It has been difficult to track the regulation of noncoding RNA. Now, researchers from Helmholtz Munich and the Technical University of Munich (TUM) report they have developed a minimally invasive reporter system that enables monitoring of RNA production of both coding and noncoding RNA.

Their findings are published in the journal Nature Cell Biology in an article titled, “Intron-encoded cistronic transcripts for minimally invasive monitoring of coding and noncoding RNAs.”

“Despite their fundamental role in assessing (patho)physiological cell states, conventional gene reporters can follow gene expression but leave scars on the proteins or substantially alter the mature messenger RNA,” wrote the researchers. “Multi-time-point measurements of noncoding RNAs are currently impossible without modifying their nucleotide sequence, which can alter their native function, half-life, and localization. Thus, we developed the intron-encoded scarless programmable extranuclear cistronic transcript (INSPECT) as a minimally invasive transcriptional reporter embedded within an intron of a gene of interest.”

“Unlike previous methods, INSPECT encodes sequences for reporter proteins in modified introns. These are sequences in the pre-mature RNA molecule that are removed naturally and eliminated by the cell during processing. INSPECT stabilizes the introns such that, rather than being degraded after removal, they are transported to the cellular cytoplasm where they are translated into reporter proteins,” explained first author Dong-Jiunn Jeffery Truong, PhD, group leader at Helmholtz Munich. The researchers can then use conventional methods to detect reporter protein signals such as fluorescence.

The new molecular biology tool not only solves the problem of tracking the generation of noncoding RNA, but also offers advantages for studying coding RNA. Current reporter systems often run the risk of damaging the RNA or proteins under investigation. Rather than modifying the completed RNA or the proteins, INSPECT modifies the introns.

“INSPECT adds an important molecular biology tool to the biomedical toolbox. It makes it easier to study the role of certain noncoding RNA molecules in cell development and to explore how their regulation can be modulated, for example, to prevent them from turning into cancer cells,” said Gil Westmeyer, PhD, professor, neurobiological engineering, TUM. “In combination with the minimally invasive reporter system EXSISERS, which we previously developed to study protein isoforms, it may be possible in the future to study an entire genetic regulation process from RNA processing to the production of specific protein variants in living cells.”

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