title: Role of miR-19b and its target mRNAs in 5-fluorouracil resistance in colon cancer cells.
abstract: BACKGROUND
  Drug resistance in colorectal cancers is assumed to be mediated by changes in the expression of microRNAs, but the specific identities and roles of microRNAs are largely unclear. We examined the effect of 5-fluorouracil (5-FU) resistance on microRNA expression.
  
  
  METHODS
  Two types of 5-FU-resistant colon cancer cells were derived from the DLD-1 and KM12C cell lines. The expressions of microRNAs were profiled with a microarray containing 723 microRNAs and validated by quantitative real-time polymerase chain reaction (qRT-PCR). To survey the downstream mediators of microRNA, we used a microRNA:mRNA immunoprecipitation (RIP)-Chip and pathway analysis tool to identify potential direct targets of microRNA.
  
  
  RESULTS
  In response to 5-FU, miR-19b and miR-21 were over-expressed in 5-FU-resistant cells. Of note, miR-19b was up-regulated 3.47-fold in the DLD-1 resistant cells, which exhibited no alteration in cell cycle profiles despite exposure to 5-FU. After transfection of miR-19b, specific mRNAs were recruited to microRNA:mRNA complexes isolated with Ago2 antibody and subjected to whole-genome transcriptional analysis. In this analysis, 66 target mRNAs were enriched by at least 5.0-fold in the microRNA:mRNA complexes from DLD-1 resistant cells. Ingenuity pathway analysis of mRNA targets significantly (P < 0.05) indicated the category "Cell Cycle" as a probable area of the molecular and cellular function related with 5-FU resistance. Among candidate mRNA targets, SFPQ and MYBL2 have been linked to cell cycle functions.
  
  
  CONCLUSIONS
  We revealed up-regulation of miR-19b in response to 5-FU and potential targets of miR-19b mediating the cell cycle under treatment with 5-FU. Our study provides an important insight into the mechanism of 5-FU resistance in colorectal cancers.
authors.affiliation: Department of Stress Science, Institute of Health Biosciences, The University of Tokushima Graduate School, Tokushima, 770-8503, Japan.
authors.forename: Ken
authors.initials: K
authors.lastname: Kurokawa
authors.forename: Toshihito
authors.initials: T
authors.lastname: Tanahashi
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Iima
authors.forename: Yuta
authors.initials: Y
authors.lastname: Yamamoto
authors.forename: Yoko
authors.initials: Y
authors.lastname: Akaike
authors.forename: Kensei
authors.initials: K
authors.lastname: Nishida
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Masuda
authors.forename: Yuki
authors.initials: Y
authors.lastname: Kuwano
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Murakami
authors.forename: Masakazu
authors.initials: M
authors.lastname: Fukushima
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Rokutan
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000627:therapeutic use*; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000188:drug therapy* / Q000235:genetics / Q000378:metabolism; D019008:Drug Resistance, Neoplasm; D005472:Fluorouracil / Q000627:therapeutic use*; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D035683:MicroRNAs / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D046228:Microarray Analysis; D012333:RNA, Messenger / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D014162:Transfection; D015854:Up-Regulation
chemical_list: D000964:Antimetabolites, Antineoplastic; C549214:MIRN19A microRNA, human; C524940:MIRN21 microRNA, human; D035683:MicroRNAs; D012333:RNA, Messenger; D005472:Fluorouracil