By exploring the evolutionary origins of an enzyme used in the CRISPR genome-editing system, researchers have unearthed more than one million other potential editors lurking in microbial genomes.
The study, published on 9 September in Science1, found the new editing enzymes among a family of proteins called IscB. These proteins are thought to be the ancestors of the enzyme Cas9 — known as CRISPR’s molecular scissors. During genome editing, Cas9 teams up with a snippet of RNA that guides the enzyme to find and cut a specific DNA sequence. The technique’s reliance on RNA as a guidance system is a key reason for its versatility and widespread use, because it allows researchers to easily target Cas9 to the region of the genome they want to alter.
The discovery of other RNA-targeted enzymes capable of cutting DNA could yield further tools for genome editing, says study lead author Feng Zhang, a molecular biologist at the Massachusetts Institute of Technology in Cambridge (MIT). “These programmable proteins are very useful, beyond basic biological interest,” he says. “And this mechanism of RNA-guided DNA recognition is likely something that nature has created independently multiple times.”
Although researchers have harnessed it for genetic engineering, CRISPR is thought to be a microbial defence system that allows bacteria and other single-celled organisms called archaea to fend off viruses and other genetic invaders by sending Cas9 to chew up their DNA. Computational studies suggested that Cas9 probably evolved from proteins in the IscB family, which are encoded by transposons, or ‘jumping genes’, that can hop around to new locations in the genome. Until now, the function of IscB proteins has been unclear.
Zhang and his colleagues found that the DNA responsible for encoding IscB proteins is often located near the DNA for a class of RNA molecules that they dubbed ωRNAs. They also discovered that some IscB proteins can cleave DNA at a site specified by the sequence of an ωRNA, much like Cas9 and its guide RNA.
The team went on to investigate another family of proteins, called TnpB, which are thought to be the ancestors of another DNA-slicing CRISPR-associated enzyme called Cas12. They found that some of these proteins could also cut DNA when guided by ωRNA.
Database searches turned up more than one million genes that could carry the code for TnpB proteins, and some organisms contain more than 100 copies of these genes, says Soumya Kannan, a molecular biologist at MIT and a co-first author of the study.
And IscB genes showed up not only in bacteria and archaea, but also in the light-harvesting chloroplast inside the cell of an alga. This is the first time such genome-editing systems have been found in a eukaryote (the group of organisms whose cells contain nuclei, which includes all plants and animals) — a surprising result that suggests that they are more widespread than previously thought. “Every time I give a talk, people always ask me if we’ve seen CRISPR activity in a eukaryotic cell,” says Zhang. “Now, I can finally say ‘yes’.”
In nature, these genes could be performing various functions, including defence or regulating the expression of other genes. And in the lab, the discovery could yield a treasure trove of editing tools. Zhang and his team found that IscB could be used to cut human DNA, although at a lower efficiency than the popular CRISPR–Cas9 system. But Zhang says the IscB system could be improved, and notes that the small size of the IscB protein might make it easier to work with for some applications.
For geneticist Gaetan Burgio at the Australian National University in Canberra, the real beauty of the study is its contribution to understanding evolution — and to finally assigning a possible function to such a large and prevalent group of proteins as IscBs. “It’s absolutely fascinating,” he says. “It fills an important gap: we didn’t really know how these CRISPR systems became CRISPR.”
Altae-Tran, H. et al. Science https://doi.org/10.1126/science.abj6856 (2021).