RIP-chip-SRM--a new combinatorial large-scale approach identifies a set of translationally regulated bantam/miR-58 targets in C. elegans

Genome Res. 2012 Jul;22(7):1360-71. doi: 10.1101/gr.133330.111. Epub 2012 Mar 27.

Abstract

MicroRNAs (miRNAs) are small, noncoding RNAs that negatively regulate gene expression. As miRNAs are involved in a wide range of biological processes and diseases, much effort has been invested in identifying their mRNA targets. Here, we present a novel combinatorial approach, RIP-chip-SRM (RNA-binding protein immunopurification + microarray + targeted protein quantification via selected reaction monitoring), to identify de novo high-confidence miRNA targets in the nematode Caenorhabditis elegans. We used differential RIP-chip analysis of miRNA-induced silencing complexes from wild-type and miRNA mutant animals, followed by quantitative targeted proteomics via selected reaction monitoring to identify and validate mRNA targets of the C. elegans bantam homolog miR-58. Comparison of total mRNA and protein abundance changes in mir-58 mutant and wild-type animals indicated that the direct bantam/miR-58 targets identified here are mainly regulated at the level of protein abundance, not mRNA stability.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Genetically Modified / genetics
  • Animals, Genetically Modified / metabolism
  • Caenorhabditis elegans / genetics*
  • Caenorhabditis elegans / metabolism
  • Crosses, Genetic
  • Immunologic Techniques / methods
  • MicroRNAs / genetics
  • MicroRNAs / metabolism*
  • Oligonucleotide Array Sequence Analysis / methods
  • Plasmids / genetics
  • Plasmids / metabolism
  • Protein Biosynthesis
  • Proteomics / methods*
  • RNA Interference
  • RNA Stability
  • RNA, Helminth / genetics
  • RNA, Helminth / metabolism
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism
  • Transgenes

Substances

  • MicroRNAs
  • RNA, Helminth
  • RNA, Messenger
  • RNA-Binding Proteins

Associated data

  • GEO/GSE32944