Next revolution in gene editing?


In just 10 years, CRISPR revolutionized gene editing. The technology has sparked a wave of new scientific discoveries in cancer, Alzheimer’s and heart disease, and several gene therapies using the gene editor will soon be available to patients.

But the first-generation CRISPR-Cas9 gene-editing system has some drawbacks. Although it allows scientists to edit the genome at a precise location, it is often difficult to control the change made and the final edit difficult to predict.

Next-generation gene editors are needed, and a promising new technology based on parasitic “jumping genes” is being developed in the lab of Sam Sternberg, PhD, assistant professor of biochemistry and molecular biophysics, and may be able to do so to overcome limitations of first-generation gene editing.

Taming jumping genes

Jumping genes – also called transposons – are mobile regions in the genome that jump randomly to other parts of the genome. Although transposons often cause damage to the organism due to this lack of specificity, they mobilize with highly efficient “integrase” enzymes capable of inserting large genetic payloads into the genome. The Sternberg lab previously discovered a unique family of bacterial transposons that naturally utilize CRISPR to control their specificity, thus offering a new mode of gene editing that exhibits both remarkable fidelity and efficient DNA insertion activity.

Sternberg and his colleagues have previously shown that the transposon-based gene editor is less likely to edit unintended genomic targets than the original CRISPR technology.

One reason for the remarkable accuracy of the new editor, the researchers report today in the journal Nature, lies in a single protein in the editing complex.

Molecular details of the TnsC protein, including its 3D structure (above), should help researchers optimize the new gene editor for genome manipulation of human cells. Image: Hoffmann, Kim, Beh et al., Nature (2022).

In the new work, Sternberg’s team, led by graduate student Florian Hoffmann and postdoctoral fellows Minjoo Kim, PhD, and Leslie Beh, PhD, used biochemical techniques to identify all sites in the genome to which the publisher’s proteins go during gene insertion bind in a bacterium. This approach revealed that while the major protein complex of the editor binds many off-target sites, only a small number of these sites can recruit the TnsC protein, which is essential for gene insertion.

The process works similar to a set of search filters that a user might apply during an advanced online search. “These gene-editing systems first take samples from a large set of possible target sites, and then TnsC is the filter that selects only the right target site for DNA integration,” says Sternberg.

The researchers also revealed the 3D structure of TnsC in work led by Israel Fernández, PhD, a collaborator at St. Jude Children’s Research Hospital, which revealed that this critical protein forms a ring around DNA and is the genomic target site for precisely positioned the transposon insertion .

Human applications?

The new editor could potentially insert complete, functional genes into patient cells.

The new details about the molecular mechanics of these transposons when editing bacterial genomes should help researchers optimize the new gene editors for human cell genetic engineering and clinical applications, says Sternberg.

The transposon-based editor’s ability to make large gene insertions with exquisite precision could be particularly useful in certain types of gene therapy. While the original CRISPR-Cas9 tool is limited to small changes, the new editor could potentially insert complete, functional genes into patient cells.

Sternberg says that although clinical applications like this require further technological development, his team has already met a challenge and successfully adapted the gene editor to work in human cells in the laboratory.


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