Millions of New Gene Editing Enzymes Discovered During Investigation CRISPR Origins

Millions of New Gene Editing Enzymes Discovered During Investigation CRISPR Origins

A new study exploring the origins of CRISPR and has shown we have only just scratched the surface and there are potentially millions of CRISPR-like systems that could potentially be harnessed and used in medicine as gene editing tools.

CRISPR, or clustered regularly interspaced short palindromic repeats to give it its full name, is a family of DNA sequences found in the genomes of certain bacteria and archaea, which are derived from bacteriophages that previously infected those organisms. They are used to detect and destroy bacteriophages the next time they are encountered. An associated enzyme, of which Cas9 is one example, is used in conjunction with an RNA to recognize a complementary section of DNA and remove it. The enzyme component acts like a pair of molecular scissors and protects against changes to DNA as a result of infection. Researchers have identified CRISPR in around half of all sequenced bacteria and around 90% of sequenced archaea.

CRISPR has been harnessed by researchers and is used as a gene editing tool for making changes to the double-stranded DNA in human cells. Rather than remove DNA inserted by viral invaders, the tool has been used to remove defective genes, or insert more beneficial genes, such as to improve yields in crop plants. CRISPR-Cas9 is the most widely used gene editing tool by researchers.

Dr. Feng Zhang has been heavily involved in the use of CRISPR-Cas9 since the system was first identified as a gene editing tool by Jennifer Doudna and Emmanuelle Charpentier, following the discovery by Francisco Mojica. Zhang was one of the pioneers who shifted attention from CRISPR in bacterial cells to the use of CRISPR in eukaryotic cells and played a key role in expanding the applications of CRISPR.

Zhang and his team investigated the origins of CRISPR to see if there were any other branches of the CRISPR family tree that could potentially be used for gene editing in medicine. They found a huge number of potential gene editing tools, including an entirely new family line dubbed Obligate Mobile Element Guided Activity (OMEGA). This variant operates in a similar way to CRISPR but is distinctly different, as it uses a new microbial IscB protein family as the molecular scissors along with previously unknown RNA guides (OMEGA-RNA) to direct the proteins to make the cuts in the right place. OMEGAs are believed to be the system from which CRISPR systems evolved.  Prior to the study, the microbial IscB protein family was not known.

IscB genes are present in bacteria and archaea but can also be fund in algal chloroplasts. The IscB genes are located close to regions of DNA that code for RNA molecules which act as guides for the proteins to direct them to sections of the DNA where cuts are made. The sheer number of alternative systems came as a big surprise. Conducting a big data search, the researchers identified more than a million potential genetic sites that encode one of the proteins used in OMEGA systems. The researchers also identified a protein family called TnpB, which also cuts the DNA when guided by RNA. The researchers suggest this family of proteins is the ancestor of the Ca13 enzyme.

The researchers demonstrated IscB proteins could make cuts to human DNA, but were far less efficient than Cas9 enzymes, although since the IscB proteins are 30% smaller than their Cas9 counterpart, they could have some important uses in certain applications.

“We are super excited about the discovery of these widespread programmable enzymes, which have been hiding under our noses all along,” said Zhang. “These results suggest the tantalizing possibility that there are many more programmable systems that await discovery and development as useful technologies.”

You can read more about the study in the paper – The widespread IS200/605 transposon family encodes diverse programmable RNA-guided endonucleases – which was recently published in Science. DOI: 10.1126/science.abj6856