A new gene-editing protein, CasX, may give CRISPR-Cas9 a run for its money. UC Berkeley scientists have determined the unique structure of CasX (grey), revealing that this pint-sized Cas enzyme is dominated by RNA (red) that directs it to specific sequences of DNA (blue), where it binds and cuts the DNA. Credit: University of California – Berkeley
While CRISPR Cas9 remains a mainstay in the genomics world, researchers have now found a much smaller protein called CasX that could be used for gene editing.
In 2017, a research team from the University of California Berkeley discovered in some of the world’s smallest bacteria a protein similar to Cas9 but substantially smaller, which could enable researchers to deliver a gene editor for both bacteria and human cells.
According to the study, the RNA-guided CRISPR-associated (Cas) proteins Cas9 and Cas12a provide adaptive immunity against invading nucleic acids, and function as powerful tools for genome editing in a wide range of organisms.
CasX is similar to both Cas9 and Cas12 in a number of ways, but different enough to where the researchers believe it evolved in bacteria independent of other Cas proteins. CasX can cut double-stranded DNA, much like Cas9, while also binding to DNA to regulate genes and can be targeted to specific DNA sequences similar to other Cas proteins.
Despite originating in bacteria that is not found in humans, the researchers combed through a database of microbes found in groundwater and sediment and discovered that the human immune system should accept CasX more easily than Cas9, as some doctors believe that Cas9 can create an immune reaction in patients treated with CRISPR therapies.
“The immunogenicity, delivery and specificity of a genome-editing tool are all vitally important,” co-lead author Benjamin Oakes, a former UC Berkeley graduate student and current Entrepreneurial Fellow in the Innovative Genomics Institute (IGI), said in a statement. “We’re excited about CasX on all of these fronts.”
The researchers used a cryo-electron microscope to capture snapshots of the CasX protein going through the motions of editing a gene. Here, they found more details on CasX’s molecular makeup and shape, proving that the protein evolved independently of Cas9 and shares no common ancestry.
“The first thing that jumps out is how the highly unique domains accomplish similar roles to what we have seen with other RNA-guided DNA-binding proteins,” Oakes said. “CasX’s minimal size, with no fat on the bone, helps to clearly demonstrate there is a basic recipe that nature uses. Understanding this recipe will help us to better evolve and engineer genome editing tools for our purposes rather than nature’s.”
The researchers will now work to further develop the next gene-editing tool.
“The culmination of biochemistry, genome editing and structure experiments within this single study is a prime example of the comprehensive efforts that are underway at the IGI,” Jennifer Doudna, IGI’s executive director, a UC Berkeley professor of molecular and cell biology and of chemistry and a Howard Hughes Medical Institute Investigator, said in a statement. “We aren’t just looking to uncover the next pair of molecular scissors. We want to build the next Swiss Army knife.”
The study was published in Nature.
