DNMT3a通过提升基因内部甲基化介导紫杉醇诱导的LINE-1异常表达
收稿日期: 2019-10-29
修回日期: 2019-12-27
网络出版日期: 2020-01-07
基金资助
国家自然科学基金项目资助编号：(81172091)
DNMT3a mediates paclitaxel-induced abnormal expression of LINE-1 by increasing the intragenic methylation
Received date: 2019-10-29
Revised date: 2019-12-27
Online published: 2020-01-07
Supported by
Supported by the National Natural Science Foundation of China No(81172091)
药物诱导的长散在重复序列LINE-1异常激活可促进细胞基因组不稳定,而基因组不稳定是促进肿瘤发生发展和耐药表型形成的重要因素。因此,探索LINE-1异常激活的分子机制具有重要的理论和临床意义。DNA甲基化是调控基因表达的重要方式,已知DNA甲基转移酶家族成员DNMT3a不仅能通过促进基因启动子甲基化抑制基因表达,还可通过增强基因内部甲基化上调基因表达。本实验室前期研究发现,将乳腺癌细胞暴露于化疗药物可诱导LINE-1异常高表达,但LINE-1启动子甲基化水平并无显著改变。本研究进一步探讨了在化疗药物压力下DNMT3a是否可通过增强LINE-1基因内部甲基化水平促进LINE-1在乳腺癌细胞中的异常高表达。ChIP实验和甲基分析结果显示,用化疗药物紫杉醇(PTX)处理乳腺癌细胞,不仅可以诱导DNMT3a表达,而且可以促进DNMT3a与LINE-1基因内部区域的结合,提升其基因内部甲基化水平, 进而上调LINE-1的表达水平。利用表达载体增加细胞内DNMT3a的表达水平,可显著上调LINE-1基因内部的甲基化及基因的表达水平,而下调DNMT3a的表达可有效抑制LINE-1表达。上述研究结果表明,DNMT3a介导的基因非启动子区甲基化在药物诱导的LINE-1异常激活中发挥重要作用,为认识LINE-1在乳腺癌化疗耐药性形成过程中异常激活的机制提供了新思路。
关键词： 乳腺癌; LINE-1异常激活; 基因内部DNA甲基化; DNMT3a
王昕源, 张雨, 杨楠, 程禾, 孙玉洁 . DNMT3a通过提升基因内部甲基化介导紫杉醇诱导的LINE-1异常表达[J]. 遗传, 2020 , 42(1) : 100 -111 . DOI: 10.16288/j.yczz.19-258
The activation of long interspersed nuclear element-1 (LINE-1) leads to genomic instability, which promotes carcinogenesis and drug resistant. Therefore, exploring the mechanism underlying LINE-1 abnormal activation has the theoretical and clinical significance. DNA methylation is an important way to regulate gene expression. DNMT3a, one member of the DNA methyltransferase family, not only inhibits gene expression by inducing promoter hypermethylation, but also activates gene expression by increasing the intragenic DNA methylation. Our previous studies found that the expression of LINE-1 did not increase significantly in the promoter methylation in breast cancer cells treated with paclitaxel (PTX), a first-line chemotherapeutic drug for breast cancer. Here we explored whether DMNMT3a could directly mediate the drug-induced activation of LINE-1 in breast cancer cells through increasing the LINE-1 intragenic methylation. Our ChIP experiments and methyl analysis showed that treatment of breast cancer cells with PTX not only induced DNMT3a expression, but also promoted the binding of DNMT3a to the inner region of the LINE-1 gene to increase its methylation, resulting in upregulation of LINE-1 expression. Using expression vectors or RNA interference to alter the DNMT3a expression levels in the cells significantly changed the intragenic methylation degree and LINE-1 expression. Moreover, down-regulation of DNMT3a expression effectively inhibited the expression of LINE-1. These results indicate that DNMT3a-mediated intragenic methylation plays an important role in drug-induced abnormal activation of LINE-1, which provides a new idea for understanding the mechanism of abnormal activation of Line-1 induced by chemotherapy drug stress in breast cancer cells.
