CRISPR-Cas9-based Photoactivatable Transcription System

Abstract

Targeted endogenous gene activation is necessary for understanding complex gene networks and has great potential in medical and industrial applications. The CRISPR-Cas system offers simple and powerful tools for this purpose. However, these CRISPR-Cas-based tools for activating user-defined genes are unable to offer precise temporal control of gene expression, despite the fact that many biological phenomena are regulated by highly dynamic patterns of gene expression. Here we created a light-inducible, user-defined, endogenous gene activation system based on CRISPR-Cas9. We demonstrated that this CRISPR-Cas9-based transcription system can allow rapid and reversible targeted gene activation by light. In addition, using this system, we have exemplified photoactivation of multiple user-defined endogenous genes in mammalian cells. The present CRISPR-Cas9-based transcription system offers simple and versatile approaches for precise endogenous gene activation in basic biological research and biotechnology applications.

dCas9: A Versatile Tool for Epigenome Editing

“…In order to control effector recruitment rather than split-(d)Cas9 reconstitution, the heterodimerization partners CRY2 and CIB1 have been used to bring together full-length dCas9 and effectors such as VP64 or p65 upon blue light irradiation (Konermann et al, 2013;Nihongaki et al, 2015b;Polstein and Gersbach, 2015) (Fig. 2.2C).…”

Genome-Editing Technologies: Principles and Applications

“…Much like zinc fingers and TALEs, methods for achieving conditional gene modulation using dCas9 have also been reported, including the fusion of a dihydrofolate reductase destabilization domain to dCas9, which can provide chemical control over activation, enabling cellular reprogramming or differentiation (Balboa et al 2015). Light-inducible dCas9-based systems capable of providing optical control of gene expression provide another means for achieving conditional control of gene expression (Nihongaki et al 2015;Polstein and Gersbach 2015).…”

CRISPR: gene editing is just the beginning

“…The resulting system triggers gene expression when cells are exposed to the light, and stops it when the light is flicked off 12 . A group led by chemical biologist Moritoshi Sato of the University of Tokyo rigged a similar system 13 , and also made an active Cas9 that edited the genome only after it was hit with blue light 14 .…”