Promoter hypermethylation regulates UCHL1 expression in cholangiocarcinoma cell lines
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Abstract
Cholangiocarcinoma (CCA) is a significant health burden in Northeastern Thailand. Aberrant DNA methylation is a characteristic feature of most human cancers. Although overexpression of UCHL1 was observed in primary hilar CCA, the regulation of UCHL1 expression by DNA methylation in CCA has not yet been confirmed. We herein detected DNA methylation and mRNA expression levels in CCA and immortalized cholangiocyte cell lines using MS-HRM and RT-PCR, respectively. The results showed inverse correlation between DNA methylation levels and mRNA expression levels in CCA cell lines. To confirm whether DNA methylation regulated UCHL1 expression in CCA, 5-azacytidine was applied to inhibit DNA methylation process. We found that UCHL1 protein was up-regulated while DNA methylation was reduced after treatment suggesting the regulation of UCHL1 expression by DNA methylation. To study the role of UCHL1, CCA cell lines were transfected with vector harboring UCHL1 gene and control vector. Cell proliferation and chemotherapeutic drug sensitivity were investigated. The results showed no difference between these two groups. Our study reveals that overexpression of UCHL1 alone is not sufficient to neither inhibit tumor cell growth nor enhance chemoresponsiveness. Nevertheless, UCHL1 expression is epigenetically control indicating its function as a tumor suppressor by which the key features of UCHL1 and its partners in CCA are worth investigating.
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References
Valinluck V, Sowers LC. Endogenous cytosine damage products alter the site selectivity of human DNA maintenance methyltransferase DNMT1. Cancer Res 2007; 67(3): 946-50.
Abu-Remaileh M, Bender S, Raddatz G, Ansari I, Cohen D, Gutekunst J, et al. Chronic inflammation induces a novel epigenetic program that is conserved in intestinal adenomas and in colorectal cancer. Cancer Res 2015; 75(10): 2120-30.
Maiuri AR, Li H, Stein BD, Tennessen JM, O’Hagan HM. Inflammation-induced DNA methylation of DNA polymerase gamma alters the metabolic profile of colon tumors. Cancer Metab 2018; 6: 9.
Greger V, Passarge E, Hopping W, Messmer E, Horsthemke B. Epigenetic changes may contribute to the formation and spontaneous regression of retinoblastoma. Hum Genet 1989; 83(2): 155-8.
Brock MV, Hooker CM, Ota-Machida E, Han Y, Guo M, Ames S, et al. DNA methylation markers and early recurrence in stage I lung cancer. N Engl J Med 2008; 358(11): 1118-28.
Reyngold M, Turcan S, Giri D, Kannan K, Walsh LA, Viale A, et al. Remodeling of the methylation landscape in breast cancer metastasis. PLoS ONE 2014; 9(8): e103896.
Zhang C, Zhang B, Meng D, Ge C. Comprehensive analysis of DNA methylation and gene expression profiles in cholangiocarcinoma. Cancer Cell Int 2019; 19: 352.
Tsai H-C, Baylin SB. Cancer epigenetics: linking basic biology to clinical medicine. Cell Res 2011; 21(3): 502-17.
Pfeifer GP. Defining driver DNA methylation changes in human cancer. Int J Mol Sci 2018; 19(4): 1166.
Bishop P, Rocca D, Henley JM. Ubiquitin C-Terminal hydrolase L1 (UCH-L1): structure, distribution and roles in brain function and dysfunction. Biochem J 2016; 473(16): 2453-62.
Luo Y, He J, Yang C, Orange M, Ren X, Blair N, et al. UCH-L1 promotes invasion of breast cancer cells through activating Akt signaling pathway. J Cell Biochem 2018; 119(1): 691-700.
Kwan SY, Au-Yeung CL, Yeung TL, Rynne-Vidal A, Wong KK, Risinger JI, et al. Ubiquitin Carboxyl-Terminal Hydrolase L1 (UCHL1) promotes uterine serous cancer cell proliferation and cell cycle progression. cancers (Basel) 2020; 12(1): 118.
