Impact of targeting the adenine- and uracil-rich element of bcl-2 mRNA with oligoribonucleotides on apoptosis, cell cycle, and neuronal differentiation in SHSY-5Y cells

Mol Pharmacol. 2008 Feb;73(2):498-508. doi: 10.1124/mol.107.038323. Epub 2007 Nov 7.

Abstract

We have identified previously a destabilizing adenine- and uracil-rich element (ARE) in the 3'-UTR of bcl-2 mRNA that interacted with ARE-binding proteins to down-regulate bcl-2 gene expression in response to apoptotic stimuli. We have also described three contiguous 2'-O-methyl oligoribonucleotides (ORNs) in both sense and antisense orientation with respect to the bcl-2 ARE that are able to regulate the bcl-2 mRNA half-life and Bcl-2 protein level in two different cell lines. Here we show that treatment of neuronal cell line (SHSY-5Y) with antisense ORNs targeting the bcl-2 ARE (bcl-2 ARE asORNs) prevents bcl-2 down-regulation in response to apoptotic stimuli with glucose/growth factor starvation (Locke medium) or oxygen deprivation and enhances the apoptotic threshold as evaluated by time-lapse videomicroscopy, fluorescence-activated cell sorting analysis, and caspase-3 activation. Additional effects of bcl-2 ARE asORNs included inhibition of cell cycle entry and a marked increase of cellular neurite number and length, a hallmark of neuronal differentiation resulting from bcl-2 up-regulation. The ability of bcl-2 ARE asORNs to enhance the apoptotic threshold and to induce neuronal differentiation implies their potential application as a novel informational tool to protect cells from ischemic damage and to prevent neuronal degeneration.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenine / physiology*
  • Apoptosis / physiology*
  • Cell Cycle / physiology
  • Cell Differentiation / genetics*
  • Cell Line, Tumor
  • Gene Targeting / methods
  • Humans
  • Neurons / cytology*
  • Neurons / physiology
  • Oligoribonucleotides / genetics*
  • Oligoribonucleotides / metabolism
  • Proto-Oncogene Proteins c-bcl-2 / genetics*
  • Proto-Oncogene Proteins c-bcl-2 / metabolism
  • RNA, Messenger / biosynthesis
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism*
  • Uracil / metabolism*

Substances

  • Oligoribonucleotides
  • Proto-Oncogene Proteins c-bcl-2
  • RNA, Messenger
  • Uracil
  • Adenine