Depletion of human micro-RNA miR-125b reveals that it is critical for the proliferation of differentiated cells but not for the down-regulation of putative targets during differentiation

J Biol Chem. 2005 Apr 29;280(17):16635-41. doi: 10.1074/jbc.M412247200. Epub 2005 Feb 18.


Micro-RNAs are small non-coding RNAs that regulate target gene expression post-transcriptionally through base pairing with the target messenger RNA. Functional characterization of micro-RNAs awaits robust experimental methods to knock-down a micro-RNA as well as to assay its function in vivo. In addition to the recently developed method to sequester micro-RNA with 2'-O-methyl antisense oligonucleotide, we report that small interfering RNA against the loop region of a micro-RNA precursor can be used to deplete the micro-RNA. The depletion of miR-125b by this method had a profound effect on the proliferation of adult differentiated cancer cells, and this proliferation defect was rescued by co-transfected mature micro-RNA. This technique has unique advantages over the 2'-O-methyl antisense oligonucleotide and can be used to determine micro-RNA function, assay micro-RNAs in vivo, and identify the contribution of a predicted micro-RNA precursor to the pool of mature micro-RNA in a given cell. miR-125b and let-7 micro-RNAs are induced, whereas their putative targets, lin-28 and lin-41, are decreased during in vitro differentiation of Tera-2 or embryonic stem cells. Experimental increase or decrease of micro-RNA concentrations did not, however, affect the levels of the targets, a finding that is explained by the fact that the down-regulation of the targets appears to be mostly at the transcriptional level in these in vitro differentiation systems. Collectively these results reveal the importance of micro-RNA depletion strategies for directly determining micro-RNA function in vivo.

Publication types

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

MeSH terms

  • Base Sequence
  • Blotting, Northern
  • Cell Differentiation
  • Cell Line, Tumor
  • Cell Nucleus / metabolism
  • Cell Proliferation
  • Down-Regulation*
  • Humans
  • MicroRNAs / chemistry*
  • Molecular Sequence Data
  • Oligonucleotides / chemistry
  • Open Reading Frames
  • RNA / chemistry
  • RNA Processing, Post-Transcriptional
  • RNA, Messenger / metabolism
  • RNA, Small Interfering / metabolism
  • Transcription, Genetic*
  • Transfection


  • MicroRNAs
  • Oligonucleotides
  • RNA, Messenger
  • RNA, Small Interfering
  • RNA