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A Reporter Based Cellular Assay for Monitoring Splicing Efficiency
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Modulation of Splicing by Single-Stranded Silencing RNAs.

Jing Liu1,2, Jiaxin Hu1,2, Jessica A Hicks1,2

  • 11Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, Dallas, Texas.

Nucleic Acid Therapeutics
|March 11, 2015
PubMed
Summary

Single-stranded silencing RNAs (ss-siRNAs) can alter gene splicing, including dystrophin. This RNA interference (RNAi) mechanism offers a new therapeutic strategy for increasing beneficial protein production.

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Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Single-stranded silencing RNAs (ss-siRNAs) are novel chemically modified oligonucleotides.
  • Their mechanism of action via RNA interference (RNAi) machinery for gene expression modulation is under investigation.
  • The potential of ss-siRNAs as a gene expression control platform is largely unexplored.

Purpose of the Study:

  • To investigate if ss-siRNAs targeting splice sites can increase the production of therapeutically relevant protein isoforms.
  • To test the hypothesis that ss-siRNAs can modulate gene splicing for therapeutic benefit.
  • To explore the role of RNAi factors in ss-siRNA-mediated splicing modulation.

Main Methods:

  • Utilizing ss-siRNAs designed to target specific splice sites.
  • Assessing changes in dystrophin splicing patterns.
  • Evaluating the requirement for seed sequence complementarity and argonaute 2 expression.

Main Results:

  • ss-siRNAs were observed to alter dystrophin splicing.
  • Altered splicing was dependent on seed sequence complementarity to the target.
  • The effect was contingent upon the expression of the RNAi factor argonaute 2.
  • Evidence suggests RNAi can occur within the mammalian cell nucleus.

Conclusions:

  • ss-siRNAs can effectively modulate gene splicing.
  • This provides a novel therapeutic development avenue for enhancing protein isoform production.
  • The findings support the occurrence of RNAi in mammalian cell nuclei, impacting classical nuclear processes like splicing.