Specific sequence determinants of miR-15/107 microRNA gene group targets

Nucleic Acids Res. 2011 Oct;39(18):8163-72. doi: 10.1093/nar/gkr532. Epub 2011 Jun 30.

Abstract

MicroRNAs (miRNAs) target mRNAs in human cells via complex mechanisms that are still incompletely understood. Using anti-Argonaute (anti-AGO) antibody co-immunoprecipitation, followed by microarray analyses and downstream bioinformatics, 'RIP-Chip' experiments enable direct analyses of miRNA targets. RIP-Chip studies (and parallel assessments of total input mRNA) were performed in cultured H4 cells after transfection with miRNAs corresponding to the miR-15/107 gene group (miR-103, miR-107, miR-16 and miR-195), and five control miRNAs. Three biological replicates were run for each condition with a total of 54 separate human Affymetrix Human Gene 1.0 ST array replicates. Computational analyses queried for determinants of miRNA:mRNA binding. The analyses support four major findings: (i) RIP-Chip studies correlated with total input mRNA profiling provides more comprehensive information than using either RIP-Chip or total mRNA profiling alone after miRNA transfections; (ii) new data confirm that miR-107 paralogs target coding sequence (CDS) of mRNA; (iii) biochemical and computational studies indicate that the 3' portion of miRNAs plays a role in guiding miR-103/7 to the CDS of targets; and (iv) there are major sequence-specific targeting differences between miRNAs in terms of CDS versus 3'-untranslated region targeting, and stable AGO association versus mRNA knockdown. Future studies should take this important miRNA-to-miRNA variability into account.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Argonaute Proteins / isolation & purification
  • Base Sequence
  • Cell Line
  • Down-Regulation*
  • Gene Expression Profiling
  • Humans
  • Immunoprecipitation
  • MicroRNAs / chemistry*
  • MicroRNAs / metabolism
  • Oligonucleotide Array Sequence Analysis
  • RNA, Messenger / chemistry
  • RNA, Messenger / metabolism*

Substances

  • Argonaute Proteins
  • MIRN107 microRNA, human
  • MIRN15 microRNA, human
  • MicroRNAs
  • RNA, Messenger