TFAP2C regulates carbonic anhydrase XII in human breast cancer


The expression of carbonic anhydrase XII (CA12) is associated with the expression of estrogen receptor alpha (ERα) in breast cancer and is linked to a good prognosis with a lower risk of metastasis. Transcription Factor Activator Protein 2γ (TFAP2C, AP-2γ) governs luminal breast cancer phenotype through direct and indirect regulation of ERα and ERα-associated genes, GATA3, FOXA1, EGFR, CDH1, DSP, KRT7, FBP1, MYB, RET, KRT8, MUC1, and ERBB2—genes which are responsible for the luminal signature in breast cancer. Herein, utilizing chromatin immunoprecipitation and direct sequencing (ChIP-seq), we show that CA12 is regulated by AP-2γ through binding with its promoter region in luminal breast cancer cell lines and indirectly through a distal estrogen-responsive region in ERα-positive cell lines by upregulation of ERα. CA12 is transcriptionally silenced in basal breast cancer cell lines through histone deacetylation and CpG methylation of the promoter region and can be re-activated with Trichostatin A (histone deacetylase inhibitor) and/or 5-aza-dC (an inhibitor of DNA methylation). Strong concordance in co-expression of CA12 and ESR1 (R2 = 0.1128, p = 0486) and TFAP2C (R2 = 0.1823, p = 0.0105) was found using a panel of primary breast tumor samples (n = 35), supporting a synergetic role of AP-2γ and ERα in activation of CA12. Our results highlight the essential role of AP-2γ in maintaining the luminal breast cancer phenotype and provide evidence that epigenetic mechanisms silence luminal gene expression in the basal phenotype. Additional studies to decipher mechanisms that drive epigenetic silencing of AP-2γ target genes are a critical area for further research.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6


  1. 1.

    U.S. Cancer Statistics Working Group. U.S. Cancer Statistics Data Visualizations Tool, based on November 2017 submission data (1999–2015). U.S. Department of Health and Human Services, Centers for Disease Control and Prevention and National Cancer Institute; 2018.

  2. 2.

    Masood S. Breast cancer subtypes: morphologic and biologic characterization. Women’s Health. 2016;12:103–19.

  3. 3.

    Sorlie T, Perou CM, Tibshirani R, Aas T, Geisler S, Johnsen H, et al. Gene expression patterns of breast carcinomas distinguish tumor subclasses with clinical implications. Proc Natl Acad Sci USA. 2001;98:10869–74.

  4. 4.

    Supuran CT. Carbonic anhydrases—an overview. Curr Pharm Des. 2008;14:603–14.

  5. 5.

    Tashian RE. The carbonic anhydrases: widening perspectives on their evolution, expression and function. Bioessays. 1989;10:186–92.

  6. 6.

    Sly WS, Hu PY. Human carbonic anhydrases and carbonic anhydrase deficiencies. Annu Rev Biochem. 1995;64:375–401.

  7. 7.

    Parkkila S, Parkkila AK, Saarnio J, Kivela J, Karttunen TJ, Kaunisto K, et al. Expression of the membrane-associated carbonic anhydrase isozyme XII in the human kidney and renal tumors. J Histochem Cytochem. 2000;48:1601–8.

  8. 8.

    Ilie MI, Hofman V, Ortholan C, Ammadi RE, Bonnetaud C, Havet K, et al. Overexpression of carbonic anhydrase XII in tissues from resectable non-small cell lung cancers is a biomarker of good prognosis. Int J Cancer. 2010;128:1614–23.

  9. 9.

    Kivela A, Parkkila S, Saarnio J, Karttunen TJ, Kivela J, Parkkila AK, et al. Expression of a novel transmembrane carbonic anhydrase isozyme XII in normal human gut and colorectal tumors. Am J Pathol. 2000;156:577–84.

  10. 10.

    Nordfors K, Haapasalo J, Korja M, Niemelä A, Laine J, Parkkila AK, et al. The tumour-associated carbonic anhydrases CA II, CA IX and CA XII in a group of medulloblastomas and supratentorial primitive neuroectodermal tumours: an association of CA IX with poor prognosis. BMC Cancer. 2010;10:148.

  11. 11.

    Hynninen P, Vaskivuo L, Saarnio J, Haapasalo H, Kivela J, Pastoreková S, et al. Expression of transmembrane carbonic anhydrases IX and XII in ovarian tumours. Histopathology. 2006;49:594–602.

  12. 12.

