Ectopic expression of systemic RNA interference defective protein in embryonic stem cells.
RNA interference (RNAi), a post-transcriptional gene silencing mechanism originally described in Caenorhabditis elegans, involves sequence-specific mRNA degradation mediated by double-stranded RNAs (dsRNAs). Passive dsRNA uptake has been uniquely observed in C. elegans due to the expression of systemic RNA interference defective-1 (SID-1). Here we investigated the ability of ectopic SID-1 expression to enable passive cellular uptake of short interfering RNA (siRNA) or double stranded RNA (dsRNA) in pluripotent mouse embryonic stem cells (mESCs). When SID-1-GFP and the Firefly luciferase reporter gene (luc(Fir)) were co-expressed in mESCs, luc(Fir) activity could be suppressed by simple incubation with dsRNAs/siRNAs that were designed to specifically target luc(Fir). By contrast, suppression was not observed in mESCs expressing luc(Fir) and GFP alone or when either GFP- or SID-1-GFP-expressing cells were incubated with control dsRNAs/siRNAs (non-silencing or Renilla luciferase-specific). These results may lead to high-throughput experimental strategies for studying ESC differentiation and novel approaches to genetically inhibit or eliminate the tumorigenicity of ESCs.
Tsang
Suk Ying
SY
Stem Cell Program, University of California, Davis, CA 95817, USA.
Moore
Jennifer C
JC
Huizen
Rika Van
RV
Chan
Camie W Y
CW
Li
Ronald A
RA
D000818:Animals; D029742:Caenorhabditis elegans Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D053595:Embryonic Stem Cells / Q000378:metabolism*; D020868:Gene Silencing / Q000502:physiology*; D006801:Humans; D007668:Kidney / Q000196:embryology* / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D034622:RNA Interference / Q000502:physiology*; D011994:Recombinant Proteins / Q000378:metabolism; D014162:Transfection / Q000379:methods*
D029742:Caenorhabditis elegans Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins; C452203:SID-1 protein, C elegans