researchers pioneer a new method for modifying genes in human cells |

SAN FRANCISCO, January 5, 2022 / PRNewswire / – Over the past decade, the CRISPR genome editing system has revolutionized molecular biology, giving scientists the ability to edit genes inside living cells for research or medical applications. Today, researchers at the Gladstone Institutes have refined an additional system for more efficient gene editing, using molecules called retrons.

Let’s go back, the group reported in the newspaper Nature Chemistry Biology, can be optimized for efficiency and used to modify genes in a variety of cell types, from fungi to human cells.

“This work really consolidates the retrons as a platform that can be used in all organisms,” says Gladstone deputy investigator Seth Ship, PhD, lead author of the new study. “We can make precise changes to genes easier, faster and more efficiently than with current approaches.”

One stop shop for gene editing

Most of the current CRISPR-based gene-editing technologies involve cutting a section of DNA from a cell’s genome and then introducing new genetic material called “template DNA” to replace it. As the cell repairs the places where an existing gene has been cut, the template DNA is integrated.

This template DNA is normally produced in the laboratory and then introduced into cells from the outside. The protein that cuts the cell’s genome, called Cas9, is shipped separately. Neither Cas9 nor template DNA enters every cell, which limits the efficiency of CRISPR gene editing.

Retrons, however, act like DNA factories, producing abundant copies of template DNA from inside cells. In addition, the retrons can be supplied with the rest of the CRISPR components so that cells simultaneously obtain all the material needed for gene editing – the genetic codes of model DNA, Cas9 and the molecules that help researchers track. the changes that have been made.

“This means that we only have to introduce one element in each cell”, explains Santiago lopez, a graduate student of the Shipman Lab and first author of the new article. “This greatly simplifies the process and opens the door to new types of experiences.”

Retron reengineering

Both retrons and CRISPR originate from bacteria; both are defense mechanisms bacteria use to modify DNA in response to infections. After the advent of CRISPR genome editing, in which the CRISPR system was co-opted to selectively target genes in other types of cells, some researchers began to investigate whether retrons could be used to provide the models for precise gene editing. However, the roles of different sections of the retron structure in its function – and how to modify those sections to improve retrons – are unknown.

“The retron system has evolved to help defend bacteria,” says Shipman, who is also an assistant professor of bioengineering and therapeutic sciences at UC San Francisco (UCSF). “But we wanted to change it from what it normally does to what we want it to do – produce models for gene editing.”

In the new study, Shipman’s group designed E. coli retrons to create hundreds of new variations. They tested each new variant and discovered a series of changes that together led to an 8-10-fold increase in the amount of template DNA ultimately produced by the retron in E. coli cells.

Next, the researchers tested the new redesigned retron system in the fungus Saccharomyces cerevisiae (baker’s yeast) and in cultured human cells, and they found that this optimized system worked in all cases. This was the first demonstration of the use of retrons in human cells and their portability between cell types.

Since the team could now refine exactly the amount of template DNA produced by retrons, they were also able to show that when retrons produce high levels of template DNA, it increases the efficiency of gene editing. .

“Our study shows for the first time that the more DNA template we can produce, the better the genome is edited,” says Shipman. “Better, more accurate editing ultimately means more efficient and safer genomic drugs and more advanced basic research.”

Take tools from bacteria

Retrons are immediately useful, according to Shipman, as a research tool to edit genes in different types of cells in the lab. While the platform is not yet ready for use in humans, it also has the potential to help modify genes for therapeutic purposes, for example by repairing genetic mutations that cause disease.

Since different bacteria contain different retrons, his group also plans to explore whether other variations of retrons have advantages over the E. coli retron they optimized in this study.

“We take a general approach where we extract parts that we find in bacteria and domesticate them for our own use,” says Shipman. “It has already been incredibly successful in developing new tools, but I think we are just starting to reap the benefits of applying these tools in biotechnology.”

About the study

The article “Accurate Genome Editing Across Realms of Life Using DNA Derived from Retrons” was published in the journal Nature Chemistry Biology to 23 December 2021. Other authors are Kate crawford, Sierra Lear, and Santi Bhattarai-Kline from the Gladstone Institutes.

The work was supported by the Simons Foundation Autism Research Initiative, the Pew Scholars Program in the Biomedical Sciences, the Sandler Program for Breakthrough Biomedical Research at UCSF, the National Institutes of Health (1DP2GM140917-01), the LK Whittier Foundation, the National Science Foundation (Graduate Research Fellowships 2019247827 and 2034836), a Berkeley Scholarship for Graduate Studies, and the Discovery Fellows program at UCSF.

About Gladstone Institutes

To ensure that our work is doing the greatest good, the Gladstone Institutes focus on conditions with profound medical, economic and social impact – unresolved illnesses. Gladstone is an independent, non-profit life science research organization that uses visionary science and technology to beat disease. He has an academic affiliation with the University of California, San Francisco.

Media contact:

Julie langelier, Associate Director, Communications, julie.langelier@gladstone.org, 415.734.5000,

1650 Owens Street, San Francisco, California 94158, gladstone.org, Twitter: @GladstoneInst

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