目的:探究miR-152-3p、DNA甲基转移酶1(DNA methyltransferases, DNMT1)在结肠癌细胞中的调控机制。方法:qRT-PCR检测miR-152-3p和DNMT1的表达;细胞增殖实验、克隆形成实验、划痕愈合实验以及Transwell实验检测各处理组癌细胞的增殖、迁移和侵袭能力;双荧光素酶实验检测miR-152-3p与DNMT1的靶向关系;Western blot实验检测DNMT1蛋白表达;流式细胞术实验检测细胞凋亡率。结果:结肠癌细胞中miR-152-3p显著低表达,DNMT1显著高表达;过表达miR-152-3p会抑制结肠癌细胞增殖、迁移和侵袭;miR-152-3p能靶向抑制DNMT1的表达,从而抑制结肠癌细胞的增殖、迁移和侵袭;过表达miR-152-3p可以靶向DNMT1并促进细胞凋亡。结论:miR-152-3p通过靶向下调DNMT1的表达,从而抑制结肠癌细胞的发生发展。
Objective: To investigate the regulatory mechanism of miR-152-3p and DNA Methyltransferases(DNMT1) in colon cancer cells. Methods: qRT-PCR was used to detect the expression of miR-152-3p and DNMT1. Cell proliferation assay, colony formation assay, scratch healing assay and Transwell assay were used to detect the proliferation, migration and invasion ability of cancer cells in each treatment group. Dual luciferase assay was used to detect the targeting relationship between miR-152-3p and DNMT1. Western blot assay was used to detect the protein expression of DNMT1. Flow cytometry assay was used to detect apoptosis rate of cells. Results: MiR-152-3p was significantly downregulated while DNMT1 was highly expressed in colon cancer cells. Overexpression of miR-152-3p inhibited the proliferation, migration and invasion of colon cancer cells. MiR-152-3p could inhibit the expression of DNMT1, thereby inhibiting the proliferation, migration and invasion of colon cancer cells. Overexpression of miR-152-3p could target DNMT1 and promote cell apoptosis. Conclusion: MiR-152-3p could downregulate the expression of DNMT1, thereby inhibiting the development of colon cancer cells.
[1] Brody H. Colorectal cancer[J]. Nature, 2015,521(7551): S1.
[2] MármoL I, Sánchez-De-Diego C, Pradilla Dieste A, et al. Colorectal carcinoma: a general overview and future perspectives in colorectal cancer[J]. Int J Mol Sci, 2017,18(1): 197.
[3] Flavahan WA, Gaskell E, Bernstein BE. Epigenetic plasticity and the hallmarks of cancer[J]. Science, 2017, 357(6348): eaal2380.
[4] Hanahan D, Weinberg RA. The hallmarks of cancer[J]. Cell, 2000, 100(1): 57-70.
[5] Wu CT, Lin WY, Chang YH, et al. DNMT1-dependent suppression of microRNA424 regulates tumor progression in human bladder cancer[J]. Oncotarget, 2015, 6(27): 24119-24131.
[6] Sun J, Tian XH, Zhang JQ, et al. Regulation of human glioma cell apoptosis and invasion by miR-152-3p through targeting DNMT1 and regulating NF2: miR-152-3p regulate glioma cell apoptosis and invasion[J]. J Exp Clin Cancer Res, 2017, 36(1): 100.
[7] Seo JS, Choi YH, Moon JW, et al. Hinokitiol induces DNA demethylation via DNMT1 and UHRF1 inhibition in colon cancer cells[J]. BMC Cell Biol, 2017, 18(1): 14.
[8] Lu ZW, Du MY, Qian LX, et al. miR-152 functioning as a tumor suppressor that interacts with DNMT1 in nasopharyngeal carcinoma[J]. Onco Targets Ther, 2018, 11: 1733-1741.
[9] Moya LR, Meijer J, Schubert S, et al. Assessment of miR-98-5p, miR-152-3p, miR-326 and miR-4289 expression as biomarker for prostate cancer diagnosis[J]. Int J Mol Sci, 2019, 20(5): 1154.
[10] Balacescu O, Sur D, Cainap C, et al. The impact of miRNA in colorectal cancer progression and its liver metastases[J]. Int J Mol Sci, 2018,19(12):3711.
[11] Ma J, Yao YL, Wang P, et al. miR-152 functions as a tumor suppressor in glioblastoma stem cells by targeting Krüppel-like factor 4[J]. Cancer Lett, 2014, 355(1): 85-95.
[12] Azizi M, Teimoori-Toolabi L, Arzanani MK, et al. microRNA-148b and microRNA-152 reactivate tumor suppressor genes through suppression of DNA methyltransferase-1 gene in pancreatic cancer cell lines[J]. Cancer Biol Ther, 2014, 15(4): 419-427.
[13] Ramalho-Carvalho J, Gonçalves CS, Graça I, et al. A multiplatform approach identifies miR-152-3p as a common epigenetically regulated onco-suppressor in prostate cancer targeting TMEM97[J]. Clin Epigenetics, 2018, 10: 40.
[14] Chen X, Wang YW, Gao P. SPIN1, negatively regulated by miR-148/152, enhances Adriamycin resistance via upregulating drug metabolizing enzymes and transporter in breast cancer[J]. J Exp Clin Cancer Res, 2018, 37(1): 100.
[15] Wang CC, Ma XJ, Zhang JY, et al. DNMT1 maintains the methylation of miR-152-3p to regulate TMSB10 expression, thereby affecting the biological characteristics of colorectal cancer cells[J]. IUBMB Life, 2020, 72(11): 2432-2443.
[16] Wong KK. DNMT1:a key drug target in triple-negative breast cancer[J]. Semin Cancer Biol, 2021,72: 198-213.
[17] Du WW, Yang WN, Li XM, et al. A circular RNA circ-DNMT1 enhances breast cancer progression by activating autophagy[J]. Oncogene, 2018, 37(44): 5829-5842.
[18] Wang P, Chu WH, Zhang X, et al. Kindlin-2 interacts with and stabilizes DNMT1 to promote breast cancer development[J]. Int J Biochem Cell Biol, 2018, 105: 41-51.
[19] Hu YB, Ma Z, He YM, et al. LncRNA-SNHG1 contributes to gastric cancer cell proliferation by regulating DNMT1[J]. Biochem Biophys Res Commun, 2017, 491(4): 926-931.
[20] Wang H, Wu JX, Meng XQ, et al. microRNA-342 inhibits colorectal cancer cell proliferation and invasion by directly targeting DNA methyltransferase 1[J]. Carcinogenesis, 2011, 32(7): 1033-1042.
[21] Han GD, Wei ZJ, Cui HB, et al. NUSAP1 gene silencing inhibits cell proliferation, migration and invasion through inhibiting DNMT1 gene expression in human colorectal cancer[J]. Exp Cell Res, 2018, 367(2): 216-221.
