Off-target effect of doublecortin family shRNA on neuronal migration associated with endogenous MicroRNA dysregulation Journal Article


Author(s): Baek, SeungTae; Kerjan, Géraldine; Bielas, Stephanie L; Lee, Jieun; Fenstermaker, Ali G; Novarino, Gaia; Gleeson, Joseph G
Article Title: Off-target effect of doublecortin family shRNA on neuronal migration associated with endogenous MicroRNA dysregulation
Affiliation
Abstract: Acute gene inactivation using short hairpin RNA (shRNA, knockdown) in developing brain is a powerful technique to study genetic function; however, discrepancies between knockdown and knockout murine phenotypes have left unanswered questions. For example, doublecortin (Dcx) knockdown but not knockout shows a neocortical neuronal migration phenotype. Here we report that in utero electroporation of shRNA, but not siRNA or miRNA, to Dcx demonstrates a migration phenotype in Dcx knockouts akin to the effect in wild-type mice, suggestingshRNA-mediated off-target toxicity. This effect wasnot limited to Dcx, as it was observed in Dclk1 knockouts, as well as with a fraction of scrambled shRNAs, suggesting a sequence-dependent but not sequence-specific effect. Profiling RNAs from electroporated cells showed a defect in endogenous let7 miRNA levels, and disruption of let7 or Dicer recapitulated the migration defect. The results suggest that shRNA-mediated knockdown can produce untoward migration effects by altering endogenous miRNA pathways.
Keywords: unclassified drug; protein kinase; MicroRNA; dicer; doublecortin; doublecortin like kinase I; let7; short hairpin RNA; small interfering RNA
Journal Title: Neuron
Volume: 82
Issue 6
ISSN: 0896-6273
Publisher: Elsevier  
Date Published: 2014-06-18
Start Page: 1255
End Page: 1262
Sponsor: This work was supported by the National Institutes of Health R01NS41537. G.K. was supported by an EMBO Long Term Fellowship, S.L.B. by the A.P. Giannini Fellowship, and A.G.F. by the Brain Behavior Research Foundation
DOI: 10.1016/j.neuron.2014.04.036
Open access: no