Tumor Biology

, Volume 37, Issue 6, pp 8293–8304 | Cite as

p21 and CK2 interaction-mediated HDAC2 phosphorylation modulates KLF4 acetylation to regulate bladder cancer cell proliferation

  • Zhuo-min Jia
  • Xing Ai
  • Jing-fei Teng
  • Yun-peng Wang
  • Bao-jun Wang
  • Xu Zhang
Original Article


Krüppel-like factor 4 (KLF4) is a transcription factor involved in both tumor suppression and oncogenesis as a transcriptional activator or repressor in a context-dependent manner. KLF4 acts as a regulator of p53 depending on p21 status in breast cancer. However, the mechanisms underlying the distinct role of KLF4 remain poorly understood. Here, we revealed that p21 depletion converted KLF4 from a cell cycle inhibitor to a promoter of bladder cancer cell proliferation. Additionally, KLF4 was acetylated in a p21-dependent manner to inhibit bladder cancer cell growth as a tumor suppressor. However, deacetylated KLF4 functioned as an oncogene promoting bladder cancer cell proliferation. Mechanistically, p21 and CK2 interaction, but not CK2 alone, enhanced HDAC2 phosphorylation and restricted KLF4 deacetylation and subsequent tumor promotion. Furthermore, we observed that KLF4 was acetylated by CBP/p300 and that overexpression of CBP resulted in KLF4 acetylation and tumor suppression even in p21-depleted bladder cancer cells. Moreover, we discovered that Notch-1 knockdown-induced KLF4 is acetylated form of KLF4, which may mediate Notch-1 function in bladder cancer cell proliferation. Our data demonstrate that KLF4 acts as a tumor suppressor or oncogene to activate or repress target gene transcription depending on its acetylation status, which is regulated by p21 and CK2 interaction-mediated HDAC2 phosphorylation. Targeting KLF4 at the post-transcriptional levels may provide novel insight for bladder cancer therapy.


CK2 HDAC2 phosphorylation KLF4 acetylation Notch-1 p21 



This project was supported by “Military General Hospital of Beijing PLA Innovation Cultivation Fund 2015-LC-13.”

