TargetLink, a new method for identifying the endogenous target set of a specific microRNA in intact living cells

Yan Xu, Yan Chen, Daliang Li, Qing Liu, Zhenyu Xuan, Wen Hong Li

Research output: Contribution to journalArticle

Abstract

MicroRNAs are small non-coding RNAs acting as posttranscriptional repressors of gene expression. Identifying mRNA targets of a given miRNA remains an outstanding challenge in the field. We have developed a new experimental approach, TargetLink, that applied locked nucleic acid (LNA) as the affinity probe to enrich target genes of a specific microRNA in intact cells. TargetLink also consists a rigorous and systematic data analysis pipeline to identify target genes by comparing LNA-enriched sequences between experimental and control samples. Using miR-21 as a test microRNA, we identified 12 target genes of miR-21 in a human colorectal cancer cell by this approach. The majority of the identified targets interacted with miR-21 via imperfect seed pairing. Target validation confirmed that miR-21 repressed the expression of the identified targets. The cellular abundance of the identified miR-21 target transcripts varied over a wide range, with some targets expressed at a rather low level, confirming that both abundant and rare transcripts are susceptible to regulation by microRNAs, and that TargetLink is an efficient approach for identifying the target set of a specific microRNA in intact cells. C20orf111, one of the novel targets identified by TargetLink, was found to reside in the nuclear speckle and to be reliably repressed by miR-21 through the interaction at its coding sequence.

Original languageEnglish (US)
Pages (from-to)259-274
Number of pages16
JournalRNA Biology
Volume14
Issue number2
DOIs
StatePublished - Feb 1 2017

Keywords

  • Colorectal cancer
  • LNA
  • locked nucleic acid
  • microRNA
  • microRNA target
  • miR-21
  • target identification
  • targetlink

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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