New method opens the door to efficient genome writing in bacteria

Bioengineers at MIT have found a new way to efficiently manipulate bacterial genomes and program memories in bacterial cells by rewriting their DNA. With this approach, various forms of spatial and temporal information can be permanently stored over generations and retrieved by sequencing the DNA of the cells.

The new DNA writing technique, which the researchers call HiSCRIBE, is much more efficient than previously developed systems for DNA editing in bacteria, which have a success rate of only about 1 in 10,000 cells per generation. In a new study, the researchers showed that this approach could be used to store memory of cellular interactions or spatial location.

This technique could also make it possible to selectively manipulate, activate, or silence genes in certain types of bacteria that live in a natural community such as the human microbiome, the researchers said.

“With this new DNA writing system, we can edit bacterial genomes within complex bacterial ecosystems precisely and efficiently without the need for any form of selection,” says Fahim Farzadfard, former MIT postdoc and lead author of the study. “This enables us to do genome editing and DNA writing outside of the laboratory environment, be it to develop bacteria, optimize features of interest in situ, or investigate evolutionary dynamics and interactions in bacterial populations.”

Timothy Lu, associate professor of electrical and computer science and bioengineering at MIT, is the lead author of the study, which is published today in. appears Cell systems. Nava Gharaei, a former PhD student at Harvard University, and Robert Citorik, a former MIT student, are also authors of the study.

Write genomes and record memories

Lu’s lab has been working on using DNA to store information like memories of cellular events for several years. In 2014, he and Farzadfard developed a way to identify bacteria as “genomic tape recorder, “Engineering E. coli to store long-term memories of events such as chemical exposure.

To do this, the researchers engineered the cells to produce a reverse transcriptase enzyme called retron, which when expressed in the cells produces a single-stranded DNA (ssDNA) and a recombinase enzyme that insert a specific sequence ( “Write”) can from single-stranded DNA to a target location in the genome. This DNA is only produced when it is activated by the presence of a predetermined molecule or some other type of input, such as light. After the DNA is produced, the recombinase inserts the DNA into a preprogrammed location that can be anywhere in the genome.

This technique, which the researchers called SCRIBE, had relatively poor writing efficiency. In every generation of 10,000 E. coli Cells would only acquire the new DNA that the researchers were trying to build into the cells. This is in part because the E. coli have cellular mechanisms that prevent single-stranded DNA from being accumulated and integrated into their genome.

In the new study, the researchers tried to increase the efficiency of the process by adding some of the E. coli ‘s Defense mechanisms against single-stranded DNA. First, they deactivated enzymes called exonucleases that break down single-stranded DNA. They also turned off genes involved in a system called mismatch repair that normally prevents single-stranded DNA from integrating into the genome.

With these modifications, the researchers managed to incorporate the genetic changes they tried to introduce almost universally, creating an unprecedented and efficient way to manipulate bacterial genomes without selection.

“Because of this improvement, we were able to implement some applications that we could not do with the previous generation of SCRIBE or other DNA writing technologies,” says Farzadfard.

Cellular Interactions

In their 2014 study, the researchers showed that they could use SCRIBE to record the duration and intensity of exposure to a particular molecule. With their new HiSCRIBE system, they can track such exposures as well as additional types of events such as interactions between cells.

As an example, the researchers showed that they could follow a process called bacterial conjugation, in which bacteria exchange pieces of DNA. By incorporating a DNA “barcode” into the genome of each cell, which can then be exchanged with other cells, researchers can determine which cells have interacted with each other by sequencing their DNA to see which barcodes they carry.

This type of mapping could help researchers study how bacteria communicate with one another in aggregates such as biofilms. If a similar approach could be applied in mammalian cells, it could one day be used to map interactions between other cell types such as neurons, Farzadfard says. Viruses that can cross neural synapses could be programmed to carry DNA barcodes that researchers could use to track connections between neurons, offering a new way to map the brain’s connectome.

“We use DNA as a mechanism to record spatial information about the interaction of bacterial cells and perhaps, in the future, tagged neurons,” says Farzadfard.

The researchers also showed that this technique allows them to specifically manipulate the genome of a bacterial species within a community of many species. In this case, they introduced the gene for an enzyme that breaks down galactose in E. coli Cells that grow in culture with several other types of bacteria.

This type of species-selective editing could offer a new way to make antibiotic-resistant bacteria more susceptible to existing drugs by silencing their resistance genes, the researchers say. However, such treatments would likely take several years more years of research to develop, they say.

The researchers also showed that using this technique they can develop a synthetic ecosystem of bacteria and bacteriophages that can continuously rewrite certain sections of their genome and develop autonomously at a faster rate than would be possible through natural evolution. In this case, they were able to optimize the cells’ ability to absorb the lactose consumption.

“This approach could be used in the evolutionary development of cellular traits or in experimental evolution studies by letting the evolutionary tape play over and over,” says Farzadfard.

Republished with permission from MIT. Photo: MIT researchers have found a way to program memories in bacterial cells by rewriting their DNA more efficiently. Photo credit: MIT News