Analysis of promoter methylation and epigenetic regulation of miR‑32 in colorectal cancer cells

Wu,Weiyun; Ye,Shicai; Tan,Wenkai; Zhou,Yu; Quan,Juanhua

doi:10.3892/etm.2019.7328

Analysis of promoter methylation and epigenetic regulation of miR‑32 in colorectal cancer cells

Authors:
- Weiyun Wu
- Shicai Ye
- Wenkai Tan
- Yu Zhou
- Juanhua Quan
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Affiliations: Department of Gastroenterology, Affiliated Hospital of Guangdong Medical University, Zhanjiang, Guangdong 524001, P.R. China
Published online on: February 28, 2019 https://doi.org/10.3892/etm.2019.7328
Pages: 3209-3214

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Abstract

MicroRNA‑32 (miR‑32) is upregulated in colorectal cancer (CRC) tissues; its overexpression leads to increased cell proliferation, migration and invasion, as well as reduced apoptosis of CRC cells, at least partly by inhibiting the target gene phosphatase and tensin homolog. However, the mechanisms of its upregulation have remained elusive. In the present study, the effects of methylation and acetylation on the expression of miR‑32 were investigated. The promoter methylation status of miR‑32 in the CRC cell lines HT‑29 and HCT‑116 and the normal colonic epithelial cell line NCM460 was investigated by bisulfate sequencing polymerase chain reaction (BSP). The potential role of methylation and histone acetylation in the regulation of miR‑32 expression in CRC cells was investigated using the demethylation reagent 5‑aza‑2'‑deoxycytidine (5‑Aza‑dC), the histone deacetylase inhibitor trichostatin A (TSA) and transfection of DNA methyltransferase 1 (DNMT1) overexpression plasmid. BSP revealed that CpG sites in the miR‑32 promoter region of CRC and normal colonic epithelial cells were all hypomethylated, with methylation rates of 0.12, 1.14 and 0.64% in HCT‑116, HT‑29 and NCM460 cells, respectively. Treatment with 5‑Aza‑dC and/or TSA and transfection with DNMT1 plasmid did not significantly alter the expression of miR‑32. Therefore, the present results suggest that methylation and histone acetylation do not affect miR‑32 expression in CRC cells.

