A new toolbox helps characterize internal ribosome entry sites within cells

Recently, beans, recognized as activity regulators of gene expression at the protein biosynthetic level, have ribosomes (one of the oldest molecular machines in evolutionary terms). This is an important process of cell development and function where genetic information is converted to proteins. The final step in which information encoded with messenger RNA (mRNA) is transferred is known as translation.

The “immunobiochemistry” research group, led by Professor Kathrin Leppek of the UKB Institute for Clinical Chemistry and Clinical Pharmacology (IKCKP), is investigating the regulation of translation using the direct interaction of ribosomes and mRNAs.

“Ribosomes, such as proteins and RNA structures and their associated factors, as essential central translation mechanisms for all lifespans, are the focus of our research,” says Professor Leppek, a member of the two clusters of immune infections at the University of Bonn. “Ribosomal composition affects selective translation, and there is growing evidence that customized ribosomes preferentially bind and translate specific mRNAs.”

The role of IRES in gene expression

As an example of such a structure that plays an important role in the initiation of translation and thus regulating gene expression, Bonn researchers are currently investigating the site of internal ribosome entry (IRES). These are special folded sequences within RNA strands, particularly well known for virus genetic material, to hijack host ribosomes after infection. For example, hepatitis C or polioviruses can begin the production of new viral proteins independent of all initiation factors thanks to the IRES element.

By recruiting ribosomes, IRES sequence allows for the initiation of translation independent of the 5′ cap of mRNA. This is a protective cap equipped with the host’s own mRNA strands, allowing translation under normal conditions, but is blocked under viral infection.

There is no uniform standard for clear characterization of IRES

IRE was first described in the viral genome where it allowed virus replication in infected cells by recruiting host ribosomes. However, in recent decades, more and more air has been reported in eukaryotic cells, unlike viruses, with nuclei.

“This reinforces the general view that these elements are also involved in regulating eukaryotic cell translation,” says Philipp Koch, a Leppek research group at UKB and a doctoral student at Bonn University. He is the first author of a paper published in the EMBO Journal.

His colleague and co-author, Martin Heyman, who is also a doctoral student at the University of Bonn, added: “However, the precise and reliable features of newly described Aires, especially from eukaryotic mRNA, were difficult due to the technical hurdles and artifacts associated with existing technologies used up to now.”

In their current research work, Bonn researchers have compiled and tested several versatile techniques that will allow for robust characterization of IRES in the future. One important method is the use of a circular RNA reporter that can be used to confirm the IRES-mediated activity of RNA elements.

Other techniques include quantitative staining techniques for individual mRNAs in mouse embryonic tissues and measuring the translation rate of mRNAs, including individual IREs.

“This comprehensive toolbox, which can be applied to cultured cells and embryonic tissue, represents a new gold standard for robust testing and characterization of Aile,” says Koch. Professor Leppek adds, “The powerful IRES elements are directly related to synthetic biology and emerging mRNA therapy.”