Compliance with ethical standard

Conflicts of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Kaufman DS, Shipley WU, Feldman AS. Bladder cancer. Lancet. 2009;374:239–49.CrossRefPubMedGoogle Scholar
  2. 2.
    Urist MJ, Di Como CJ, Lu ML, Charytonowicz E, Verbel D, Crum CP, et al. Loss of p63 expression is associated with tumor progression in bladder cancer. The American Journal of Pathology. 2002;161:1199–206.CrossRefPubMedPubMedCentralGoogle Scholar
  3. 3.
    Junttila TT, Laato M, Vahlberg T, Soderstrom KO, Visakorpi T, Isola J, et al. Identification of patients with transitional cell carcinoma of the bladder overexpressing ErbB2, ErbB3, or specific erbb4 isoforms: real-time reverse transcription-pcr analysis in estimation of ErbB receptor status from cancer patients. Clinical Cancer Research. 2003;9:5346–57.PubMedGoogle Scholar
  4. 4.
    Habuchi T, Marberger M, Droller MJ, Hemstreet 3rd GP, Grossman HB, Schalken JA, et al. Prognostic markers for bladder cancer: International Consensus Panel on bladder tumor markers. Urology. 2005;66:64–74.CrossRefPubMedGoogle Scholar
  5. 5.
    Wolff EM, Liang G, Jones PA. Mechanisms of disease: genetic and epigenetic alterations that drive bladder cancer. Nature Clinical Practice Urology. 2005;2:502–10.CrossRefPubMedGoogle Scholar
  6. 6.
    Muto S, Horie S, Takahashi S, Tomita K, Kitamura T. Genetic and epigenetic alterations in normal bladder epithelium in patients with metachronous bladder cancer. Cancer Research. 2000;60:4021–5.PubMedGoogle Scholar
  7. 7.
    Tien YT, Chang MH, Chu PY, Lin CS, Liu CH, Liao AT. Downregulation of the KLF4 transcription factor inhibits the proliferation and migration of canine mammary tumor cells. Vet J. 2015.Google Scholar
  8. 8.
    An J, Golech S, Klaewsongkram J, Zhang Y, Subedi K, Huston GE, et al. Kruppel-like factor 4 (KLF4) directly regulates proliferation in thymocyte development and IL-17 expression during Th17 differentiation. FASEB Journal. 2011;25:3634–45.CrossRefPubMedPubMedCentralGoogle Scholar
  9. 9.
    Yusuf I, Kharas MG, Chen J, Peralta RQ, Maruniak A, Sareen P, et al. KLF4 is a FOXO target gene that suppresses b cell proliferation. International Immunology. 2008;20:671–81.CrossRefPubMedGoogle Scholar
  10. 10.
    Yang Y, Goldstein BG, Chao HH, Katz JP. KLF4 and KLF5 regulate proliferation, apoptosis and invasion in esophageal cancer cells. Cancer Biology & Therapy. 2005;4:1216–21.CrossRefGoogle Scholar
  11. 11.
    Zhang W, Geiman DE, Shields JM, Dang DT, Mahatan CS, Kaestner KH, et al. The gut-enriched Kruppel-like factor (Kruppel-like factor 4) mediates the transactivating effect of p53 on the P21WAF1/Cip1 promoter. The Journal of Biological Chemistry. 2000;275:18391–8.CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Yoon HS, Yang VW. Requirement of Kruppel-like factor 4 in preventing entry into mitosis following DNA damage. The Journal of Biological Chemistry. 2004;279:5035–41.CrossRefPubMedGoogle Scholar
  13. 13.
    Rowland BD, Bernards R, Peeper DS. The KLF4 tumour suppressor is a transcriptional repressor of p53 that acts as a context-dependent oncogene. Nature Cell Biology. 2005;7:1074–82.CrossRefPubMedGoogle Scholar
  14. 14.
    Black AR, Black JD, Azizkhan-Clifford J. Sp1 and Kruppel-like factor family of transcription factors in cell growth regulation and cancer. Journal of Cellular Physiology. 2001;188:143–60.CrossRefPubMedGoogle Scholar
  15. 15.
    Ohnishi S, Ohnami S, Laub F, Aoki K, Suzuki K, Kanai Y, et al. Downregulation and growth inhibitory effect of epithelial-type Kruppel-like transcription factor KLF4, but not KLF5, in bladder cancer. Biochemical and Biophysical Research Communications. 2003;308:251–6.CrossRefPubMedGoogle Scholar
  16. 16.
    Dang DT, Mahatan CS, Dang LH, Agboola IA, Yang VW. Expression of the gut-enriched Kruppel-like factor (Kruppel-like factor 4) gene in the human colon cancer cell line RKO is dependent on CDX2. Oncogene. 2001;20:4884–90.CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Zhao W, Hisamuddin IM, Nandan MO, Babbin BA, Lamb NE, Yang VW. Identification of Kruppel-like factor 4 as a potential tumor suppressor gene in colorectal cancer. Oncogene. 2004;23:395–402.CrossRefPubMedPubMedCentralGoogle Scholar
  18. 18.
    Wei D, Gong W, Kanai M, Schlunk C, Wang L, Yao JC, et al. Drastic down-regulation of Kruppel-like factor 4 expression is critical in human gastric cancer development and progression. Cancer Research. 2005;65:2746–54.CrossRefPubMedGoogle Scholar
  19. 19.
    Dang DT, Chen X, Feng J, Torbenson M, Dang LH, Yang VW. Overexpression of Kruppel-like factor 4 in the human colon cancer cell line RKO leads to reduced tumorigenecity. Oncogene. 2003;22:3424–30.CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Foster KW, Frost AR, McKie-Bell P, Lin CY, Engler JA, Grizzle WE, et al. Increase of GKLF messenger RNA and protein expression during progression of breast cancer. Cancer Research. 2000;60:6488–95.PubMedGoogle Scholar
  21. 21.
    Foster KW, Ren S, Louro ID, Lobo-Ruppert SM, McKie-Bell P, Grizzle W, et al. Oncogene expression cloning by retroviral transduction of adenovirus E1A-immortalized rat kidney RK3E cells: transformation of a host with epithelial features by c-myc and the zinc finger protein GKLF. Cell Growth & Differentiation. 1999;10:423–34.Google Scholar