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1	Vogelstein B, Fearon ER, Hamilton SR, Kern SE, Preisinger AC, Leppert M, Nakamura Y, White R, Smits AM and Bos JL: Genetic alterations during colorectal-tumor development. N Engl J Med. 319:525–532. 1988. View Article : Google Scholar : PubMed/NCBI
2	Ha M and Kim VN: Regulation of microRNA biogenesis. Nat Rev Mol Cell Biol. 15:509–524. 2014. View Article : Google Scholar : PubMed/NCBI
3	Zhang W, Liu J and Wang G: The role of microRNAs in human breast cancer progression. Tumour Biol. 35:6235–6344. 2014. View Article : Google Scholar : PubMed/NCBI
4	Zheng Q, Chen C, Guan H, Kang W and Yu C: Prognostic role of microRNAs in human gastrointestinal cancer: A systematic review and meta-analysis. Oncotarget. 8:46611–46623. 2017.PubMed/NCBI
5	Moore LD, Le T and Fan G: DNA methylation and its basic function. Neuropsychopharmacology. 38:23–38. 2013. View Article : Google Scholar : PubMed/NCBI
6	Tse JWT, Jenkins LJ, Chionh F and Mariadason JM: Aberrant DNA Methylation in colorectal cancer: What should we target? Trends Cancer. 3:698–712. 2017. View Article : Google Scholar : PubMed/NCBI
7	Wang W, Ji G, Xiao X, Chen X, Qin WW, Yang F, Li YF, Fan LN, Xi WJ, Huo Y, et al: Epigenetically regulated miR-145 suppresses colon cancer invasion and metastasis by targeting LASP1. Oncotarget. 7:68674–68687. 2016.PubMed/NCBI
8	Yan-Fang T, Jian N, Jun L, Na W, Pei-Fang X, Wen-Li Z, Dong W, Li P, Jian W, Xing F and Jian P: The promoter of miR-663 is hypermethylated in Chinese pediatric acute myeloid leukemia (AML). BMC Med Genet. 14:742013. View Article : Google Scholar : PubMed/NCBI
9	Spange S, Wagner T, Heinzel T and Krämer OH: Acetylation of non-histone proteins modulates cellular signalling at multiple levels. Int J Biochem Cell Biol. 41:185–198. 2009. View Article : Google Scholar : PubMed/NCBI
10	Poddar S, Kesharwani D and Datta M: Histone deacetylase inhibition regulates miR-449a levels in skeletal muscle cells. Epigenetics. 11:579–587. 2016. View Article : Google Scholar : PubMed/NCBI
11	Wu W, Yang J, Feng X, Wang H, Ye S, Yang P, Tan W, Wei G and Zhou Y: MicroRNA-32 (miR-32) regulates phosphatase and tensin homologue (PTEN) expression and promotes growth, migration, and invasion in colorectal carcinoma cells. Mol Cancer. 12:302013. View Article : Google Scholar : PubMed/NCBI
12	Wu W, Yang P, Feng X, Wang H, Qiu Y, Tian T, He Y, Yu C, Yang J, Ye S and Zhou Y: The relationship between and clinical significance of MicroRNA-32 and phosphatase and tensin homologue expression in colorectal cancer. Genes Chromosomes Cancer. 52:1130–1140. 2013. View Article : Google Scholar
13	Mizuno Y, Maemura K, Tanaka Y, Hirata A, Futaki S, Hamamoto H, Taniguchi K, Hayashi M, Uchiyama K, Shibata MA, et al: Expression of delta-like 3 is downregulated by aberrant DNA methylation and histone modification in hepatocellular carcinoma. Oncol Rep. 39:2209–2216. 2018.PubMed/NCBI
14	Guo LH, Li H, Wang F, Yu J and He JS: The tumor suppressor roles of miR-433 and miR-127 in gastric cancer. Int J Mol Sci. 14:14171–14184. 2013. View Article : Google Scholar : PubMed/NCBI
15	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(−Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
16	Weber M and Schubeler D: Genomic patterns of DNA methylation: Targets and function of an epigenetic mark. Curr Opin Cell Biol. 19:273–280. 2007. View Article : Google Scholar : PubMed/NCBI
17	Lopez-Serra P and Esteller M: DNA methylation-associated silencing of tumor-suppressor microRNAs in cancer. Oncogene. 31:1609–1622. 2012. View Article : Google Scholar : PubMed/NCBI
18	Wang Z, Yao H, Lin S, Zhu X, Shen Z, Lu G, Poon WS, Xie D, Lin MC and Kung HF: Transcriptional and epigenetic regulation of human microRNAs. Cancer Lett. 331:1–10. 2013. View Article : Google Scholar : PubMed/NCBI
19	Xia Z, Qiu D, Deng J, Jiao X, Yang R, Sun Z, Wan X and Li J: Methylation-induced downregulation and tumor-suppressive role of microRNA-98 in glioma through targeting Sal-like protein 4. Int J Mol Med. 41:2651–2659. 2018.PubMed/NCBI
20	Shindo T, Niinuma T, Nishiyama N, Shinkai N, Kitajima H, Kai M, Maruyama R, Tokino T, Masumori N and Suzuki H: Epigenetic silencing of miR-200b is associated with cisplatin resistance in bladder cancer. Oncotarget. 9:24457–24469. 2018. View Article : Google Scholar : PubMed/NCBI
21	Song WF, Wang L, Huang WY, Cai X, Cui JJ and Wang LW: MiR-21 upregulation induced by promoter zone histone acetylation is associated with chemoresistance to gemcitabine and enhanced malignancy of pancreatic cancer cells. Asian Pac J Cancer Prev. 14:7529–7536. 2013. View Article : Google Scholar : PubMed/NCBI
22	Wang F, Liu M, Li X and Tang H: MiR-214 reduces cell survival and enhances cisplatin-induced cytotoxicity via down-regulation of Bcl2l2 in cervical cancer cells. FEBS Lett. 587:488–495. 2013. View Article : Google Scholar : PubMed/NCBI
23	Ambs S, Prueitt RL, Yi M, Hudson RS, Howe TM, Petrocca F, Wallace TA, Liu CG, Volinia S, Calin GA, et al: Genomic profiling of microRNA and messenger RNA reveals deregulated microRNA expression in prostate cancer. Cancer Res. 68:6162–6170. 2008. View Article : Google Scholar : PubMed/NCBI
24	Erdogan B, Bosompem A, Peng D, Han L, Smith E, Kennedy ME, Alford CE, Wu H, Zhao Z, Mosse CA, et al: Methylation of promoters of microRNAs and their host genes in myelodysplastic syndromes. Leuk Lymphoma. 54:2720–2727. 2013. View Article : Google Scholar : PubMed/NCBI
25	Rodriguez A, Griffiths-Jones S, Ashurst JL and Bradley A: Identification of mammalian microRNA host genes and transcription units. Genome Res. 14:1902–1910. 2004. View Article : Google Scholar : PubMed/NCBI
26	Yeung CL, Tsang TY, Yau PL and Kwok TT: Human papillomavirus type 16 E6 suppresses microRNA-23b expression in human cervical cancer cells through DNA methylation of the host gene C9orf3. Oncotarget. 8:12158–12173. 2017. View Article : Google Scholar : PubMed/NCBI
27	Chen CQ, Chen CS, Chen JJ, Zhou LP, Xu HL, Jin WW, Wu JB and Gao SM: Histone deacetylases inhibitor trichostatin A increases the expression of Dleu2/miR-15a/16-1 via HDAC3 in non-small cell lung cancer. Mol Cell Biochem. 383:137–148. 2013. View Article : Google Scholar : PubMed/NCBI
28	Yang H, Lan P, Hou Z, Guan Y, Zhang J, Xu W, Tian Z and Zhang C: Histone deacetylase inhibitor SAHA epigenetically regulates miR-17–92 cluster and MCM7 to upregulate MICA expression in hepatoma. Br J Cancer. 112:112–121. 2015. View Article : Google Scholar : PubMed/NCBI