    Wykoff CC, Beasley N, Watson PH, Campo L, Chia SK, English R, et al. Expression of the hypoxia-inducible and tumorassociated carbonic anhydrases in ductal carcinoma in situ of the breast. Am J Pathol. 2001;158:1011–9.

  13. 13.

    Haapasalo J, Hilvo M, Nordfors K, Haapasalo H, Parkkila S, Hyrskyluoto A, et al. Identification of an alternatively spliced isoform of carbonic anhydrase XII in diffusely infiltrating astrocytic gliomas. Neuro Oncol. 2008;10:131–8.

  14. 14.

    Watson PH, Chia SK, Wykoff CC, Han C, Leek RD, Sly WS, et al. Carbonic anhydrase XII is a marker of good prognosis in invasive breast carcinoma. Br J Cancer. 2003;88:1065–70.

  15. 15.

    Ilie MI, Hofman V, Ortholan C, Ammadi RE, Bonnetaud C, Havet K, et al. Overexpression of carbonic anhydrase XII in tissues from resectable non-small cell lung cancers is a biomarker of good prognosis. Int J Cancer. 2010;128:1614–23.

  16. 16.

    Kivela AJ, Parkkila S, Saarnio J, Karttunen TJ, Kivela J, Parkkila AK, et al. Expression of von Hippel-Lindau tumor suppressor and tumor-associated carbonic anhydrases IX and XII in normal and neoplastic colorectal mucosa. World J Gastroenterol. 2005;11:2616–25.

  17. 17.

    Barnett DH, Sheng S, Charn TH, Waheed A, Sly WS, Lin CY, et al. Estrogen receptor regulation of carbonic anhydrase XII through a distal enhancer in breast cancer. Cancer Res. 2008;68:3505–15.

  18. 18.

    Woodfield GW, Hitchler MJ, Chen Y, Domann FE, Weigel RJ. Interaction of TFAP2C with the estrogen receptor-alpha promoter is controlled by chromatin structure. Clin Cancer Res. 2009;15:3672–9.

  19. 19.

    Kulak MV, Cyr AR, Woodfield GW, Bogachek M, Spanheimer PM, Li T, et al. Transcriptional regulation of the GPX1 gene by TFAP2C and aberrant CpG methylation in human breast cancer. Oncogene. 2013;32:4043–51.

  20. 20.

    Cyr AR, Kulak MV, Park JM, Bogachek MV, Spanheimer PM, Woodfield GW, et al. TFAP2C governs the luminal epithelial phenotype in mammary development and carcinogenesis. Oncogene. 2015;34:436–44.

  21. 21.

    De Andrade JP, Park JM, Gu VW, Woodfield GW, Kulak MV, Lorenzen AW, et al. EGFR is regulated by TFAP2C in luminal breast cancer and is a target for vandetanib. Mol Cancer Ther. 2016;15:503–11.

  22. 22.

    Liu Q, Kulak MV, Borcherding N, Maina PK, Zhang W, Weigel RJ, et al. A novel HER2 gene body enhancer contributes to HER2 expression. Oncogene. 2018;37:687–94.

  23. 23.

    Kopecka J, Campia I, Jacobs A, Frei AP, Ghigo D, Wollscheid B, et al. Carbonic anhydrase XII is a new therapeutic target to overcome chemoresistance in cancer cells. Oncotarget. 2015;6:6776–93.

  24. 24.

    Woodfield GW, Horan AD, Chen Y, Weigel RJ. TFAP2C controls hormone response in breast cancer cells through multiple pathways of estrogen signaling. Cancer Res. 2007;67:8439–43.

  25. 25.

    McPherson LA, Weigel RJ. AP2alpha and AP2gamma: a comparison of binding site specificity and transactivation of the estrogen receptor promoter and single site promoter constructs. Nucleic Acids Res. 1999;27:4040–9.

  26. 26.

    Spanheimer PM, Woodfield GW, Cyr AR, Kulak MV, White-Baer LS, Bair TB, et al. Expression of the RET proto-oncogene is regulated by TFAP2C in breast cancer independent of the estrogen receptor. Ann Surg Oncol. 2013;20:2204–12.

  27. 27.

    Dai X, Cheng H, Bai Z, Li J. Breast cancer cell line classification and its relevance with breast tumor subtyping. J Cancer. 2017;8:3131–41.

  28. 28.

    Yersal O, Barutca S. Biological subtypes of breast cancer: prognostic and therapeutic implications. World J Clin Oncol. 2014;5:412–24.

