Molecules and Cells

  • 제44권9호
  • /
  • Pages.658-669
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  • 2021
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  • 1016-8478(pISSN)
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  • 0219-1032(eISSN)
한국분자세포생물학회 (Korean Society for Molecular and Cellular Biology)
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Identification and Functional Characterization of Two Noncoding RNAs Transcribed from Putative Active Enhancers in Hepatocellular Carcinoma

  • Lee, Ye-Eun (Division of Biomedical Convergence, College of Biomedical Science, Institute of Bioscience & Biotechnology, Kangwon National University) ;
  • Lee, Jiyeon (Severance Biomedical Science Institute, Graduate School of Medical Science, Brain Korea 21 Project, Gangnam Severance Hospital, Yonsei University College of Medicine) ;
  • Lee, Yong Sun (Department of Cancer Biomedical Science, Graduate School of Cancer Science and Policy, National Cancer Center) ;
  • Jang, Jiyoung Joan (Department of Cancer Biomedical Science, Graduate School of Cancer Science and Policy, National Cancer Center) ;
  • Woo, Hyeonju (Department of Life Science and the Research Center for Cellular Homeostasis, Ewha Womans University) ;
  • Choi, Hae In (Department of Bionanotechnology, Hanyang University) ;
  • Chai, Young Gyu (Department of Molecular and Life Science, Hanyang University) ;
  • Kim, Tae-Kyung (Department of Life Sciences, Pohang University of Science and Technology (POSTECH)) ;
  • Kim, TaeSoo (Department of Life Science and the Research Center for Cellular Homeostasis, Ewha Womans University) ;
  • Kim, Lark Kyun (Severance Biomedical Science Institute, Graduate School of Medical Science, Brain Korea 21 Project, Gangnam Severance Hospital, Yonsei University College of Medicine) ;
  • Choi, Sun Shim (Division of Biomedical Convergence, College of Biomedical Science, Institute of Bioscience & Biotechnology, Kangwon National University)
  • 투고 : 2021.07.07
  • 심사 : 2021.08.13
  • 발행 : 2021.09.30
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초록

Enhancers have been conventionally perceived as cis-acting elements that provide binding sites for trans-acting factors. However, recent studies have shown that enhancers are transcribed and that these transcripts, called enhancer RNAs (eRNAs), have a regulatory function. Here, we identified putative eRNAs by profiling and determining the overlap between noncoding RNA expression loci and eRNA-associated histone marks such as H3K27ac and H3K4me1 in hepatocellular carcinoma (HCC) cell lines. Of the 132 HCC-derived noncoding RNAs, 74 overlapped with the eRNA loci defined by the FANTOM consortium, and 65 were located in the proximal regions of genes differentially expressed between normal and tumor tissues in TCGA dataset. Interestingly, knockdown of two selected putative eRNAs, THUMPD3-AS1 and LINC01572, led to downregulation of their target mRNAs and to a reduction in the proliferation and migration of HCC cells. Additionally, the expression of these two noncoding RNAs and target mRNAs was elevated in tumor samples in the TCGA dataset, and high expression was associated with poor survival of patients. Collectively, our study suggests that noncoding RNAs such as THUMPD3-AS1 and LINC01572 (i.e., putative eRNAs) can promote the transcription of genes involved in cell proliferation and differentiation and that the dysregulation of these noncoding RNAs can cause cancers such as HCC.

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과제정보

This research was supported by National Research Foundation of Korea (NRF) (NRF-2019R1A5A6099645, NRF-2017M3A9G7073033) and by the Ministry of Education, Science, and Technology (MEST) (2019R1A2C 1002350).

참고문헌

  1. Alexander, J.J., Bey, E.M., Geddes, E.W., and Lecatsas, G. (1976). Establishment of a continuously growing cell line from primary carcinoma of the liver. S. Afr. Med. J. 50, 2124-2128.
