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Move over Cas9: Introducing Cascade-Cas3 system

It has been over two weeks since Professors Emmanuelle Charpentier and Jennifer A. Doudna received The Nobel Prize in Chemistry 2020 for their work on the technology of genome editing. Now, researchers from UC San Francisco have reported of a new gene editing tool – Cascade-Cas3 – for the editing of larger pieces of DNA.

CRISPR-Cas

CRISPR-Cas systems are a diverse group of RNA-guided nucleases. They are derived from prokaryotes that use these systems against viral invaders. Most gene-editing applications focus on single subunit Class 2 CRISPR systems, for example, Cas9. However, Class 1 systems hold great potential as editing technologies. Cas3 is a signature gene in Class 1 Type I systems. It is a 3′–5′ single-strand DNA helicase-nuclease enzyme that, unlike Cas9, degrades target DNA processively. This property of Cascade (CRISPR-associated complex for antiviral defence)–Cas3 systems also provides the possibility of developing a tool for larger genomic deletions.

Joseph Bondy-Denomy, principal investigator, stated:

“Cas3 is like Cas9 with a motor – after finding its specific DNA target, it runs on DNA and chews it up like a Pac-Man.”

Cascade-Cas3

In a study, published in Nature Methods, researchers optimised a Type I-C CRISPR system from Pseudomonas aeruginosa (PaeCas3c). Specifically, Type I-C is streamlined and only requires four proteins.

The team found that DNA cleavage guided by a single CRISPR RNA generated large deletions (7-424 kilobases) in P. aeruginosa with near-100% efficiency. Cas3 is also able to generate bidirectional deletions originating from the programmed site.

Bond-Denomy added:

“Previously, there was no easy and reliable way to delete very large regions of DNA in bacteria for research or therapeutic purposes.

Now, instead of making 100 different small DNA deletions, we can just make one deletion and ask, ‘What changed?'”

This system will allow researchers to delete or replace longer stretches of DNA. In turn, this will enable them to efficiently assess the importance of genomic regions. Additionally, it will facilitate the manipulation of repetitive and noncoding regions. The team believe that the intrinsic properties of Cas3 make it a promising tool to fill in the gaps of current gene-editing capabilities.

Image credit: wildpixel – canva.com


More on these topics

CRISPR / Gene Editing / Nobel Prize