Different
"events" such as infections by viruses, as well as the exposure to
environmental toxins or other forms of stress, change the activity of genes
thereby leaving molecular traces inside the cell. These changes happen mainly
at the level of messenger RNA (mRNA). These are molecules that encode genetic
information when genes become activated and read, a process known as
transcription. Researchers can accurately investigate the activity of a gene by
measuring the mRNA molecules present in a cell. However, the traces of gene
transcription disappear rapidly: mRNA is highly instable, and cells often
degrade it after a short time.
ETH
researcher Randall Platt and his colleagues in the Department of Biosystems
Science and Engineering have now developed a molecular recording system that
writes transcriptional events into DNA where they can be permanently stored and
later accessed to by sequencing.
To
create their "recording device," Platt's doctoral students Florian
Schmidt and Mariia Cherepkova employed the CRISPR-Cas system. CRIPSR-Cas is an
adaptive immune system in bacteria and archaea. The system functions like an
immunological memory device by recording genetic information about pathogens
infecting the cell. This genetic information is recorded in a specific stretch
of DNA known as a CRISPR array -- a process called acquisition.
CRISPR
arrays are capable of storing short sequences of DNA, known as 'spacers',
originating from a pathogen. Spacers are separated from each other by short
identical DNA sequence called direct repeats, just like pearls on a string.
The
researchers worked with the gut bacterium Escherichia coli, introducing the
genes for the CRISPR-Cas system from a different bacterial species. One of
those Cas genes is fused to a reverse transcriptase, an enzyme that uses an RNA
molecule to produce DNA encoding the same information -- in other words, it
transcribes RNA back into DNA.
The
Escherichia coli cells supplied with the foreign genes for this CRISPR-Cas were
able to produce a protein complex that binds short mRNA molecules. The reverse
transcriptase translates these RNA spacers into DNA, containing the same
information as the original RNA, and subsequently storing them in the CRISPR
array. This process can occur multiple times such that new spacers are added to
the CRISPR array in reverse chronological order, so the most recently acquired
piece of DNA is always first.
In
principle, this makes it possible to record any number spacers within a CRISPR
array. Since DNA is very stable, the information recorded in them is stored for
a long time and is also passed on from one generation of bacteria to the next.
"Our
system is a biological data logger. It records the genetic response of bacteria
to external influences and enables us to access that information even after
many bacterial generations" says Florian Schmidt, the lead author of the
study, which was recently published in the journal Nature.
ETH
Professor Randall Platt says, "Researchers have been working on creating
forms of synthetic cellular memory for a long time, but we are the first to
develop one that can record information about the expression of each gene in a
cell over time." The researchers have spent over two years working on this
system.
Until
now, researchers were limited to measuring mRNA at only a single snapshot in
time. Taking these snapshots generally means destroying the cell, extracting
its mRNA, and then quantifying them. In contrast, the new CRISPR-Cas RNA
recording system records the history of cell, allowing researchers to
effectively access the entire cellular log book rather than just a single point
in time.
As
part of their study, the ETH researchers recorded the reaction of E. coli
bacteria equipped with the data logger to the herbicide paraquat. This
substance provokes changes in mRNA transcription within the cells, and the
scientists could read out this response from the CRISPR arrays even days after
the herbicide exposure. Without the data logger, any molecular traces of the
bacteria's contact with the herbicide would have long since been broken down
and the information lost.
Biological
data loggers like this, in addition to being interesting for research purposes,
could also conceivable be used as a kind of sensor, to measure environmental
toxins such as the herbicide, or in diagnostics. The present study intriguingly
demonstrates the feasibility of such an approach, however practical
applications are still a long way off. Randall Platt's research team in Basel
is already working on transferring the system to other cell types and paving the
way for its effective use as a diagnostic tools.
See:
Florian
Schmidt, Mariia Y. Cherepkova, Randall J. Platt. Transcriptional recording by
CRISPR spacer acquisition from RNA. Nature, 2018; DOI:
10.1038/s41586-018-0569-1
Posted by Dr. Tim Sandle,
Pharmaceutical Microbiology
Posted by Dr. Tim Sandle,
Pharmaceutical Microbiology
Posted by Dr. Tim Sandle,
Pharmaceutical Microbiology