  2. Ali, T. and Grote, P. (2020). Beyond the RNA-dependent function of LncRNA genes. Elife 9, e60583. https://doi.org/10.7554/eLife.60583
  3. Alvarez-Dominguez, J.R., Knoll, M., Gromatzky, A.A., and Lodish, H.F. (2017). The super-enhancer-derived alncRNA-EC7/Bloodlinc potentiates red blood cell development in trans. Cell Rep. 19, 2503-2514. https://doi.org/10.1016/j.celrep.2017.05.082
  4. Anders, S., Pyl, P.T., and Huber, W. (2015). HTSeq-a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166-169. https://doi.org/10.1093/bioinformatics/btu638
  5. Andersson, R., Gebhard, C., Miguel-Escalada, I., Hoof, I., Bornholdt, J., Boyd, M., Chen, Y., Zhao, X., Schmidl, C., Suzuki, T., et al. (2014). An atlas of active enhancers across human cell types and tissues. Nature 507, 455-461. https://doi.org/10.1038/nature12787
  6. Andrews, S. (2010). FastQC: a quality control tool for high throughput sequence data. Retrieved January 20, 2020, from https://www.bioinformatics.babraham.ac.uk/projects/fastqc/
  7. Arner, E., Daub, C.O., Vitting-Seerup, K., Andersson, R., Lilje, B., Drablos, F., Lennartsson, A., Ronnerblad, M., Hrydziuszko, O., Vitezic, M., et al. (2015). Transcribed enhancers lead waves of coordinated transcription in transitioning mammalian cells. Science 347, 1010-1014. https://doi.org/10.1126/science.1259418
  8. Banerji, J., Rusconi, S., and Schaffner, W. (1981). Expression of a β-globin gene is enhanced by remote SV40 DNA sequences. Cell 27(2 Pt 1), 299-308. https://doi.org/10.1016/0092-8674(81)90413-x
  9. Bhattarai, S., Akella, A., Gandhi, A., and Dharap, A. (2021). Modulation of brain pathology by enhancer RNAs in cerebral ischemia. Mol. Neurobiol. 58, 1482-1490. https://doi.org/10.1007/s12035-020-02194-9
  10. Bolger, A.M., Lohse, M., and Usadel, B. (2014). Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30, 2114-2120. https://doi.org/10.1093/bioinformatics/btu170
  11. Bulger, M. and Groudine, M. (2010). Enhancers: the abundance and function of regulatory sequences beyond promoters. Dev. Biol. 339, 250-257. https://doi.org/10.1016/j.ydbio.2009.11.035
  12. Carullo, N.V., Phillips Iii, R.A., Simon, R.C., Soto, S.A.R., Hinds, J.E., Salisbury, A.J., Revanna, J.S., Bunner, K.D., Ianov, L., Sultan, F.A., et al. (2020). Enhancer RNAs predict enhancer-gene regulatory links and are critical for enhancer function in neuronal systems. Nucleic Acids Res. 48, 9550-9570. https://doi.org/10.1093/nar/gkaa671
  13. Chen, H., Li, C., Peng, X., Zhou, Z., Weinstein, J.N., Cancer Genome Atlas Research Network, and Liang, H. (2018). A pan-cancer analysis of enhancer expression in nearly 9000 patient samples. Cell 173, 386-399.e12. https://doi.org/10.1016/j.cell.2018.03.027
  14. Chen, Y., Sheppard, D., Dong, X., Hu, X., Chen, M., Chen, R., Chakrabarti, J., Zavros, Y., Peek, R.M., and Chen, L. (2020). H. pylori infection confers resistance to apoptosis via Brd4-dependent BIRC3 eRNA synthesis. Cell Death Dis. 11, 667. https://doi.org/10.1038/s41419-020-02894-z
  15. Creyghton, M.P., Cheng, A.W., Welstead, G.G., Kooistra, T., Carey, B.W., Steine, E.J., Hanna, J., Lodato, M.A., Frampton, G.M., Sharp, P.A., et al. (2010). Histone H3K27ac separates active from poised enhancers and predicts developmental state. Proc. Natl. Acad. Sci. U. S. A. 107, 21931-21936. https://doi.org/10.1073/pnas.1016071107
