Nrue sub-journal: Molecular monitoring of RNA regulation

The better we understand cellular processes such as RNA regulation, the better molecular therapies
we can develop.
Until now, it has been particularly difficult to track the regulation of non-coding RNAs, which are RNAs
that are not further converted into proteins.
A research team from the Helmholtz University of Munich and the Technical University of Munich (TUM) has now developed a minimally invasive reporting system capable of highly sensitive monitoring
of RNA production of coding and non-coding RNAs.

During cellular processes, our genetic DNA information is transcribed into RNA, which is then further processed, and then it either serves as a blueprint for proteins or performs cellular functions themselves
.
The type and amount of RNA produced reveals a lot about our cells
.
For example, in the case of infection, cells produce more RNA molecules that encode proteins
involved in the immune response.

When DNA molecules are translated into proteins via RNA, researchers can track the process
with existing reporter systems.
However, not all human genes code for proteins
.
Most human genes are non-coding, including long non-coding RNA (lncRNA) genes
.
These RNA molecules have more than 200 building blocks, and they do not serve as blueprints
for proteins.
Instead, they control important processes
in cells.
Preliminary research suggests that lncRNAs are involved in processes such as regulating RNA production, the organization of the nuclear structure, or the switching of certain enzymes
.

Despite the importance of lncRNAs in cellular processes, studying lncRNAs with existing methods has been difficult
.
So far, this is only partially possible, such as in fixed cells at specific time points, because classical reporting systems based on translation into proteins cannot be used
.

INSPECT allows monitoring of non-coding RNAs

A solution to a new journalistic system has now been found: inspections
.
A team working with Gil Westmeyer, professor of neurobioengineering at the Technical University of Munich and director of the Helmholz Institute for Synthetic Biomedicine in Munich, has now published the newly developed reporting system
in the journal Nature Cell Biology.

"Unlike previous methods, INSPECT encodes to modify reporter protein sequences
in introns.
These are sequences in immature RNA molecules that are naturally removed
by the cell during processing.
INSPECT stabilizes introns so that they are not degraded after removal, but instead transported into the cytoplasm of cells translated into reporter proteins," explains
first author Dong-Jiunn Jeffery Truong.
Researchers can then use traditional methods to detect reporter protein signals, such as fluorescence
.

INSPECT modifies neither the finished RNA nor the protein

This new molecular biology tool not only solves the problem of tracking the generation of non-coding RNAs, but also provides advantages
for the study of coding RNAs.
Current reporter systems often run the risk of damaging the RNA or protein being studied, for example, because they must fuse directly with the RNA being studied in order to be co-translated into proteins
.
Instead of modifying the finished RNA or protein, INSPECT modifies the intron
.

The team demonstrated the functionality of INSPECT with a variety of examples of
coding and non-coding RNAs.
They tracked the production of RNA from interleukin 2, a protein that is produced in large quantities
in the response to infection.
They also performed highly sensitive monitoring of the production of two lncRNAs and tracked regulatory changes
during the investigation.

INSPECT adds an important molecular biology tool
to the biomedical toolbox.
This makes it easier to study the role of certain non-coding RNA molecules in cell development and explore how to regulate their regulation, for example, by preventing them from becoming cancerous
.
Combined with our previously developed minimally invasive reporter system EXSISERS for studying protein isoforms, it may be possible to investigate the entire genetic regulatory process
from RNA processing to the production of specific protein variants in living cells in the future.
”

Intron-encoded cistronic transcripts for minimally-invasive monitoring of coding and non-coding RNAs, Nature Cell Biology (2022)