By Anne Trafton, Massachusetts Institute of Technology
August 8, 2021
MIT
researchers have devised a way to program memories into bacterial cells
by rewriting their DNA more efficiently. Credit: MIT News, iStockphoto
Technique for editing bacterial genomes can record
interactions between cells, may offer a way to edit genes in the human
microbiome.
La técnica de edición de genomas bacterianos puede registrar las interacciones entre las células y podría ofrecer una forma de editar los genes del microbioma humano.
Los ingenieros biológicos del MIT han ideado una nueva forma de editar eficazmente los genomas bacterianos y programar memorias en las células bacterianas reescribiendo su ADN. Con este método, se pueden almacenar permanentemente varias formas de información espacial y temporal durante generaciones y recuperarlas mediante la secuenciación del ADN de las células.
La nueva técnica de escritura del ADN, que los investigadores denominan HiSCRIBE, es mucho más eficaz que los sistemas desarrollados anteriormente para editar el ADN de las bacterias, que sólo tenían una tasa de éxito de aproximadamente 1 de cada 10.000 células por generación. En un nuevo estudio, los investigadores demostraron que este método podría utilizarse para almacenar la memoria de las interacciones celulares o la localización espacial.
Según los investigadores, esta técnica también podría permitir editar, activar o silenciar genes de forma selectiva en determinadas especies de bacterias que viven en una comunidad natural como el microbioma humano.
"Con este nuevo sistema de escritura del ADN, podemos editar con precisión y eficacia los genomas bacterianos sin necesidad de ninguna forma de selección, dentro de ecosistemas bacterianos complejos", afirma Fahim Farzadfard, antiguo postdoctorado del MIT y autor principal del artículo. "Esto nos permite llevar a cabo la edición del genoma y la escritura del ADN fuera de los entornos de laboratorio, ya sea para la ingeniería de las bacterias, la optimización de los rasgos de interés in situ, o el estudio de la dinámica evolutiva y las interacciones en las poblaciones bacterianas."
Timothy Lu, profesor asociado de ingeniería eléctrica e informática y de ingeniería biológica del MIT, es el autor principal del estudio, que se publicó el 5 de agosto de 2021 en Cell Systems. Nava Gharaei, ex estudiante de posgrado de la Universidad de Harvard, y Robert Citorik, ex estudiante de posgrado del MIT, también son autores del estudio.
Escritura del genoma y grabación de memorias
Durante varios años, el laboratorio de Lu ha estado trabajando en formas de utilizar el ADN para almacenar información, como la memoria de eventos celulares. En 2014, él y Farzadfard desarrollaron una forma de emplear las bacterias como una "grabadora genómica", diseñando E. coli para almacenar recuerdos a largo plazo de eventos como una exposición química.
Para conseguirlo, los investigadores diseñaron las células para que produjeran una enzima transcriptasa inversa llamada retron, que produce un ADN monocatenario (ssDNA) cuando se expresa en las células, y una enzima recombinasa, que puede insertar ("escribir") una secuencia específica de ADN monocatenario en un lugar determinado del genoma. Este ADN sólo se produce cuando se activa por la presencia de una molécula predeterminada u otro tipo de entrada, como la luz. Una vez producido el ADN, la recombinasa lo inserta en un lugar preprogramado, que puede estar en cualquier parte del genoma.
Esta técnica, que los investigadores llamaron SCRIBE, tenía una eficacia de escritura relativamente baja. En cada generación, de cada 10.000 células de E. coli, sólo una adquiría el nuevo ADN que los investigadores intentaban incorporar a las células. Esto se debe, en parte, a que las E. coli tienen mecanismos celulares que impiden que el ADN monocatenario se acumule e integre en sus genomas.
En el nuevo estudio, los investigadores trataron de aumentar la eficacia del proceso eliminando algunos de los mecanismos de defensa de E. coli contra el ADN monocatenario. En primer lugar, desactivaron unas enzimas llamadas exonucleasas, que descomponen el ADN monocatenario. También eliminaron los genes que intervienen en un sistema denominado reparación de desajustes, que normalmente impide la integración del ADN monocatenario en el genoma.
Con esas modificaciones, los investigadores pudieron lograr la incorporación casi universal de los cambios genéticos que intentaban introducir, creando una forma inédita y eficiente de editar genomas bacterianos sin necesidad de selección.
"Gracias a esa mejora, pudimos realizar algunas aplicaciones que no podíamos hacer con la generación anterior de SCRIBE o con otras tecnologías de escritura de ADN", afirma Farzadfard.
Biological engineers at MIT have devised a new way to efficiently edit bacterial genomes and program memories into bacterial cells by rewriting their DNA.
Using this approach, various forms of spatial and temporal information
can be permanently stored for generations and retrieved by sequencing
the cells’ DNA.
The new DNA writing technique, which the researchers call HiSCRIBE,
is much more efficient than previously developed systems for editing DNA
in bacteria, which had a success rate of only about 1 in 10,000 cells
per generation. In a new study, the researchers demonstrated that this
approach could be used for storing memory of cellular interactions or
spatial location.