| [1] | Wei R, Lau SSS, Cheung PSY . Breast carcinoma in chinese women: Does age affect treatment choice and outcome? Asian J Surg, 2010,33(2):97-102. |
| [2] | Ferlay J, Shin HR, Bray F, Forman D, Mathers C, Parkin DM . Estimates of worldwide burden of cancer in 2008: GLOBOCAN 2008. Int J Cancer, 2010,127(12):2893-2917. |
| [3] | Clarke R, Tyson JJ, Dixon JM. Endocrine resistance in breast cancer--An overview and update. Mol Cell Endocrinol, 2015, 418 Pt 3: 220-234. |
| [4] | Kuiper GG, Enmark E, Pelto-Huikko M, Nilsson S, Gustafsson JA . Cloning of a novel receptor expressed in rat prostate and ovary. Proc Natl Acad Sci USA, 1996,93(12):5925-5930. |
| [5] | Rivera E . Management of metastatic breast cancer: monotherapy options for patients resistant to anthracyclines and taxanes. Am J Clin Oncol, 2010,33(2):176-185. |
| [6] | Bao WD, Kojima KK, Kohany O . Repbase update, a database of repetitive elements in eukaryotic genomes. Mob DNA, 2015,6:11. |
| [7] | Belancio VP, Hedges DJ, Deininger P . Mammalian non-LTR retrotransposons: for better or worse, in sickness and in health. Genome Res, 2008,18(3):343-358. |
| [8] | Beck CR, Garcia-Perez JL, Badge RM, Moran JV . LINE-1 elements in structural variation and disease. Annu Rev Genomics Hum Genet, 2011,12:187-215. |
| [9] | Liu Q, Wang JH, Li XY, Cen S,. The connection between LINE-1 retrotransposition and human tumorigenesis. Hereditas(Beijing), 2016,38(2):93-102. |
| [9] | 刘茜, 王瑾晖, 李晓宇, 岑山 . 逆转录转座子LINE-1与肿瘤的发生和发展. 遗传, 2016,38(2):93-102. |
| [10] | Belgnaoui SM, Gosden RG, Semmes OJ, Haoudi A . Human LINE-1 retrotransposon induces DNA damage and apoptosis in cancer cells. Cancer Cell Int, 2006,6:13. |
| [11] | Feng Q, Moran JV, Kazazian HH Jr, Boeke JD . Human L1 retrotransposon encodes a conserved endonuclease required for retrotransposition. Cell, 1996,87(5):905-916. |
| [12] | Martin SL, Li J, Epperson LE, Lieberman B . Functional reverse transcriptases encoded by A-type mouse LINE-1: defining the minimal domain by deletion analysis. Gene, 1998,215(1):69-75. |
| [13] | Farkash EA, Kao GD, Horman SR, Prak ET . Gamma radiation increases endonuclease-dependent L1 retrotransposition in a cultured cell assay. Nucleic Acids Res, 2006,34(4):1196-1204. |
| [14] | Gasior SL, Wakeman TP, Xu B, Deininger PL . The human LINE-1 retrotransposon creates DNA double-strand breaks. . Mol Biol, 2006,357(5):1383-1393. |
| [15] | Garcia-Perez JL, Doucet AJ, Bucheton A, Moran JV, Gilbert N . Distinct mechanisms for trans-mediated mobilization of cellular RNAs by the LINE-1 reverse transcriptase. Genome Res, 2007,17(5):602-611. |
| [16] | Wei W, Gilbert N, Ooi SL, Lawler JF, Ostertag EM, Kazazian HH, Boeke JD, Moran JV . Human L1 retrotransposition: cis preference versus trans complementation. Mol Cell Biol, 2001,21(4):1429-1439. |
| [17] | Kines KJ, Sokolowski M, deHaro DL, Christian CM, Baddoo M, Smither ME, Belancio VP,. The endonuclease domain of the LINE-1 ORF2 protein can tolerate multiple mutations. Mob DNA, 2016,7:8. |
| [18] | Okano M, Xie S, Li E . Cloning and characterization of a family of novel mammalian DNA (cytosine-5) methyltransferases. Nat Genet, 1998,19(3):219-220. |
