Specific interference with gene expression and gene function mediated by long dsRNA in neural cells

J Neurosci Methods. 2002 Dec 15;121(2):151-7. doi: 10.1016/s0165-0270(02)00230-3.

Abstract

Double-stranded (ds) RNA-induced sequence-specific interference with gene expression, RNA interference (RNAi), has been extensively used in invertebrates, allowing for efficient and high-throughput gene silencing and gene function analysis. In vertebrates, however, use of RNAi to study gene function has been limited due to non-specific effects induced by double-stranded RNA (dsRNA)-dependent protein kinase and interferon activation. dsRNA-induced specific inhibition of vertebrate gene expression has only been shown in embryonic and non-differentiated mammalian cells. In this report, we demonstrate dsRNA-induced specific interference of gene expression and gene function in partially as well as fully differentiated mouse neuroblastoma cells. Specific silencing was observed in the expression of an integrated transgene coding for green fluorescent protein and a variety of endogenous genes. Moreover, we show that RNAi-mediated inhibition of poly (ADP-ribose) polymerase (PARP) expression induced cellular resistance to oxygen-glucose deprivation, consistent with the role of PARP in ischemia-induced brain damage. Our results indicate that RNAi can be used as a powerful tool to study gene function in neural cells.

Publication types

  • Comparative Study

MeSH terms

  • Animals
  • Cell Culture Techniques
  • Gene Silencing* / drug effects
  • Green Fluorescent Proteins
  • Luminescent Proteins / metabolism
  • Mice
  • Microtubule-Associated Proteins / drug effects
  • Microtubule-Associated Proteins / genetics
  • Microtubule-Associated Proteins / metabolism
  • Neuroblastoma
  • Neurons / drug effects*
  • Neurons / physiology
  • Poly(ADP-ribose) Polymerases / drug effects
  • Poly(ADP-ribose) Polymerases / genetics
  • Poly(ADP-ribose) Polymerases / metabolism
  • RNA Interference / physiology*
  • RNA, Double-Stranded / pharmacology*
  • RNA, Small Interfering / pharmacology
  • Time Factors
  • Tretinoin / pharmacology
  • Tumor Cells, Cultured

Substances

  • Luminescent Proteins
  • Microtubule-Associated Proteins
  • RNA, Double-Stranded
  • RNA, Small Interfering
  • Green Fluorescent Proteins
  • Tretinoin
  • Poly(ADP-ribose) Polymerases