  16. De Santa, F., Barozzi, I., Mietton, F., Ghisletti, S., Polletti, S., Tusi, B.K., Muller, H., Ragoussis, J., Wei, C., and Natoli, G. (2010). A large fraction of extragenic RNA pol II transcription sites overlap enhancers. PLoS Biol. 8, e1000384. https://doi.org/10.1371/journal.pbio.1000384
  17. Dobin, A., Davis, C.A., Schlesinger, F., Drenkow, J., Zaleski, C., Jha, S., Batut, P., Chaisson, M., and Gingeras, T.R. (2013). STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15-21. https://doi.org/10.1093/bioinformatics/bts635
  18. Ha, S., Cho, W., DeKoter, R.P., and Kim, S.O. (2019). The transcription factor PU. 1 mediates enhancer-promoter looping that is required for IL-1β eRNA and mRNA transcription in mouse melanoma and macrophage cell lines. J. Biol. Chem. 294, 17487-17500. https://doi.org/10.1074/jbc.RA119.010149
  19. Hah, N., Murakami, S., Nagari, A., Danko, C.G., and Kraus, W.L. (2013). Enhancer transcripts mark active estrogen receptor binding sites. Genome Res. 23, 1210-1223. https://doi.org/10.1101/gr.152306.112
  20. Hauberg, M.E., Fullard, J.F., Zhu, L., Cohain, A.T., Giambartolomei, C., Misir, R., Reach, S., Johnson, J.S., Wang, M., Mattheisen, M., et al. (2019). Differential activity of transcribed enhancers in the prefrontal cortex of 537 cases with schizophrenia and controls. Mol. Psychiatry 24, 1685-1695. https://doi.org/10.1038/s41380-018-0059-8
  21. Heintzman, N.D., Hon, G.C., Hawkins, R.D., Kheradpour, P., Stark, A., Harp, L.F., Ye, Z., Lee, L.K., Stuart, R.K., Ching, C.W., et al. (2009). Histone modifications at human enhancers reflect global cell-type-specific gene expression. Nature 459, 108-112. https://doi.org/10.1038/nature07829
  22. Heintzman, N.D., Stuart, R.K., Hon, G., Fu, Y., Ching, C.W., Hawkins, R.D., Barrera, L.O., Van Calcar, S., Qu, C., Ching, K.A., et al. (2007). Distinct and predictive chromatin signatures of transcriptional promoters and enhancers in the human genome. Nat. Genet. 39, 311-318. https://doi.org/10.1038/ng1966
  23. Heinz, S., Benner, C., Spann, N., Bertolino, E., Lin, Y.C., Laslo, P., Cheng, J.X., Murre, C., Singh, H., and Glass, C.K. (2010). Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol. Cell 38, 576-589. https://doi.org/10.1016/j.molcel.2010.05.004
  24. Hsieh, C.L., Fei, T., Chen, Y., Li, T., Gao, Y., Wang, X., Sun, T., Sweeney, C.J., Lee, G.S., Chen, S., et al. (2014). Enhancer RNAs participate in androgen receptor-driven looping that selectively enhances gene activation. Proc. Natl. Acad. Sci. U. S. A. 111, 7319-7324. https://doi.org/10.1073/pnas.1324151111
  25. Jiao, X., Sherman, B.T., Huang, D.W., Stephens, R., Baseler, M.W., Lane, H.C., and Lempicki, R.A. (2012). DAVID-WS: a stateful web service to facilitate gene/protein list analysis. Bioinformatics 28, 1805-1806. https://doi.org/10.1093/bioinformatics/bts251
  26. Kim, T., Hemberg, M., Gray, J.M., Costa, A.M., Bear, D.M., Wu, J., Harmin, D.A., Laptewicz, M., Barbara-Haley, K., Kuersten, S., et al. (2010). Widespread transcription at neuronal activity-regulated enhancers. Nature 465, 182-187. https://doi.org/10.1038/nature09033
  27. Ku, J.L. and Park, J.G. (2005). Biology of SNU cell lines. Cancer Res. Treat. 37, 1-19. https://doi.org/10.4143/crt.2005.37.1.1
