title: Deregulated microRNAs in triple-negative breast cancer revealed by deep sequencing.
abstract: BACKGROUND
  MicroRNAs (miRNAs) are short, non-coding RNA molecules that play critical roles in human malignancy. However, the regulatory characteristics of miRNAs in triple-negative breast cancer, a phenotype of breast cancer that does not express the genes for estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2, are still poorly understood.
  
  
  METHODS
  In this study, miRNA expression profiles of 24 triple-negative breast cancers and 14 adjacent normal tissues were analyzed using deep sequencing technology. Expression levels of miRNA reads were normalized with the quantile-quantile scaling method. Deregulated miRNAs in triple-negative breast cancer were identified from the sequencing data using the Student's t-test. Quantitative reverse transcription PCR validations were carried out to examine miRNA expression levels. Potential target candidates of a miRNA were predicted using published target prediction algorithms. Luciferase reporter assay experiments were performed to verify a putative miRNA-target relationship. Validated molecular targets of the deregulated miRNAs were retrieved from curated databases and their associations with cancer progression were discussed.
  
  
  RESULTS
  A novel 25-miRNA expression signature was found to effectively distinguish triple-negative breast cancers from surrounding normal tissues in a hierarchical clustering analysis. We documented the evidence of seven polycistronic miRNA clusters preferentially harboring deregulated miRNAs in triple-negative breast cancer. Two of these miRNA clusters (miR-143-145 at 5q32 and miR-497-195 at 17p13.1) were markedly down-regulated in triple-negative breast cancer, while the other five miRNA clusters (miR-17-92 at 13q31.3, miR-183-182 at 7q32.2, miR-200-429 at 1p36.33, miR-301b-130b at 22q11.21, and miR-532-502 at Xp11.23) were up-regulated in triple-negative breast cancer. Moreover, miR-130b-5p from the miR-301b-130b cluster was shown to directly repress the cyclin G2 (CCNG2) gene, a crucial cell cycle regulator, in triple-negative breast cancer cells. Luciferase reporter assays showed that miR-130b-5p-mediated repression of CCNG2 was dependent on the sequence of the 3'-untranslated region. The findings described in this study implicate a miR-130b-5p-CCNG2 axis that may be involved in the malignant progression of triple-negative breast cancer.
  
  
  CONCLUSIONS
  Our work delivers a clear picture of the global miRNA regulatory characteristics in triple-negative breast cancer and extends the current knowledge of microRNA regulatory network.
authors.affiliation: Department of Electrical Engineering, Graduate Institute of Biomedical Electronics and Bioinformatics, National Taiwan University, Taipei, Taiwan. yinchang15@gmail.com.
authors.forename: Yao-Yin
authors.initials: YY
authors.lastname: Chang
authors.affiliation: Department of Surgery, College of Medicine, National Taiwan University, Taipei, Taiwan. npcancer@yahoo.com.tw.
authors.forename: Wen-Hung
authors.initials: WH
authors.lastname: Kuo
authors.affiliation: Department of Electrical Engineering, Graduate Institute of Biomedical Electronics and Bioinformatics, National Taiwan University, Taipei, Taiwan. paholisi.nctu@gmail.com.|Bioinformatics and Biostatistics Core, NTU Center of Genomic Medicine, Taipei, Taiwan. paholisi.nctu@gmail.com.
authors.forename: Jui-Hui
authors.initials: JH
authors.lastname: Hung
authors.affiliation: Department of Electrical Engineering, Graduate Institute of Biomedical Electronics and Bioinformatics, National Taiwan University, Taipei, Taiwan. kinomoto@sakura.idv.tw.|Bioinformatics and Biostatistics Core, NTU Center of Genomic Medicine, Taipei, Taiwan. kinomoto@sakura.idv.tw.
authors.forename: Chien-Yueh
authors.initials: CY
authors.lastname: Lee
authors.affiliation: Bioinformatics and Biostatistics Core, NTU Center of Genomic Medicine, Taipei, Taiwan. mr.beaneye@gmail.com.
authors.forename: Yung-Hua
authors.initials: YH
authors.lastname: Lee
authors.affiliation: Institute of Biotechnology, College of Bio-resources and Agriculture, National Taiwan University, Taipei, Taiwan. r99b22016@ntu.edu.tw.
authors.forename: Ya-Chu
authors.initials: YC
authors.lastname: Chang
authors.affiliation: Institute of Biotechnology, College of Bio-resources and Agriculture, National Taiwan University, Taipei, Taiwan. d00b48010@ntu.edu.tw.
authors.forename: Wen-Chun
authors.initials: WC
authors.lastname: Lin
authors.affiliation: Institute of Biotechnology, College of Bio-resources and Agriculture, National Taiwan University, Taipei, Taiwan. scying1123@gmail.com.
authors.forename: Cheng-Ying
authors.initials: CY
authors.lastname: Shen
authors.affiliation: Department of Surgery, College of Medicine, National Taiwan University, Taipei, Taiwan. huangcs@ntu.edu.tw.
authors.forename: Chiun-Sheng
authors.initials: CS
authors.lastname: Huang
authors.affiliation: Department of Obstetrics and Gynecology, College of Medicine, National Taiwan University, Taipei, Taiwan. fjhsieh@ntu.edu.tw.
authors.forename: Fon-Jou
authors.initials: FJ
authors.lastname: Hsieh
authors.affiliation: Department of Physiology, College of Medicine, National Taiwan University, Taipei, Taiwan. llai@ntu.edu.tw.
authors.forename: Liang-Chuan
authors.initials: LC
authors.lastname: Lai
authors.affiliation: Institute of Biotechnology, College of Bio-resources and Agriculture, National Taiwan University, Taipei, Taiwan. motiont@gmail.com.
authors.forename: Mong-Hsun
authors.initials: MH
authors.lastname: Tsai
authors.affiliation: Department of Surgery, College of Medicine, National Taiwan University, Taipei, Taiwan. kingjen@ntu.edu.tw.|Department of Surgery, Cheng Ching General Hospital, Taichung, Taiwan. kingjen@ntu.edu.tw.
authors.forename: King-Jen
authors.initials: KJ
authors.lastname: Chang
authors.affiliation: Department of Electrical Engineering, Graduate Institute of Biomedical Electronics and Bioinformatics, National Taiwan University, Taipei, Taiwan. chuangey@ntu.edu.tw.|Bioinformatics and Biostatistics Core, NTU Center of Genomic Medicine, Taipei, Taiwan. chuangey@ntu.edu.tw.
authors.forename: Eric Y
authors.initials: EY
authors.lastname: Chuang
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D018797:Cell Cycle Proteins / Q000235:genetics; D002460:Cell Line; D045744:Cell Line, Tumor; D056767:Cyclin G2 / Q000235:genetics; D015536:Down-Regulation / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics*; D057809:HEK293 Cells; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D008875:Middle Aged; D059467:Transcriptome / Q000235:genetics*; D064726:Triple Negative Breast Neoplasms / Q000235:genetics*; D015854:Up-Regulation / Q000235:genetics
chemical_list: D020413:3' Untranslated Regions; C101593:CCNG2 protein, human; D018797:Cell Cycle Proteins; D056767:Cyclin G2; D035683:MicroRNAs