| [19] | Brusa G, Benvenuti M, Mazzacurati L, Mancini M, Pattacini L, Martinelli G, Barbieri E, Greenberger JS, Baccarani M, Santucci MA. p53 loss of function enhances genomic instability and accelerates clonal evolution of murine myeloid progenitors expressing the p(210)BCR-ABL tyrosine kinase. Haematologica, 2003,88(6):622-630. |
| [20] | Greaves M, Maley CC . Clonal evolution in cancer. Nature, 2012,481(7381):306-313. |
| [21] | Ng CK, Pemberton HN, Reis-Filho JS . Breast cancer intratumor genetic heterogeneity: causes and implications. Expert Rev Anticancer Ther, 2012,12(8):1021-1032. |
| [22] | Chen L, Dahlstrom JE, Chandra A, Board P, Rangasamy D . Prognostic value of LINE-1 retrotransposon expression and its subcellular localization in breast cancer. Breast Cancer Res Treat, 2012,136(1):129-142. |
| [23] | Park SY, Seo AN, Jung HY, Gwak JM, Jung N, Cho NY, Kang GH . Alu and LINE-1 hypomethylation is associated with HER2 enriched subtype of breast cancer. PLoS One, 2014,9(6):e100429. |
| [24] | Feng F, Lu YY, Zhang F, Gao XD, Zhang CF, Meredith A, Xu ZX, Yang YT, Chang XJ, Wang H, Qu JH, Zeng Z, Yang JL, Wang CP, Zhu YF, Cui JJ, Yang YP . Long interspersed nuclear element ORF-1 protein promotes proliferation and resistance to chemotherapy in heaptocellular carcinoma. World J Gastroenterol, 2013,19(7):1068-1078. |
| [25] | Chen Y, Zeng Q, Liu X, Fu J, Zeng Z, Zhao Z, Liu Z, Bai W, Dong Z, Liu H, Lu X, Zhu Y, Lu Y . LINE-1 ORF-1p enhances the transcription factor activity of pregnenolone X receptor and promotes sorafenib resistance in hepatocellular carcinoma cells. Cancer Manag Res, 2018,10:4421-4438. |
| [26] | Suzuki MM, Bird A . DNA methylation landscapes: provocative insights from epigenomics. Nat Rev Genet, 2008,9(6):465-476. |
| [27] | Jin B, Robertson KD . DNA methyltransferases, DNA damage repair, and cancer. Adv Exp Med Biol, 2013,754:3-29. |
| [28] | Yang L, Rau R, Goodell MA . DNMT3A in haematological malignancies. Nat Rev Cancer, 2015,15(3):152-165. |
| [29] | Jeong M, Sun D, Luo M, Huang Y, Challen GA, Rodriguez B, Zhang X, Chavez L, Wang H, Hannah R, Kim SB, Yang L, Ko M, Chen R, G?ttgens B, Lee JS, Gunaratne P, Godley LA, Darlington GJ, Rao A, Li W, Goodell MA . Large conserved domains of low DNA methylation maintained by Dnmt3a. Nat Genet, 2014,46(1):17-23. |
| [30] | Wu H, Coskun V, Tao J, Xie W, Ge W, Yoshikawa K, Li E, Zhang Y, Sun YE . Dnmt3a-dependent nonpromoter DNA methylation facilitates transcription of neurogenic genes. Science, 2010,329(5990):444-448. |
| [31] | Yang X, Han H, De Carvalho DD, Lay FD, Jones PA, Liang G . Gene body methylation can alter gene expression and is a therapeutic target in cancer. Cancer Cell, 2014,26(4):577-590. |
| [32] | Yoder JA, Walsh CP, Bestor TH . Cytosine methylation and the ecology of intragenomic parasites. Trends Genet, 1997,13(8):335-340. |
| [33] | Ray Chaudhuri A, Callen E, Ding X, Gogola E, Duarte AA, Lee JE, Wong N, Lafarga V, Calvo JA, Panzarino NJ, John S, Day A, Crespo AV, Shen B, Starnes LM, de Ruiter JR, Daniel JA, Konstantinopoulos PA, Cortez D, Cantor SB, Fernandez-Capetillo O, Ge K, Jonkers J, Rottenberg S, Sharan SK, Nussenzweig A, . Replication fork stability confers chemoresistance in BRCA-deficient cells. Nature, 2016,535(7612):382-387. |