  28. Lam, M.T., Cho, H., Lesch, H.P., Gosselin, D., Heinz, S., Tanaka-Oishi, Y., Benner, C., Kaikkonen, M.U., Kim, A.S., Kosaka, M., et al. (2013). Rev-Erbs repress macrophage gene expression by inhibiting enhancer-directed transcription. Nature 498, 511-515. https://doi.org/10.1038/nature12209
  29. Langmead, B., Trapnell, C., Pop, M., and Salzberg, S.L. (2009). Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol. 10, R25. https://doi.org/10.1186/gb-2009-10-3-r25
  30. Leek, J.T., Johnson, W.E., Parker, H.S., Jaffe, A.E., and Storey, J.D. (2012). The sva package for removing batch effects and other unwanted variation in high-throughput experiments. Bioinformatics 28, 882-883. https://doi.org/10.1093/bioinformatics/bts034
  31. Li, W., Notani, D., Ma, Q., Tanasa, B., Nunez, E., Chen, A.Y., Merkurjev, D., Zhang, J., Ohgi, K., Song, X., et al. (2013). Functional roles of enhancer RNAs for oestrogen-dependent transcriptional activation. Nature 498, 516-520. https://doi.org/10.1038/nature12210
  32. Li, X., Zhang, Q., Shi, Q., Liu, Y., Zhao, K., Shen, Q., Shi, Y., Liu, X., Wang, C., Li, N., et al. (2017). Demethylase Kdm6a epigenetically promotes IL-6 and IFN-β production in macrophages. J. Autoimmun. 80, 85-94. https://doi.org/10.1016/j.jaut.2017.02.007
  33. Love, M.I., Huber, W., and Anders, S. (2014). Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550. https://doi.org/10.1186/s13059-014-0550-8
  34. Maston, G.A., Evans, S.K., and Green, M.R. (2006). Transcriptional regulatory elements in the human genome. Annu. Rev. Genomics Hum. Genet. 7, 29-59. https://doi.org/10.1146/annurev.genom.7.080505.115623
  35. Melgar, M.F., Collins, F.S., and Sethupathy, P. (2011). Discovery of active enhancers through bidirectional expression of short transcripts. Genome Biol. 12, R113. https://doi.org/10.1186/gb-2011-12-11-r113
  36. Melo, C.A., Drost, J., Wijchers, P.J., van de Werken, H., de Wit, E., Oude Vrielink, J.A., Elkon, R., Melo, S.A., Leveille, N., Kalluri, R., et al. (2013). eRNAs are required for p53-dependent enhancer activity and gene transcription. Mol. Cell 49, 524-535. https://doi.org/10.1016/j.molcel.2012.11.021
  37. Moreau, P., Hen, R., Wasylyk, B., Everett, R., Gaub, M., and Chambon, P. (1981). The SV40 72 base repair repeat has a striking effect on gene expression both in SV40 and other chimeric recombinants. Nucleic Acids Res. 9, 6047-6068. https://doi.org/10.1093/nar/9.22.6047
  38. Mousavi, K., Zare, H., Dell'Orso, S., Grontved, L., Gutierrez-Cruz, G., Derfoul, A., Hager, G.L., and Sartorelli, V. (2013). eRNAs promote transcription by establishing chromatin accessibility at defined genomic loci. Mol. Cell 51, 606-617. https://doi.org/10.1016/j.molcel.2013.07.022
  39. Ong, C. and Corces, V.G. (2011). Enhancer function: new insights into the regulation of tissue-specific gene expression. Nat. Rev. Genet. 12, 283-293. https://doi.org/10.1038/nrg2957
  40. Orom, U.A., Derrien, T., Beringer, M., Gumireddy, K., Gardini, A., Bussotti, G., Lai, F., Zytnicki, M., Notredame, C., Huang, Q., et al. (2010). Long noncoding RNAs with enhancer-like function in human cells. Cell 143, 46-58. https://doi.org/10.1016/j.cell.2010.09.001
  41. Picard Toolkit (2019). Broad Institute, GitHub repository. Retrieved January 20, 2020, from http://broadinstitute.github.io/picard/