Nanok C, Jearanaikoon P, Proungvitaya S, Limpaiboon T. Aberrant methylation of HTATIP2 and UCHL1 as a predictive biomarker for cholangiocarcinoma. Mol Med Rep 2017; 17(3): 4145-53.
Zhong J, Zhao M, Ma Y, Luo Q, Liu j, Wang J, et al. UCHL1 acts as a colorectal cancer oncogene via activation of the β-catenin/TCF pathway through its deubiquitinating activity. Int J Mol Med 2012; 30(2): 430-6.
Xiang T, Li L, Yin X, Yuan C, Tan C, Su X, et al. The ubiquitin peptidase UCHL1 induces G0/G1 cell cycle arrest and apoptosis through stabilizing p53 and is frequently silenced in breast cancer. PLoS One 2012; 7(1): e29783.
Sripa B, Leungwattanawanit S, Nitta T, Nitta T, Wongkham C, Bhudhisawasdi V, et al. Establishment and characterization of an opisthorchiasis-associated cholangiocarcinoma cell line (KKU-100). World J Gastroenterol 2005; 11(22): 3392-7.
Maruyama M, Kobayashi N, Westerman KA, Sakaguchi M, Allain JE, Totsugawa T, et al. Establishment of a highly differentiated immortalized human cholangiocyte cell line with SV40T and hTERT. Transplantation 2004; 77(3): 446-51.
Schmittgen TD, Livak KJ. Analyzing real-time PCR data by the comparative CT method. Nature protocols 2008; 3: 1101-8.
Ummanni R, Jost E, Braig M, Lohmann F, Mundt F, Barett C, et al. Ubiquitin carboxyl-terminal hydrolase 1 (UCHL1) is a potential tumour suppressor in prostate cancer and is frequently silenced by promoter methylation. Mol Cancer 2011; 10(1): 129.
Kagara I, Enokida H, Kawakami K, Matsuda R, Toki K, Nishimura H, et al. CpG hypermethylation of the UCHL1 gene promoter is associated with pathogenesis and poor prognosis in renal cell carcinoma. J Urol 2008; 180(1): 343-51.
Okochi-Takada E, Nakazawa K, Wakabayashi M, Mori A, Ichimura S, Yasugi T, et al. Silencing of the UCHL1 gene in human colorectal and ovarian cancers. Int J Cancer 2006; 119(6): 1338-44.
Jin C, Yu W, Lou X, Zhou F, Han X, Zhao N, et al. UCHL1 is a putative tumor suppressor in ovarian cancer cells and contributes to cisplatin resistance. J Cancer 2013; 4(8): 662-70.
Li L, Tao Q, Jin H, Hasselt AV, Poon FF, Wang X, et al. The tumor suppressor UCHL1 forms a complex with p53/MDM2/ARF to promote p53 signaling and is frequently silenced in nasopharyngeal carcinoma. Clin Cancer Res 2010; 16(11): 2949-58.
Hematulin A, Meethang S, Utapom K, Wongkham S, Sagan D. Etoposide radiosensitizes p53-defective cholangiocarcinoma cell lines independent of their G2 checkpoint efficacies. Oncol Lett 2018; 15(3): 3895-903.
Yokochi T, Robertson KD. Dimethyl sulfoxide stimulates the catalytic activity of de novo DNA methyltransferase 3a (Dnmt3a) in vitro. Bioorg Chem 2004; 32(4): 234-43.
Yang G, Fan G, Zhang T, Ma K, Huang J, Liu M, et al. Upregulation of ubiquitin carboxylterminal hydrolase L1 (UCHL1) mediates the reversal effect of verapamil on chemoresistance to adriamycin of hepatocellular carcinoma. Med Sci Monit 2018; 24: 2072-82.
Chinnasri P, Pairojkul C, Jearanaikoon P, Sripa B, Bhudhisawasdi V, Tantimavanich S, et al. Preferentially different mechanisms of inactivation of 9p21 gene cluster in liver fluke–related cholangiocarcinoma. Hum Pathol 2009; 40(6): 817-26.