When a virus attacks, the body’s immune response has two basic layers of defense: the innate and the adaptive immune systems. While the adaptive immune system grows stronger against a specific pathogen as the body is exposed to it multiple times and which forms the basis of vaccinations, the innate immune system is an assortment of molecular mechanisms that work against a broad range of pathogens at a basic level. The Kobe University virologist SHOJI Ikuo says, “The new coronavirus, however, is so infectious that we wondered what clever mechanisms the virus employs to evade the innate immune system so effectively.”

Shoji’s team previously worked on the immune response to hepatitis viruses and investigated the role of a molecular tag called “ISG15” the innate immune system attaches to the virus’s building blocks. Having learned that the novel coronavirus has an enzyme that is especially effective in removing this tag, he decided to use his team’s expertise to elucidate the effect of the ISG15 tag on the coronavirus and the mechanism of the virus’s countermeasures.

In a paper in the Journal of Virology, the Kobe University-led team is now the first to report that the ISG15 tag gets attached to a specific location on the virus’s nucleocapsid protein, the scaffold that packages the pathogen’s genetic material. For the virus to assemble, many copies of the nucleocapsid protein need to attach to each other, but the ISG15 tag prevents this, which is the mechanism behind the tag’s antiviral action. “However, the novel coronavirus also has an enzyme that can remove the tags from its nucleocapsid, recovering its ability to assemble new viruses and thus overcoming the innate immune response,” explains Shoji.

The novel coronavirus shares many traits with the SARS and MERS viruses, which all belong to the same family of viruses. And these viruses, too, have an enzyme that can remove the ISG15 tag. However, Shoji’s team found that their versions are less efficient at it than the one in the novel coronavirus. And in fact, it has been reported recently that the previous viruses’ enzymes have a different primary target. “These results suggest that the novel coronavirus is simply better at evading this aspect of the innate immune system’s defense mechanism, which explains why it is so infectious,” says Shoji.

But understanding just why the novel coronavirus is so effective also points the way to developing more effective treatments. The Kobe University researcher explains: “We may be able to develop new antiviral drugs if we can inhibit the function of the viral enzyme that removes the ISG15 tag. Future therapeutic strategies may also include antiviral agents that directly target the nucleocapsid protein, or a combination of these two approaches.”

This research was funded by the Kansai Economic Federation, the Hyogo Science and Technology Association (grant 3501) and the Ministry of Education, Culture, Sports, Science and Technology Japan (grant 18042-203556). It was conducted in collaboration with researchers from Universitas Gadjah Mada, Niigata University, the University of Yamanashi, Hokkaido University and Osaka University.

The term kokumi derives from Japanese and refers to a full-bodied and long-lasting taste experience. The taste impression is particularly pronounced in aged cheeses, mainly due to the increasing concentration of gamma-glutamyl dipeptides. These are small molecules that consist of a link between glutamic acid and another amino acid.

Depending on how the two amino acids are linked, researchers distinguish between gamma-, alpha-, and X-glutamyl dipeptides, with the latter two not contributing to the kokumi effect. The high polarity of the glutamyl dipeptides, as well as their great structural similarity with different flavor contributions, represent a major challenge for food analysis.

Efficient analysis method developed

Nevertheless, the team led by principal investigator Andreas Dunkel of the Leibniz Institute has succeeded in developing a new efficient analysis method based on ultra-high performance liquid chromatography-mass spectrometry. For the first time, it can precisely and selectively determine the concentrations of all 56 gamma-glutamyl dipeptide variants in just 22 minutes. Optimized sample preparation makes it possible to analyze 60 cheese samples per day.

“This is a significant improvement compared to other methods. Our tests have shown that our method is faster, more efficient, and yet reliable — it delivers reproducible results and detects even the smallest concentrations,” says first author Sonja Maria Fröhlich, a doctoral student at the Leibniz Institute. To further investigate the influence of ripening time on gamma-glutamyl dipeptide concentrations, the researchers applied the method to 122 cheese samples from Europe and the USA after the test phase. The ripening times of the cheese ranged from two weeks to 15 years.

Mold cultures accelerate flavor development

The results show that, as expected, the concentrations of glutamyl dipeptides increase with increasing ripeness. “Interestingly, the addition of blue and white mold cultures led to significantly higher gamma-glutamyl dipeptide concentrations, even at shorter ripening times,” says Andreas Dunkel, who heads the Integrative Food Systems Analysisresearch group at the Leibniz Institute.

The food chemist adds: “The concentration profiles we have determined for different stages of ripening and different types of cheese can be used in the future as a database for prediction models. The latter could, for example, be used to objectively monitor flavor development during cheese ripening, to shorten ripening times, or to develop new plant-based cheese products with high consumer acceptance.”

“In the sense of an interdisciplinary, food systems biology research approach, one of our goals is to combine analytical research results with bioinformatic methods to develop predictive models suitable to support sustainable food production. This is also the starting point of the project led by Andreas Dunkel,” concludes Veronika Somoza, director of the Leibniz Institute.

However, Van Overwalle’s research extends beyond motor functions, exploring the cerebellum’s involvement in social and cognitive abilities. His findings reveal that abnormalities in the cerebellum not only lead to motor deficits but are also linked to emotional and behavioral disorders. He references research on individuals with autism, demonstrating how non-invasive brain stimulation techniques like magnetic stimulation can improve social task performance.

“We’ve seen improvements in the sequence of cognitive tasks in people with autism through magnetic stimulation,” says Van Overwalle. “We’re now testing more complex tasks to see if these effects can be further enhanced, with the ultimate goal of developing practical treatments for people with autism.”

A notable breakthrough is the use of transcranial electrical stimulation (tES), a more affordable and accessible technique compared to magnetic stimulation. While the effects of tES are still limited, the research group is committed to further development, seeing its potential for wide-scale application in the future.

This research offers a fresh perspective on the cerebellum’s role and paves the way for new treatments for psychiatric and neurological conditions, such as autism spectrum disorders. “Our hope is to refine these techniques further to improve social and cognitive functions in people with autism,” concludes Van Overwalle.