  22. 22.
    Pandya AY, Talley LI, Frost AR, Fitzgerald TJ, Trivedi V, Chakravarthy M, et al. Nuclear localization of KLF4 is associated with an aggressive phenotype in early-stage breast cancer. Clinical Cancer Research. 2004;10:2709–19.CrossRefPubMedGoogle Scholar
  23. 23.
    Evans PM, Zhang W, Chen X, Yang J, Bhakat KK, Liu C. Kruppel-like factor 4 is acetylated by p300 and regulates gene transcription via modulation of histone acetylation. The Journal of Biological Chemistry. 2007;282:33994–4002.CrossRefPubMedGoogle Scholar
  24. 24.
    Huang Y, Chen J, Lu C, Han J, Wang G, Song C, et al. HDAC1 and Klf4 interplay critically regulates human myeloid leukemia cell proliferation. Cell Death & Disease. 2014;5:e1491.CrossRefGoogle Scholar
  25. 25.
    Meng F, Han M, Zheng B, Wang C, Zhang R, Zhang XH, et al. All-trans retinoic acid increases Klf4 acetylation by inducing HDAC2 phosphorylation and its dissociation from Klf4 in vascular smooth muscle cells. Biochemical and Biophysical Research Communications. 2009;387:13–8.CrossRefPubMedGoogle Scholar
  26. 26.
    Hu YY, Zheng MH, Zhang R, Liang YM, Han H. Notch signaling pathway and cancer metastasis. Advances in Experimental Medicine and Biology. 2012;727:186–98.CrossRefPubMedGoogle Scholar
  27. 27.
    Ai X, Jia Z, Liu S, Wang J, Zhang X. Notch-1 regulates proliferation and differentiation of human bladder cancer cell lines by inhibiting expression of Kruppel-like factor 4. Oncology Reports. 2014;32:1459–64.PubMedGoogle Scholar
  28. 28.
    Lesch HP, Laitinen A, Peixoto C, Vicente T, Makkonen KE, Laitinen L, et al. Production and purification of lentiviral vectors generated in 293T suspension cells with baculoviral vectors. Gene Therapy. 2011;18:531–8.CrossRefPubMedGoogle Scholar
  29. 29.
    Chen HY, Lin YM, Chung HC, Lang YD, Lin CJ, Huang J, et al. miR-103/107 promote metastasis of colorectal cancer by targeting the metastasis suppressors DAPK and KLF4. Cancer Research. 2012;72:3631–41.CrossRefPubMedGoogle Scholar
  30. 30.
    Wang J, Place RF, Huang V, Wang X, Noonan EJ, Magyar CE, et al. Prognostic value and function of KLF4 in prostate cancer: RNAa and vector-mediated overexpression identify KLF4 as an inhibitor of tumor cell growth and migration. Cancer Research. 2010;70:10182–91.CrossRefPubMedPubMedCentralGoogle Scholar
  31. 31.
    Yu F, Li J, Chen H, Fu J, Ray S, Huang S, et al. Kruppel-like factor 4 (KLF4) is required for maintenance of breast cancer stem cells and for cell migration and invasion. Oncogene. 2011;30:2161–72.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Yao K, Ki MO, Chen H, Cho YY, Kim SH, Yu DH, et al. Jnk1 and 2 play a negative role in reprogramming to pluripotent stem cells by suppressing KLF4 activity. Stem Cell Research. 2014;12:139–52.CrossRefPubMedGoogle Scholar
  33. 33.
    Segre CV, Chiocca S. Regulating the regulators: the post-translational code of class I HDAC1 and HDAC2. Journal of Biomedicine & Biotechnology. 2011;2011:690848.CrossRefGoogle Scholar
  34. 34.
    Adenuga D, Rahman I. Protein kinase CK2-mediated phosphorylation of HDAC2 regulates co-repressor formation, deacetylase activity and acetylation of HDAC2 by cigarette smoke and aldehydes. Archives of Biochemistry and Biophysics. 2010;498:62–73.CrossRefPubMedPubMedCentralGoogle Scholar
  35. 35.
    Tsai SC, Seto E. Regulation of histone deacetylase 2 by protein kinase CK2. The Journal of Biological Chemistry. 2002;277:31826–33.CrossRefPubMedGoogle Scholar
  36. 36.
    Dotto GP. P21(WAF1/CIP1): more than a break to the cell cycle? Biochimica et Biophysica Acta. 2000;1471:M43–56.PubMedGoogle Scholar
  37. 37.
    Gotz C, Wagner P, Issinger OG, Montenarh M. P21WAF1/CIP1 interacts with protein kinase CK2. Oncogene. 1996;13:391–8.PubMedGoogle Scholar
  38. 38.
    Glozak MA, Sengupta N, Zhang X, Seto E. Acetylation and deacetylation of non-histone proteins. Gene. 2005;363:15–23.CrossRefPubMedGoogle Scholar
  39. 39.
    Zhang XH, Zheng B, Gu C, Fu JR, Wen JK. TGF-beta1 downregulates AT1 receptor expression via PKC-delta-mediated Sp1 dissociation from KLF4 and Smad-mediated PPAR-gamma association with KLF4. Arteriosclerosis, Thrombosis, and Vascular Biology. 2012;32:1015–23.CrossRefPubMedGoogle Scholar
  40. 40.
    Kim MO, Kim SH, Cho YY, Nadas J, Jeong CH, Yao K, et al. ERK1 and ERK2 regulate embryonic stem cell self-renewal through phosphorylation of KLF4. Nature Structural & Molecular Biology. 2012;19:283–90.CrossRefGoogle Scholar
  41. 41.
    Kouzarides T. Acetylation: a regulatory modification to rival phosphorylation? The EMBO Journal. 2000;19:1176–9.CrossRefPubMedPubMedCentralGoogle Scholar
  42. 42.
    Zheng B, Han M, Shu YN, Li YJ, Miao SB, Zhang XH, et al. HDAC2 phosphorylation-dependent KLF5 deacetylation and RARalpha acetylation induced by RAR agonist switch the transcription regulatory programs of p21 in VSMCs. Cell Research. 2011;21:1487–508.CrossRefPubMedPubMedCentralGoogle Scholar
  43. 43.
    Ortega CE, Seidner Y, Dominguez I. Mining CK2 in cancer. PloS One. 2014;9:e115609.CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2016

Authors and Affiliations

  1. 1.Department of UrologyClinical Division of Surgery, Chinese PLA General HospitalBeijingChina
  2. 2.Department of UrologyMilitary General Hospital of Beijing PLABeijingChina

Personalised recommendations