Screening for possible miRNA-mRNA associations in a colon cancer cell line.
MicroRNAs (miRNAs) are small non-coding RNAs mediating the regulation of gene expression in various biological contexts, including carcinogenesis. Here, we screened putative associations between 34, 45, and 103 miRNAs and 164, 391, and 81 mRNAs via Argonaute1 (Ago1) or Ago2 immunoprecipitation (IP) experiments in a colon cancer cell line. We used a combination of RIP Seq analysis. RNAs that were co-immunoprecipitated with Ago1 or Ago2 were used for massively parallel small RNA and mRNA sequencing. The detected miRNAs and mRNAs were further associated with one another based on in silico target predictions. Analysis of the putative associations indicated that, although Ago1 and Ago2 shared a similar repertory of miRNAs, the mRNAs possibly regulated by those miRNAs seemed different. The mRNAs detected with Ago1 IP were indicated to be frequently associated with genes having constitutive cellular functions, regulated by a smaller number of miRNAs, and appeared to receive more stringent translational regulation. In contrast, putative miRNA-mRNA associations detected with Ago2 IP appeared to be related to signal transduction genes, which had a larger number of possible miRNA binding sites. We then conducted a similar analysis using the colon cancer cells cultured under hypoxia and identified potential hypoxia-induced miRNA-mRNA associations, which included several well-characterized cancer-related genes as novel putative miRNA targets.
Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa, Chiba 277-8562, Japan.
Sotaro
S
Kanematsu
Kousuke
K
Tanimoto
Yutaka
Y
Suzuki
Sumio
S
Sugano
D060565:Argonaute Proteins / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D003110:Colonic Neoplasms / Q000235:genetics*; D004843:Epistasis, Genetic; D039642:Eukaryotic Initiation Factors / Q000378:metabolism; D057166:High-Throughput Screening Assays; D006801:Humans; D000860:Hypoxia / Q000235:genetics / Q000378:metabolism; D047468:Immunoprecipitation; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*
C115848:AGO1 protein, human; C466422:AGO2 protein, human; D060565:Argonaute Proteins; D039642:Eukaryotic Initiation Factors; D035683:MicroRNAs; D012333:RNA, Messenger
ATRX; Ago; Argonaute; B-cell CLL/lymphoma 2; BCL2; BCL2-interacting mediator of cell death; BIM; CAMKK2; Cancer; DACT1; HIC2; HIF1A; HMGA2; Hypoxia; MicroRNA; Next generation sequencing; RNA immunoprecipitation assay; RNA-induced silencing complex; RPLP2; TSC; VEGF; alpha thalassemia/mental retardation syndrome X-linked; calcium/calmodulin-dependent protein kinase kinase 2; dapper, antagonist of beta-catenin homolog 1; high mobility group AT-hook 2; hypermethylated in cancer 2; hypoxia inducible factor 1 alpha subunit; mTOR; mechanistic target of rapamycin; miRNA-silencing effector protein complexes; miRNPs; ribosomal protein large P2; tuberous sclerosis; vascular endothelial growth factor