title: Long noncoding RNA TUG1 is downregulated in non-small cell lung cancer and can regulate CELF1 on binding to PRC2.
abstract: Long noncoding RNAs (lncRNAs) play crucial roles in tumorigenesis, and lncRNA taurine-upregulated gene 1 (TUG1) has been proven to be associated with several human cancers. However, the mechanisms of TUG1-involved regulation remain largely unknown.
  
  
  
  We examined the expressions of TUG1 in a cohort of 89 patients with non-small cell lung cancer (NSCLC) to determine the association between TUG1 expression and clinical parameters. We used circular chromosome conformation capture (4C) coupled with next-generation sequencing to explore the genome regions that interact with TUG1 and the TUG1-mediated regulation.
  
  
  
  TUG1 was significantly downregulated, and the TUG1 downregulation correlated with sex (p = 0.006), smoking status (p = 0.016), and tumor differentiation grade (p = 0.001). Knockdown of TUG1 significantly promoted the proliferation of NSCLC cells. According to the bioinformatic analysis result of TUG1 4C sequencing data, 83 candidate genes and their interaction regions were identified. Among these candidate genes, CUGBP and Elav-like family member 1 (CELF1) are potential targets of TUG1 in-trans regulation. To confirm the interaction between TUG1 and CELF1, relative expressions of CELF1 were examined in TUG1 knockdown H520 cells; results showed that CELF1 was significantly upregulated in TUG1 knockdown H520 cells. RNA immunoprecipitation was then performed to examine whether TUG1 RNA was bound to PRC2, a TUG1-involved regulation mechanism reported in previous studies. The results demonstrated that TUG1 RNA was bound to enhancer of zeste protein 2/embryonic ectoderm development (EZH2/EED), which is essential for PRC2. Finally, our designed ChIP assay revealed that the EZH2/EED was bound to the promotor region of CELF1 within 992 bp upstream of the transcript start site.
  
  
  
  TUG1 is downregulated in NSCLC. Using TUG1 4C sequencing and bioinformatic analysis, we found CELF1 to be a potential target of TUG1 RNA in in-trans regulation. Moreover, subsequent experiments showed that TUG1 RNA could bind to PRC2 in the promotor region of CELF1 and negatively regulate CELF1 expressions in H520 cells. Our results may facilitate developing new treatment modalities targeting TUG1/PRC2/CELF1 interactions in patients with NSCLC.
authors.affiliation: Graduate Institute of Clinical Medicine, Kaohsiung Medical University, No. 100, Shih-Chuan 1st Road, Kaohsiung, Taiwan.|Department of Pediatrics, School of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.|Division of Hematology and Oncology, Department of Pediatrics, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan.
authors.forename: Pei-Chin
authors.initials: PC
authors.lastname: Lin
authors.affiliation: Department of Biological Science and Technology, Institute of Bioinformatics and Systems Biology, National Chiao Tung University, Hsinchu, Taiwan.
authors.forename: Hsien-Da
authors.initials: HD
authors.lastname: Huang
authors.affiliation: Graduate Institute of Clinical Medicine, Kaohsiung Medical University, No. 100, Shih-Chuan 1st Road, Kaohsiung, Taiwan.|Division of Chest Medicine, Department of Internal Medicine, Changhua Christian Hospital, Changhua, Taiwan.
authors.forename: Chun-Chi
authors.initials: CC
authors.lastname: Chang
authors.affiliation: Epigenome Research Center, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung, Taiwan.
authors.forename: Ya-Sian
authors.initials: YS
authors.lastname: Chang
authors.affiliation: Epigenome Research Center, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung, Taiwan.
authors.forename: Ju-Chen
authors.initials: JC
authors.lastname: Yen
authors.affiliation: Epigenome Research Center, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung, Taiwan.
authors.forename: Chien-Chih
authors.initials: CC
authors.lastname: Lee
authors.affiliation: Division of Hematology and Oncology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan.
authors.forename: Wen-Hsin
authors.initials: WH
authors.lastname: Chang
authors.affiliation: Graduate Institute of Clinical Medicine, Kaohsiung Medical University, No. 100, Shih-Chuan 1st Road, Kaohsiung, Taiwan. d730093@kmu.edu.tw.|Division of Hematology and Oncology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. d730093@kmu.edu.tw.|Department of Laboratory Medicine, Kaohsiung Medical University Hospital, Kaohsiung Medical University, Kaohsiung, Taiwan. d730093@kmu.edu.tw.
authors.forename: Ta-Chih
authors.initials: TC
authors.lastname: Liu
authors.affiliation: Epigenome Research Center, China Medical University Hospital, No. 2, Yuh-Der Road, Taichung, Taiwan. d6781@mail.cmuh.org.tw.|Department of Laboratory Medicine, China Medical University Hospital, Taichung, Taiwan. d6781@mail.cmuh.org.tw.|School of Medicine, China Medical University, Taichung, Taiwan. d6781@mail.cmuh.org.tw.
authors.forename: Jan-Gowth
authors.initials: JG
authors.lastname: Chang
mesh_terms: D000067879:CELF1 Protein / Q000235:genetics* / Q000378:metabolism; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics* / Q000473:pathology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D047369:Chromatin Immunoprecipitation; D015536:Down-Regulation; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics* / Q000473:pathology; D008297:Male; D063151:Polycomb Repressive Complex 2 / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D062085:RNA, Long Noncoding / Q000235:genetics* / Q000378:metabolism; D016019:Survival Analysis; D015533:Transcriptional Activation
chemical_list: D000067879:CELF1 Protein; C102843:CELF1 protein, human; D062085:RNA, Long Noncoding; C000598536:TUG1 long noncoding RNA, human; D063151:Polycomb Repressive Complex 2
keywords: CUGBP and Elav-like family member 1 (CELF1); Circular chromosome conformation capture (4C); Long noncoding RNA (lncRNA); Non-small cell lung cancer (NSCLC); Taurine-upregulated gene 1 (TUG1)