  42. Quinlan, A.R. and Hall, I.M. (2010). BEDTools: a flexible suite of utilities for comparing genomic features. Bioinformatics 26, 841-842. https://doi.org/10.1093/bioinformatics/btq033
  43. Riethoven, J.M. (2010). Regulatory regions in DNA: promoters, enhancers, silencers, and insulators. Methods Mol. Biol. 674, 33-42. https://doi.org/10.1007/978-1-60761-854-6_3
  44. Schaukowitch, K., Joo, J., Liu, X., Watts, J.K., Martinez, C., and Kim, T. (2014). Enhancer RNA facilitates NELF release from immediate early genes. Mol. Cell 56, 29-42. https://doi.org/10.1016/j.molcel.2014.08.023
  45. Sloan, C.A., Chan, E.T., Davidson, J.M., Malladi, V.S., Strattan, J.S., Hitz, B.C., Gabdank, I., Narayanan, A.K., Ho, M., Lee, B.T., et al. (2016). ENCODE data at the ENCODE portal. Nucleic Acids Res. 44(D1), D726-D732. https://doi.org/10.1093/nar/gkv1160
  46. Tang, Z., Li, C., Kang, B., Gao, G., Li, C., and Zhang, Z. (2017). GEPIA: a web server for cancer and normal gene expression profiling and interactive analyses. Nucleic Acids Res. 45(W1), W98-W102. https://doi.org/10.1093/nar/gkx247
  47. Trapnell, C., Pachter, L., and Salzberg, S.L. (2009). TopHat: discovering splice junctions with RNA-Seq. Bioinformatics 25, 1105-1111. https://doi.org/10.1093/bioinformatics/btp120
  48. Trapnell, C., Williams, B.A., Pertea, G., Mortazavi, A., Kwan, G., Van Baren, M.J., Salzberg, S.L., Wold, B.J., and Pachter, L. (2010). Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat. Biotechnol. 28, 511-515. https://doi.org/10.1038/nbt.1621
  49. Tsai, P., Dell'Orso, S., Rodriguez, J., Vivanco, K.O., Ko, K., Jiang, K., Juan, A.H., Sarshad, A.A., Vian, L., Tran, M., et al. (2018). A muscle-specific enhancer RNA mediates cohesin recruitment and regulates transcription in trans. Mol. Cell 71, 129-141.e8. https://doi.org/10.1016/j.molcel.2018.06.008
  50. Vucicevic, D., Corradin, O., Ntini, E., Scacheri, P.C., and Orom, U.A. (2015). Long ncRNA expression associates with tissue-specific enhancers. Cell Cycle 14, 253-260. https://doi.org/10.4161/15384101.2014.977641
  51. Wang, Y., Song, F., Zhang, B., Zhang, L., Xu, J., Kuang, D., Li, D., Choudhary, M.N., Li, Y., Hu, M., et al. (2018). The 3D Genome Browser: a web-based browser for visualizing 3D genome organization and long-range chromatin interactions. Genome Biol. 19, 151. https://doi.org/10.1186/s13059-018-1519-9
  52. Wu, H., Nord, A.S., Akiyama, J.A., Shoukry, M., Afzal, V., Rubin, E.M., Pennacchio, L.A., and Visel, A. (2014). Tissue-specific RNA expression marks distant-acting developmental enhancers. PLoS Genet. 10, e1004610. https://doi.org/10.1371/journal.pgen.1004610
  53. Yi, M. (2010). Hepatitis C virus: propagation, quantification, and storage. Curr. Protoc. Microbiol. 19, 15D.1.1-15D.1.11.
  54. Zhang, Y., Liu, T., Meyer, C.A., Eeckhoute, J., Johnson, D.S., Bernstein, B.E., Nusbaum, C., Myers, R.M., Brown, M., Li, W., et al. (2008). Model-based analysis of ChIP-Seq (MACS). Genome Biol. 9, R137. https://doi.org/10.1186/gb-2008-9-9-r137
  55. Zhang, Z., Lee, J., Ruan, H., Ye, Y., Krakowiak, J., Hu, Q., Xiang, Y., Gong, J., Zhou, B., Wang, L., et al. (2019). Transcriptional landscape and clinical utility of enhancer RNAs for eRNA-targeted therapy in cancer. Nat. Commun. 10, 4562. https://doi.org/10.1038/s41467-019-12543-5

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