Transcriptional Activation via CRISPR-dCas9 Mimics Overexpression Phenotypes in Arabidopsis
The CRISPR-Cas9 system allows effective gene modification through RNA-guided
DNA targeting. Researchers have significantly altered Cas9, resulting in
partially or completely deactivated Cas9. The deactivated Cas9 (dCas9)
offers a platform to regulate transcriptional expression with the addition
of an activator or repressor.
The team of Jong-Jin Park of the Oak Ridge National Laboratory redesigned a
CRISPR-Cas9 activation system by adding several activators (p65
transactivating subunit of NF-kappa B and heat-shock factor 1, or HSF
activator) to the dCas9 for application in plants. The redesigned
CRISPR-Cas9 activation system was tested in Arabidopsis to increase
endogenous transcriptional levels of production of anthocyanin pigment 1
(PAP1) and Arabidopsis thaliana vacuolar H+-pyrophosphatase (AVP1).
The expression of PAP1 was increased significantly and the activated plants
exhibited purple leaves similar to that of PAP1 overexpressing lines. The
AVP1 gene expression was also significantly increased in the CRISPR-edited
plants. Compared to wild types, the AVP1 activated plants had increased leaf
numbers, larger single-leaf areas and improved tolerance to drought stress,
similar phenotypes to AVP1 overexpressors.
Therefore, the redesigned CRISPR-Cas9 activation system containing modified
p65-HSF provides a simple approach for producing activated plants by
upregulating endogenous transcription levels.