CRISPR artificial splicing factors
The engineering of CRISPR Artificial Splicing Factors (CASFx) based on RNA-targetingCRISPR-Cas systems are reported and it is shown that simultaneous exon inclusion and exclusion can be induced at distinct targets by differential positioning of CASFx.
Abstract
<jats:title>Abstract</jats:title> <jats:p> Alternative splicing allows expression of mRNA isoforms from a single gene, expanding the diversity of the proteome. Its prevalence in normal biological and disease processes warrant precise tools for modulation. Here we report the engineering of CRISPR Artificial Splicing Factors (CASFx) based on RNA-targeting CRISPR-Cas systems. We show that simultaneous exon inclusion and exclusion can be induced at distinct targets by differential positioning of CASFx. We also create inducible CASFx (iCASFx) using the FKBP-FRB chemical-inducible dimerization domain, allowing small molecule control of alternative splicing. Finally, we demonstrate the activation of <jats:italic>SMN2</jats:italic> exon 7 splicing in spinal muscular atrophy (SMA) patient fibroblasts, suggesting a potential application of the CASFx system. </jats:p>