This technique could also make it possible to selectively edit,
activate, or silence genes in certain species of bacteria living in a
natural community such as the human microbiome, the researchers say.
“With this new DNA writing system, we can precisely and efficiently
edit bacterial genomes without the need for any form of selection,
within complex bacterial ecosystems,” says Fahim Farzadfard, a former
MIT postdoc and the lead author of the paper. “This enables us to
perform genome editing and DNA writing outside of laboratory settings,
whether to engineer bacteria, optimize traits of interest in situ, or
study evolutionary dynamics and interactions in the bacterial
populations.”
Timothy Lu, an MIT associate professor of electrical engineering and
computer science and of biological engineering, is the senior author of
the study, which was published on August 5, 2021, in Cell Systems.
Nava Gharaei, a former graduate student at Harvard University, and
Robert Citorik, a former MIT graduate student, are also authors of the
study.
Genome writing and recording memories
For several years, Lu’s lab has been working on ways to use DNA to
store information such as memory of cellular events. In 2014, he and
Farzadfard developed a way to employ bacteria as a “genomic tape recorder,” engineering E. coli to store long-term memories of events such as a chemical exposure.
To achieve that, the researchers engineered the cells to produce a
reverse transcriptase enzyme called retron, which produces a
single-stranded DNA (ssDNA) when expressed in the cells, and a
recombinase enzyme, which can insert (“write”) a specific sequence of
single-stranded DNA into a targeted site in the genome. This DNA is
produced only when activated by the presence of a predetermined molecule
or another type of input, such as light. After the DNA is produced, the
recombinase inserts the DNA into a preprogrammed site, which can be
anywhere in the genome.
That technique, which the researchers called SCRIBE, had a relatively low writing efficiency. In each generation, out of 10,000 E. coli
cells, only one would acquire the new DNA that the researchers tried to
incorporate 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 genomes.
In the new study, the researchers tried to boost the efficiency of the process by eliminating some of E. coli’s
defense mechanisms against single-stranded DNA. First, they disabled
enzymes called exonucleases, which break down single-stranded DNA. They
also knocked out genes involved in a system called mismatch repair,
which normally prevents integration of single-stranded DNA into the
genome.
With those modifications, the researchers were able to achieve
near-universal incorporation of the genetic changes that they tried to
introduce, creating an unparalleled and efficient way for editing
bacterial genomes without the need for selection.
“Because of that improvement, we were able to do some applications
that we were not able to do with the previous generation of SCRIBE or
with other DNA writing technologies,” Farzadfard says.
Cellular interactions
In their 2014 study, the researchers showed that they could use
SCRIBE to record the duration and intensity of exposure to a specific
molecule. With their new HiSCRIBE system, they can trace those kinds of
exposures as well as additional types of events, such as interactions
between cells.
As one example, the researchers showed that they could track a
process called bacterial conjugation, during which bacteria exchange
pieces of DNA. By integrating a DNA “barcode” into each cell’s genome,
which can then be exchanged with other cells, the researchers can
determine which cells have interacted with each other by sequencing
their DNA to see which barcodes they carry.
This kind of mapping could help researchers study how bacteria
communicate with each other within aggregates such as biofilms. If a
similar approach could be deployed in mammalian cells, it could someday
be used to map interactions between other types of cells such as
neurons, Farzadfard says. Viruses that can cross neural synapses could
be programmed to carry DNA barcodes that researchers could use to trace
connections between neurons, offering a new way to help map the brain’s
connectome.
“We are using DNA as the mechanism to record spatial information
about the interaction of bacterial cells, and maybe in the future,
neurons that have been tagged,” Farzadfard says.
The researchers also showed that they could use this technique to
specifically edit the genome of one species of bacteria within a
community of many species. In this case, they introduced the gene for an
enzyme that breaks down galactose into E. coli cells growing in culture with several other species of bacteria.
This kind of species-selective editing could offer a novel way to
make antibiotic-resistant bacteria more susceptible to existing drugs by
silencing their resistance genes, the researchers say. However, such
treatments would likely require several years more years of research to
develop, they say.
The researchers also showed that they could use this technique to
engineer a synthetic ecosystem made of bacteria and bacteriophages that
can continuously rewrite certain segments of their genome and evolve
autonomously with a rate higher than would be possible by natural
evolution. In this case, they were able to optimize the cells’ ability
to consume lactose consumption.
“This approach could be used for evolutionary engineering of cellular
traits, or in experimental evolution studies by allowing you to replay
the tape of evolution over and over,” Farzadfard says.
Reference: “Efficient retroelement-mediated DNA writing in bacteria”
by Fahim Farzadfard, Nava Gharaei, Robert J. Citorik and Timothy K. Lu, 5
August 2021, Cell Systems.
DOI: 10.1016/j.cels.2021.07.001
The research was funded by the National Institutes of Health, the
Office of Naval Research, the National Science Foundation, the Defense
Advanced Research Projects Agency, the MIT Center for Microbiome
Informatics and Therapeutics, the NSF Expeditions in Computing Program
Award, and the Schmidt Science Fellows Program.
https://scitechdaily.com/mit-researchers-devised-a-way-to-program-memories-into-bacterial-cells-by-rewriting-their-dna/
