CRISPR Interference-Based Specific and Efficient Gene Inactivation in the Brain

Nat Neurosci. 2018 Mar;21(3):447-454. doi: 10.1038/s41593-018-0077-5. Epub 2018 Feb 5.

Abstract

CRISPR-Cas9 has been demonstrated to delete genes in postmitotic neurons. Compared to the establishment of proliferative cell lines or animal strains, it is more challenging to acquire a highly homogeneous consequence of gene editing in a stable neural network. Here we show that dCas9-based CRISPR interference (CRISPRi) can efficiently silence genes in neurons. Using a pseudotarget fishing strategy, we demonstrate that CRISPRi shows superior targeting specificity without detectable off-target activity. Furthermore, CRISPRi can achieve multiplex inactivation of genes fundamental for neurotransmitter release with high efficiency. By developing conditional CRISPRi tools targeting synaptotagmin I (Syt1), we modified the excitatory to inhibitory balance in the dentate gyrus of the mouse hippocampus and found that the dentate gyrus has distinct regulatory roles in learning and affective processes in mice. We therefore recommend CRISPRi as a useful tool for more rapid investigation of gene function in the mammalian brain.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Affect / physiology
  • Animals
  • Brain Chemistry / genetics*
  • CRISPR-Cas Systems / genetics*
  • Cell Proliferation
  • Cognition / physiology
  • Dentate Gyrus / metabolism
  • Fear / psychology
  • Gene Silencing
  • Hindlimb Suspension / psychology
  • Learning / physiology
  • Male
  • Maze Learning
  • Memory / physiology
  • Mice
  • Mice, Inbred C57BL
  • RNA Interference
  • Synaptotagmin I / genetics

Substances

  • Synaptotagmin I
  • Syt1 protein, mouse