Cell: Nobel laureate's latest Cell paper: A large number of viruses exist in the CRISPR system, from which smaller and more efficient Cas enzymes are mined

On June 28, 2012, Jennifer Doudna, Emmanuelle Charpentier et al.
published an epoch-making paper in the journal Science ^[1].

The detailed mechanism of action of the CRISPR-Cas system is revealed and its potential
as a genome editing tool is noted.
Since then, under the promotion of Zhang Feng, George Church, Qi Lei, Liu Ruqian and others, CRISPR gene editing technology has developed rapidly, becoming the most simple and efficient gene editing tool, and has made breakthroughs
in gene function research, drug target screening, genetic disease treatment, cancer research, crop breeding and other fields.

The CRISPR-Cas system is best known as a genome editing tool, but it's actually a class of immune systems
that are widespread in nature.
40% of bacteria and 85% of archaea have the CRISPR-Cas system, and these prokaryotes can capture fragments of the genome of invading viruses (bacteriophages) or plasmids and store them in CRISPR arrays in their own
genomes.
When these viruses invade again, the CRISPR array acts as a template to transcribe RNA, directing the Cas enzyme to cut the corresponding DNA of the invading virus and thus defend against the invading virus
.

Interestingly, viruses sometimes "steal" genome fragments from host cells, and if these "stolen" DNA give the virus a competitive advantage, they may be retained and gradually modified to better serve the virus
.
For example, a 2013 Nature paper showed that a bacteriophage infected with Vibrio cholerae has the CRISPR-Cas system and is used to escape the innate immune system of Vibrio cholerae ^[2].

In the "arms race" between bacteriophages and bacteria/archaea, there is a lack of systematic research
on how many phages have evolved their own CRISPR-Cas systems.

On November 23, 2022, Jennifer Doudna, Nobel Prize winner and CRISPR gene editing pioneer, and others published a research paper in the journal Cell entitled: Diverse virus-encoded CRISPR-Cas systems include streamlined genome editors ^[3].

The study is an important step
forward in discovering the enormous diversity of CRISPR-Cas systems.

The Jennifer Doudna Lab at the University of California, Berkeley, and the Jillian Banfield Lab used genomic resolution metagenomics to analyze the microbiome from nature, humans, and animals to comprehensively study the CRISPR-Cas system in bacteriophages, and they were surprised to find about 6,000 bacteriophages with the CRISPR-Cas system (0.
4% of known phages), covering all six known species CRSIPR-Cas system types (type I.
-VI.
, such as Cas9 belongs to type II.
, Cas12 belongs to type V.
, Cas13 belongs to type VI.
).

Importantly, some of these Casλ enzymes from bacteriophages (type V, just over 700 amino acids in size) are capable of editing plant and human cell genomes with the advantages
of miniaturization and high editing efficiency.

The study also found that the CRISPR-Cas systems in these phages have extensive variation in structure, with some systems missing components and some very compact
.
Even though bacteriophages carrying CRISPR-Cas systems are rare, the absolute number of phage CRSIPR-Cas systems will be considerable
, given the highly diverse and widespread distribution of bacteriophages.
It also reminds us that nature is always full of surprises
.

Viral (bacteriophage) genomes are much more compact than bacteria/archaea genomes, so some phages have small CRISPR-Cas systems, and miniaturized CRSIPR-Cas systems are exactly what
scientists dream of.
For gene editing, miniaturized systems can be better delivered via adeno-associated virus (AAV) vectors for in vivo gene editing
.

bacteriophage

In the study, Jennifer Doudna's lab focused on a newly discovered class of miniaturized Cas enzymes called Casλ, some of which they found could be used to edit the genomes
of Arabidopsis, wheat, and human cells.

Casλ versus Cas12a-edited human cell genomes

In recent years, many research teams have worked to mine novel gene editing tools from bacteria and archaea in nature, and have also discovered many miniaturized CAS enzymes, however, to date, these newly discovered miniaturized CAS enzymes are usually relatively inefficient
in practical gene editing applications.
In contrast, some of the Casλases from bacteriophages found in this study have the dual advantages
of miniaturization and high efficiency.
These findings suggest that viruses (bacteriophages) are an important source for the discovery of
novel gene-editing tools.

Structure of the Casλ-gRNA-DNA complex

In addition to in vitro bacteriophages, plasmids are also genetic elements of invading bacteria/archaea, so these plasmids transferred between microorganisms may also capture the host's CRISPR-Cas system, and they may also be the source of the discovery of novel CRISPR-Cas systems
.

Original source:

1) Seed K，Lazinski D，Calderwood S， et al.
A bacteriophage encodes its own CRISPR/Cas adaptive response to evade host innate immunity.
Nature 494， 489–491 (2013).
https：//doi.
org/10.
1038/nature11927.

2) MARTIN JINEK， KRZYSZTOF CHYLINSKI， INES FONFARA，et al.
A Programmable Dual-RNA–Guided DNA Endonuclease in Adaptive Bacterial Immunity.
SCIENCE， 28， Jun 2012， Vol 337， Issue 6096， pp.
816-821.

3) Basem Al-Shayeb， et al.
Diverse virus-encoded CRISPR-Cas systems include streamlined genome editors.
Cell， 2022.