This work has been submitted in partial fulfillment of the requirement for the PhD. MDM is supported by the Philip S Orsino Chair in Leukemia Research.

Author information

Authors and Affiliations

  1. Department of Medical Oncology and Hematology, Princess Margaret Cancer Center, University Health Network, Toronto, Canada

    Atom J. Wang, Youqi Han & Mark D. Minden

  2. Department of Medical Biophysics, Faculty of Medicine, University of Toronto, Toronto, Canada

    Atom J. Wang & Mark D. Minden

  3. Department of Computer Science, Faculty of Arts and Science, University of Toronto, Toronto, Canada

    Nanyang Jia & Peikun Chen

Authors
  1. Atom J. Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Youqi Han
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Nanyang Jia
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Peikun Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mark D. Minden
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

AJW, YH, PC, and NJ performed the experiments; AJW analyzed the results and made the figures; AJW and MDM designed the research and wrote the paper.

Corresponding author

Correspondence to Mark D. Minden.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Wang, A.J., Han, Y., Jia, N. et al. NPM1c impedes CTCF functions through cytoplasmic mislocalization in acute myeloid leukemia. Leukemia 34, 1278–1290 (2020). https://doi.org/10.1038/s41375-019-0681-8

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41375-019-0681-8

This article is cited by

Search

Advanced search

Quick links