  29. 29.

    Waheed A, Sly WS. Carbonic anhydrase XII functions in health and disease. Gene. 2017;623:33–40. Review.

  30. 30.

    von Neubeck B, Gondi G, Riganti C, Pan C, Parra Damas A, Scherb H, et al. An inhibitory antibody targeting carbonic anhydrase XII abrogates chemoresistance and significantly reduces lung metastases in an orthotopic breast cancer model in vivo. Int J Cancer. 2018;143:2065–75.

  31. 31.

    Kopecka J, Rankin GM, Salaroglio IC, Poulsen SA, Riganti C. P-glycoprotein-mediated chemoresistance is reversed by carbonic anhydrase XII inhibitors. Oncotarget. 2016;7:85861–75.

  32. 32.

    Kopecka J, Campia I, Jacobs A, Frei AP, Ghigo D, Wollscheid B, et al. Carbonic anhydrase XII is a new therapeutic target to overcome chemoresistance in cancer cells. Oncotarget. 2015;6:6776–93.

  33. 33.

    Palmer MB, Majumder P, Green MR, Wade PA, Boss JM. A 3′ enhancer controls snail expression in melanoma cells. Cancer Res. 2007;67:6113–20.

  34. 34.

    Neve RM, Chin K, Fridlyand J, Yeh J, Baehner FL, Fevr T, et al. A collection of breast cancer cell lines for the study of functionally distinct cancer subtypes. Cancer cell. 2006;10:515–27.

  35. 35.

    Kao J, Salari K, Bocanegra M, Choi YL, Girard L, Gandhi J, et al. Molecular profiling of breast cancer cell lines defines relevant tumor models and provides a resource for cancer gene discovery. PLoS ONE. 2009;4:e6146.

  36. 36.

    Weigelt B, Baehner FL, Reis-Filho JS. The contribution of gene expression profiling to breast cancer classification, prognostication and prediction: a retrospective of the last decade. J Pathol. 2010;220:263–80.

  37. 37.

    Sotiriou C, Pusztai L. Gene-expression signatures in breast cancer. N Engl J Med. 2009;360:790–800. Review.

  38. 38.

    O’Brien KM, Cole SR, Tse CK, Perou CM, Carey LA, Foulkes WD, et al. Intrinsic breast tumor subtypes, race, and long-term survival in the Carolina Breast Cancer Study. Clin Cancer Res. 2010;16:6100–10.

  39. 39.

    Brenton JD, Carey LA, Ahmed AA, Caldas C. Molecular classification and molecular forecasting of breast cancer: ready for clinical application? J Clin Oncol. 2005;23:7350–60.

  40. 40.

    Riaz M, van Jaarsveld MT, Hollestelle A, Prager-van der Smissen WJ, Heine AA, Boersma AW, et al. miRNA expression profiling of 51 human breast cancer cell lines reveals subtype and driver mutation-specific miRNAs. Breast cancer Res. 2013;15:R33.

  41. 41.

    Lacroix M, Haibe-Kains B, Hennuy B, Laes JF, Lallemand F, Gonze I, et al. Gene regulation by phorbol 12-myristate 13-acetate in MCF-7 and MDA-MB-231, two breast cancer cell lines exhibiting highly different phenotypes. Oncol Rep. 2004;12:701–7.

  42. 42.

    Yau C, Esserman L, Moore DH, Waldman F, Sninsky J, Benz CC. A multigene predictor of metastatic outcome in early stage hormone receptor-negative and triple-negative breast cancer. Breast Cancer Res. 2010;12:R85.

  43. 43.

    Edgar R, Domrachev M, Lash AE. Gene expression omnibus: NCBI gene expression and hybridization array data repository. Nucleic Acids Res. 2002;30:207–10.

  44. 44.

    Woodfield GW, Chen Y, Bair TB, Domann FE, Weigel RJ. Identification of primary gene targets of TFAP2C in hormone responsive breast carcinoma cells. Genes Chromosomes Cancer. 2010;49:948–62.

Download references


The authors would like to thank Zhaoming Wang for his excellent technical skills in cell culture. Also, we appreciate technical support provided by Keith D. Brendes.

Author information

Correspondence to Mikhail V. Kulak.

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

Verify currency and authenticity via CrossMark

Cite this article

Franke, C.M., Gu, V.W., Grimm, B.G. et al. TFAP2C regulates carbonic anhydrase XII in human breast cancer. Oncogene 39, 1290–1301 (2020).

Download citation