| [34] | Du M, Su XM, Zhang T, Xing YJ . Aberrant promoter DNA methylation inhibits bone morphogenetic protein 2 expression and contributes to drug resistance in breast cancer. Mol Med Rep, 2014,10(2):1051-1055. |
| [35] | Kulis M, Heath S, Bibikova M, Queirós AC, Navarro A, Clot G, Martínez-Trillos A, Castellano G, Brun-Heath I, Pinyol M, Barberán-Soler S, Papasaikas P, Jares P, Beà S, Rico D, Ecker S, Rubio M, Royo R, Ho V, Klotzle B, Hernández L, Conde L, López-Guerra M, Colomer D, Villamor N, Aymerich M, Rozman M, Bayes M, Gut M, Gelpí JL, Orozco M, Fan JB, Quesada V, Puente XS, Pisano DG, Valencia A, López-Guillermo A, Gut I, López-Otín C, Campo E, Martín-Subero JI . Epigenomic analysis detects widespread gene-body DNA hypomethylation in chronic lymphocytic leukemia. Nat Genet, 2012,44(11):1236-1242. |
| [36] | Maunakea AK, Nagarajan RP, Bilenky M, Ballinger TJ, D'Souza C, Fouse SD, Johnson BE, Hong C, Nielsen C, Zhao Y, Turecki G, Delaney A, Varhol R, Thiessen N, Shchors K, Heine VM, Rowitch DH, Xing X, Fiore C, Schillebeeckx M, Jones SJ, Haussler D, Marra MA, Hirst M, Wang T, Costello JF,. Conserved role of intragenic DNA methylation in regulating alternative promoters. Nature, 2010,466(7303):253-257. |
| [37] | Varley KE, Gertz J, Bowling KM, Parker SL, Reddy TE, Pauli-Behn F, Cross MK, Williams BA, Stamatoyannopoulos JA, Crawford GE, Absher DM, Wold BJ, Myers RM . Dynamic DNA methylation across diverse human cell lines and tissues. Genome Res, 2013,23(3):555-567. |
| [38] | Sridhar VV, Kapoor A, Zhang K, Zhu J, Zhou T, Hasegawa PM, Bressan RA, Zhu JK . Control of DNA methylation and heterochromatic silencing by histone H2B deubiquitination. Nature. 2007,447(7145):735-738. |
| [39] | Schuettengruber B, Cavalli G . Recruitment of polycomb group complexes and their role in the dynamic regulation of cell fate choice. Development, 2009,136(21):3531-3542. |
| [40] | Hawkins RD, Hon GC, Lee LK, Ngo Q, Lister R, Pelizzola M, Edsall LE, Kuan S, Luu Y, Klugman S, Antosiewicz-Bourget J, Ye Z, Espinoza C, Agarwahl S, Shen L, Ruotti V, Wang W, Stewart R, Thomson JA, Ecker JR, Ren B . Distinct epigenomic landscapes of pluripotent and lineage-committed human cells. Cell Stem Cell, 2010,6(5):479-491. |
| [41] | Wagner EJ, Carpenter PB . Understanding the language of Lys36 methylation at histone H3. Nat Rev Mol Cell Biol, 2012,13(2):115-126. |
| [42] | Venkatesh S, Smolle M, Li H, Gogol MM, Saint M, Kumar S, Natarajan K, Workman JL . Set2 methylation of histone H3 lysine 36 suppresses histone exchange on transcribed genes. Nature, 2012,489(7416):452-455. |
| [43] | Chen BF, Chan WY . The de novo DNA methyltransferase DNMT3A in development and cancer. Epigenetics, 2014,9(5):669-677. |
| [44] | Deng DJ . DNA methylation and demethylation: current status and future per-spective. Hereditas(Beijing), 2014,36(5):403-410. |
| [44] | 邓大君 . DNA甲基化和去甲基化的研究现状及思考. 遗传. 2014,36(5):403-410. |
/
| 〈 |
|
〉 |
www.chinagene.cn
备案号：京ICP备09063187号-4
总访问:,今日访问:,当前在线:
