The findings will improve clinical diagnostic services for patients with neurodevelopmental disorders.

Through rigorous genetic analysis, the researchers discovered that mutations in a small non-coding gene called RNU4-2 cause a collection of developmental symptoms that had not previously been tied to a distinct genetic disorder. Non-coding genes are parts of DNA that do not produce proteins. The investigators used whole-genome sequencing data in the United Kingdom’s National Genomic Research Library to compare the burden of rare genetic variants in 41,132 non-coding genes between 5,529 unrelated cases with intellectual disability and 46,401 unrelated controls.

The discovery is significant, as it represents one of the most common single-gene genetic causes of such disorders, ranking second only to Rett syndrome among patients sequenced by the United Kingdom’s Genomic Medicine Service. Notably, these mutations are typically spontaneous and not inherited, providing important insights into the nature of the condition.

“We performed a large genetic association analysis to identify rare variants in non-coding genes that might be responsible for neurodevelopmental disorders,” says the study’s first author Daniel Greene, PhD, Assistant Professor of Genetics and Genomics Sciences at Icahn Mount Sinai and a Visitor at the University of Cambridge. “Nowadays, finding a single gene that harbors genetic variants responsible for tens of thousands of patients with a rare disease is exceptionally unusual. Our discovery eluded researchers for years due to various sequencing and analytical challenges.”

More than 99 percent of genes known to harbor mutations that cause neurodevelopmental disorders encode proteins. The researchers hypothesized that non-coding genes, which don’t produce proteins, could also host mutations leading to intellectual disability. Neurodevelopmental disorders, which often appear before grade school, involve developmental deficits affecting personal, social, academic, or occupational functioning. Intellectual disability specifically includes significant limitations in intellectual functioning (e.g., learning, reasoning, problem-solving) and adaptive behavior (e.g., social and practical skills).

“The genetic changes we found affect a very short gene, only 141 units long, but this gene plays a crucial role in a basic biological function of cells, called gene splicing, which is present in all animals, plants and fungi,” says senior study author Ernest Turro, PhD, Associate Professor of Genetics and Genomic Sciences at Icahn Mount Sinai and a Visitor at the University of Cambridge. “Most people with a neurodevelopmental disorder do not receive a molecular diagnosis following genetic testing. Thanks to this study, tens of thousands of families will now be able to obtain a molecular diagnosis for their affected family members, bringing many diagnostic odysseys to a close.”

Next, the researchers plan to explore the molecular mechanisms underlying this syndrome experimentally. This deeper understanding aims to provide biological insights that could one day lead to targeted interventions.

“What I found remarkable is how such a common cause of a neurodevelopmental disorder has been missed in the field because we’ve been focusing on coding genes,” says Heather Mefford, MD, PhD, of the Center for Pediatric Neurological Disease Research at St. Jude Children’s Research Hospital who was not involved with the research. “This study’s discovery of mutations in non-coding genes, especially RNU4-2, highlights a significant and previously overlooked cause. It underscores the need to look beyond coding regions, which could reveal many other genetic causes, opening new diagnostic possibilities and research opportunities.”

The paper is titled “Mutations in the U4 snRNA gene RNU4-2 cause one of the most prevalent monogenic neurodevelopmental disorders.”

The remaining authors of the paper are Chantal Thys (KU Leuven, Belgium); Ian R. Berry, MD (University of Bristol, UK); Joanna Jarvis, MD (Birmingham Womens’ Hospital, UK); Els Ortibus, MD, PhD (KU Leuven, Belgium); Andrew D. Mumford, MD (University of Bristol, UK); and Kathleen Freson, PhD (KU Leuven, Belgium).

The work was supported, in part, by NIH awards R01HL161365 and R03HD111492.

The virus, called Apocryptovirus odysseus, along with 18 others that are closely related to it, was discovered through a computational screen of human neuron data — an effort aimed at elucidating the connection between RNA viruses and neuroinflammatory disease. The virus is associated with severe inflammation in humans infected with the parasite Toxoplasma gondii, leading the team to hypothesize that it exacerbates toxoplasmosis disease.

“We discovered A. odysseus in human neurons using the open-science Serratus platform to search through more than 150,000 RNA viruses” said Purav Gupta, first author on the study, recent high school graduate and current undergraduate student at U of T’s Donnelly Centre for Cellular and Biomolecular Research. “Serratus identifies RNA viruses from public data by flagging an enzyme called RNA-dependent RNA polymerase, which facilitates replication of viral RNA. This enzyme allows the virus to reproduce itself and for the infection to spread.”

The study was published recently in the journal Virus Evolution.

The parasite T. gondii is far-reaching, infecting an estimated one-third of the global population. It can live in any non-blood cell type, including neurons, forming cysts inside cells. The parasite is transmitted to nearby cells when the infected cell ruptures.

T. gondii infections often go unnoticed because they only lead to symptoms in rare cases. Regardless, toxoplasmosis merits investigation considering how widespread it is and the potential effects it may have on pregnant women and those who are immunocompromised, Gupta said.

“We believe the virus and parasite work hand-in-hand to cause disease in the human host, where the virus hides inside the parasite, like a soldier in a trojan horse, to gain entry to the human brain,” said Gupta. “Our research marks the first time that scientists have connected toxoplasmosis to a virus.”

The newly discovered A. odysseus is found in two hypervirulent strains of the T. gondii parasite, referred to as RUB and COUGAR.

RUB has been documented in French Guinea to cause severe fever and organ failure, while COUGAR has been shown in British Columbia to be connected to ocular toxoplasmosis — the leading cause of infectious blindness. Researchers found the strains in different geographical locations at different times, demonstrating their potentially wide-ranging impacts.

Symptoms of toxoplasmosis can be aggravated by a hyperactivated human immune response. The virus-carrying parasite triggers this type of response when the immune system senses the foreign RNA of the virus.

“The group of 19 RNA viruses we found are strong biomarkers for parasitic infection,” said Artem Babaian, principal investigator on the study and assistant professor of molecular genetics at the Donnelly Centre and the Temerty Faculty of Medicine. “It’s obvious now that the A. odysseus virus could be a valuable marker of disease-causing infections, like severe toxoplasmosis, in humans or other animals. The next step is to test if this raises the possibility that treating a parasite’s viruses could be an effective means of treating symptoms that arise from parasitic infections.”

Zoonotic viruses that infect other living things in our environment in order to reach us are expected to cause the majority of emerging infectious diseases in humans, Babaian noted. “This study underscores the importance of looking beyond the viruses that infect humans directly into the extended virome,” he said.

This research was supported by the Canadian Institutes of Health Research.

Additionally, the brain’s response to serotonin stimulation is strongly linked to the distribution of serotonin receptors (proteins activated by serotonin) and the connection patterns of DRN serotonin neurons. “We clearly see from the high-field MRI images which areas in the brain are activated and deactivated during the awake state and under anesthesia when we activate serotonin neurons in the DRN,” lead author Dr. Hiroaki Hamada said. “A previous study showed that the cerebral cortex and the basal ganglia were mostly deactivated under anesthesia, which we also observed, however, in awake states these areas are significantly activated.”

Our brains are made of tens of billions of nerve cells called neurons. These cells communicate with each other through biomolecules called neurotransmitters. Serotonin, a type of neurotransmitter, is produced by serotonin neurons in our brains and influences many of our behavioral and cognitive functions such as memory, sleep, and mood.

“Learning about the brain’s serotonin system can help us understand how we adapt our behaviors and how mood therapy medication works. But it was hard to study how serotonin from the DRN affects the entire brain. First, because electric stimulation of the DRN can also activate neurons that don’t use serotonin to communicate with each other, and second, using drugs can affect other serotonin in the brain,” explained Dr. Hiroaki Hamada, a former PhD student at OIST’s Neural Computation Unit and lead author of a paper on this study published in the journal Nature Communications.

Previous studies by researchers at the Neural Computation Unit have shown that serotonin neurons in the DRN promote adaptive behaviors in mice associated with future rewards. Dr. Hamada and his collaborators wanted to understand the mechanisms in the brain that cause these adaptive behaviors.

“We knew that DRN serotonin activation has strong effects on behavior, but we didn’t know how this serotonin activation affects different parts of the brain,” Prof. Kenji Doya, leader of the Neural Computation Unit, stated.

Observing the entire brain’s response to DRN serotonin activation

The researchers used a novel technique called opto-functional MRI to address this question. They used a method called optogenetics to selectively activate serotonin neurons in the DRN with light and observed the entire brain’s response using functional MRI (Magnetic Resonance Imaging). They utilized the latest MRI scanner with a strong magnetic field to achieve the high resolution needed to study the small brains of mice. The mice were put in the MRI scanner and serotonin neurons were stimulated at regular intervals to see how this affected the whole brain.

They found that DRN serotonin stimulation causes activation of the cerebral cortex and the basal ganglia, brain areas involved in many cognitive functions. This result was very different from a previous study performed under anesthesia. Additionally, the brain’s response to serotonin stimulation is strongly linked to the distribution of serotonin receptors (proteins activated by serotonin) and the connection patterns of DRN serotonin neurons.

“We clearly see from the high-field MRI images which areas in the brain are activated and deactivated during the awake state and under anesthesia when we activate serotonin neurons in the DRN,” Dr. Hamada said. “A previous study showed that the cerebral cortex and the basal ganglia were mostly deactivated under anesthesia, which we also observed, however, in awake states these areas are significantly activated.”

The cerebral cortex and the basal ganglia are parts of the brain critical for many cognitive processes, including motor activity and behaviors to gain rewards such as food and water. Activation of DNR serotonin neurons can therefore lead to changes in motivation and behavior.

Patience and stimulating your own serotonin

Combining the new technique of high field MRI and optogenetics presented many obstacles that Dr. Hamada had to overcome. “We introduced and adapted a method previously used by our collaborators and established many new procedures at OIST. For me, the main challenge was using the new MRI machine at the time, so I needed to have patience and stimulate my own serotonin. I started doing a lot of exercise after that,” he laughed.

Seeing activations in the DRN for the first time was a standout moment for Dr. Hamada. In the beginning, he used the same light intensity that his collaborators used, but this was too weak to see the brain responses in the MRI. He then used bigger optical fibers and increased the intensity to stimulate the DRNs.

Prof. Doya noted that the next important milestone to achieve is understanding exactly how this brain-wide activation of serotonin occurs: “It’s important to find out what is the actual molecular mechanism allowing this activation in our brain. People who would like to get better at adjusting their behavior and thinking in different situations could also find it helpful to learn more about how serotonin helps control our moods.”

Today’s commercial charging infrastructure can be a jungle. The market is dynamic and complex with a variety of subscriptions and free competition between providers. At some fast charging stations, congestion and long queues may even occur. In a new study, researchers at Chalmers have created a mathematical model to investigate how fast charging stations controlled by artificial intelligence, AI, can help by offering electric car drivers personalised prices, which the drivers can choose to accept or refuse. The AI uses algorithms that can adjust prices based on individual factors, such as battery level and the car’s geographic location.

“The electric car drivers can choose to share information with the charging station providers and receive a personal price proposal from a smart charging station. In our study, we could show how rational and self-serving drivers react by only accepting offers that are beneficial to themselves. This leads to both price and waiting times being minimized,” says Balázs Kulcsár, professor at the department of electrical engineering at Chalmers.

In the study, the drivers always had the option to refuse the personal price, and choose a conventional charging station with a fixed price instead. The personal prices received by the drivers could differ significantly from each other, but were almost always lower than the market prices. For the providers of charging stations, the iterative AI algorithm can find out which individual prices are accepted by the buyer, and under which conditions. However, during the course of the study, the researchers noted that on some occasions the algorithm raised the price significantly when the electric car’s batteries were almost completely empty, and the driver consequently had no choice but to accept the offer.

“Smart charging stations can solve complex pricing in a competitive market, but our study shows that they need to be developed and introduced with privacy protection for consumers, well in line with responsible-ethical AI paradigms,” says Balázs Kulcsár.

More about the study

The researchers created a mathematical model of the interaction between profit-maximising fast charging stations and electric car users. The “charging stations” could offer public market prices or AI-driven profit-maximising personal prices, which the “electric car users” could then accept or reject based on their own conditions and needs. In most cases, the results were promising, as the AI-generated prices were lower than the market prices.

However, despite extensive efforts, regenerative therapies involving PSCs still have many hurdles to overcome. One being the formation of tumors (via the process of tumorigenesis) after the transplantation of PSCs. Once the PSCs differentiate into a specific type for stem cell therapy, there is a high probability of tumor formation after differentiated stem cells are introduced to the target organ. For the success of PSC-based therapies, the need of the hour is to minimize the risk of tumorigenesis by identifying potentially problematic cells in cultures, prior to transplantation.

Against this backdrop, a research team led by Atsushi Intoh and Akira Kurisaki from Nara Institute of Science and Technology, Japan, has recently achieved a breakthrough discovery regarding stem cell therapy and tumorigenesis. “Our findings present advancements that could bridge the gap between stem cell research and clinical application,” says Intoh, talking about the potential of their findings. Their study was published in Stem Cells Translational Medicine and focuses on a membrane protein called EPHA2, which was previously found to be elevated in PSCs prior to differentiation by the team.

Through several experiments involving both mouse and human stem cell cultures, the researchers gained insights into the role of EPHA2 in preserving the potency of PSCs to develop into several cell types. They found that EPHA2 in stem cells is co-expressed with OCT4 — a transcription factor protein which controls the expression of genes which are critically involved in the differentiation of embryonic stem cells. Interestingly, when the EPHA2 gene was knocked down from the cells, cultured stem cells spontaneously differentiated. These results suggest that EPHA2 plays a central role in keeping stem cells in an undifferentiated state.

The researchers thus theorized that EPHA2-expressing stem cells, which would fail to differentiate, might be responsible for tumorigenesis upon transplantation into the target organ.

To test this hypothesis, the researchers prepared PSC cultures and artificially induced their differentiation into liver cells. Using a magnetic antibody targeting EPHA2, they extracted EPHA2-positive cells from a group of cultures prior to transplantation into mice. Interestingly, the formation of tumors in mice receiving transplants from cultures from which EPHA2 had been removed was vastly suppressed.

Taken together, these results point to the importance of EPHA2 in emerging stem cell-based therapies. “EPHA2 conclusively emerges as a potential marker for selecting undifferentiated stem cells, providing a valuable method to decrease tumorigenesis risks after stem cell transplantation in regenerative treatments,” remarks Kurisaki.

Further in-depth studies on this protein may lead to the development of protocols that make PSCs safer to use. Luckily, however, these findings pave the way towards a future where we will be able to finally restore damaged organs and even overcome degenerative conditions.