Transgenic mice for in vivo epigenome editing with CRISPR-based systems

Area of Science:

• Molecular Biology
• Genetics
• Epigenetics

Background:

• CRISPR-Cas9 enables precise DNA targeting.
• Epigenome editing uses deactivated Cas9 (dCas9) fused to chromatin modifiers.
• Delivery of large dCas9 fusion proteins hinders in vivo applications.

Purpose of the Study:

• To generate and characterize conditional transgenic mouse lines for epigenome editing.
• To enable facile, temporally controlled, and tissue-restricted epigenome editing in vivo.

Main Methods:

• Generation of Rosa26:LSL-dCas9-p300 (activation) and Rosa26:LSL-dCas9-KRAB (repression) mouse lines.
• Delivery of guide RNAs to target specific genomic loci (promoters, enhancers).
• Analysis of gene expression, epigenetic states, and phenotypic changes in various tissues and cell types.

Main Results:

• Demonstrated successful gene activation and repression in vivo (brain, liver) and ex vivo (T cells, fibroblasts).
• Showcased regulation of target genes and associated epigenetic modifications.
• Confirmed downstream phenotypic alterations in response to epigenome editing.

Conclusions:

• The developed mouse lines provide a convenient platform for in vivo epigenome editing.
• These tools facilitate temporal and tissue-specific control of gene expression.
• They are valuable for studying gene function and epigenetic regulation in complex biological systems.