Unmet Need:
CRISPR-Cas9 from Streptococcus pyogenes is the most commonly used due to Cas9’s ability to act as a singular protein and its high gene cutting activity (see Knott and Nakade). Despite the widespread use of CRISPR-Cas9, limitations remain in the efficiency with which precise genome editing occurs (see Banan). CRISPR-Cas9 creates a double-stranded break in the gene it is targeted to, which often leads to loss of the gene’s functional products (see Liu). While sometimes desirable, this loss can also be detrimental to the cell. Designing a CRISPR-Cas9 system that does not create double-stranded breaks could enable alteration of gene expression via the precise sequence targeting capabilities of CRISPR-Cas9 without the potentially unwanted loss of gene product. As such, there remains a need for development of a CRISPR-Cas9 method for altering gene expression without directly cutting the targeted gene.
Technology Overview:
Researchers at Johns Hopkins have discovered an additional functionality of CRISPR-Cas9 which allows the system to target a specific gene promotor and repress its transcription without cutting the gene. They found that a longer version of the tracrRNA (usually needed for activation of Cas9 targeting and gene cleavage) can enable Cas9 binding without cutting. This development can be used as either an alterative or complementary method to the traditionally used gene cutting capability of CRISPR-Cas9.
Stage of Development:
The discovered long tracrRNA has been shown to accurately target and repress gene transcription in bacteria. Experiments are currently underway to demonstrate use in mammalian cells as a genome editing technology.
Publication:
Workman R.E. et al. Cell. 2021. 184, 3.
