The choreography of development is a delicate dance. Beginning in utero, chromosomes, DNA, genes and RNA twirl, tap, and sashay their way in a precise pattern. A misstep or a missing step that changes the routine causes body and brain functions to go awry – as is the case with many intellectual and developmental disabilities (IDD). Fragile X syndrome is the most common known single-gene cause of inherited IDDs, including autism. Scientists know the misstep in this syndrome is in the gene FMR1. FMR1 is responsible for making the protein FMRP, which is necessary for typical brain development.
Lynne Maquat, Ph.D., founding director of the Center for RNA Biology at the University of Rochester, and professor of Biochemistry & Biophysics, Oncology, and Pediatrics, did not set out to study fragile X. It was through another line of research – her seminal discovery of and decades’ worth of work on nonsense-mediated mRNA decay (NMD) – that fragile X syndrome entered her radar. NMD is a cellular quality-control mechanism that plays a role in both healthy and disease states, and her lab discovered that it is overactive in people with fragile X.
“It was complete serendipity,” Maquat said. “No one ever thought to look at NMD and fragile X. So now we’re trying to figure out what happens at the molecular level when FMRP is absent; we want to understand the network of altered gene expression by identifying mis-regulated messenger RNAs (mRNAs).”
“It is key to finding any kind of hope for a future therapy.”
One of the most prominent surveillance systems in the body that protects against mistakes in gene expression that lead to disease, NMD is a complex pathway that is at the heart of many of the collaborations between Maquat and other University of Rochester scientists. Together, with funding from the National Institutes of Health (NIH) and the FRAXA Research Foundation, they aim to gain a deeper understanding of the sophisticated mechanisms related to NMD that will contribute to developing new drug therapies for genetic disorders such as fragile X syndrome, cystic fibrosis, and hundreds of others.
INTO THE BRAIN
Associate professor of Biomedical Genetics Christoph Pröschel, Ph.D., has spent much of his career interested in neurogenetic diseases that primarily affect the white matter of the brain, which carries signals throughout the organ. His lab started working with induced pluripotent stem cells (iPSCs) to understand different neural cell types, providing a solid foundation for their IDD research. “My lab and the Maquat lab have a mutual interest in the molecular mechanism of fragile X,” said Proschel. “It is key to finding any kind of hope for a future therapy.”
The Pröschel lab makes and differentiates neural stem cells that mimic fragile X syndrome, allowing his team to test hypotheses and understand how different therapies impact cell biology and function. He and Tatsuaki Kurosaki, Ph.D., research assistant professor in the Maquat lab, used these neural stem cells to understand the relationship between FMRP and NMD. They discovered that NMD controls the amounts of messenger RNAs deriving from a wide range of genes throughout the brain, including genes that govern motor control and cognitive processes related to attention, learning, and language. They also found that when FMRP is absent from cells, as it is in people with fragile X syndrome, NMD shifts into overdrive.
This work was part of a 2021 study published in Nature Cell Biology led by Maquat that revealed that tamping down NMD with small molecule inhibitors restored a large proportion of neurological functions in these cells.
Most recently, Pröschel co-authored research published in Molecular Cell led by Maquat and co-authored by Hana Cho, Ph.D., and Elizabeth Abshire, Ph.D., of her lab. The study highlighted a complex molecular dance between NMD and the enzyme AKT, which plays a key role in cell growth and survival. Both AKT and NMD are overactive in fragile X. Using neural stem cells that lack the FMRP protein, they tested a drug called Afuresertib, which inhibits AKT. They discovered that blocking AKT in the fragile X cells decreased its activity and decreased NMD. These cells then acted more like typical, non-disease cells.
There is still a lot the team doesn’t know about how AKT and NMD interact, because they both influence and regulate multiple activities in cells, but this work provides good direction that could inform the development of future treatments for fragile X syndrome.
“This has been one of the real fun chapters of my career – working with this group,” said Pröschel. “Everyone brings such a different perspective to the project.”
FROM SURGERY TO THE LAB
As a neurotologist (subspecialist of Otolaryngology), Hitomi Sakano, M.D., Ph.D., spends time in the clinic with patients with hearing issues or hearing loss. In the lab, she aims to understand how the brain adapts to sound information.
Her work with fragile X syndrome began as a resident at the University of Washington when she took interest in FMRP, which is highly expressed in the auditory brainstem nuclei of a typical brain and is the same protein missing in fragile X patients. When Sakano came to the Medical Center in 2018, she brought the fragile X mouse model to study this and joined the Center for RNA Biology.
“I also use the [knockout] mouse model to study hyperacusis – extreme sensitivity to sound,” said Sakano. “We know that fragile X patients have sensory and auditory sensitivity, so this model is a great tool to study both.” Hyperacusis is also very common in the general population (some report up to 15 percent) so understanding the mechanism could potentially impact our broader community.
Sakano hypothesizes that FMRP regulates genes that enable neuroplasticity to maintain
From left: Hitomi Sakano, M.D., Ph.D., and Lynne Maquat, Ph.D.
normal processing of auditory information. If true, there may be therapeutic targets for symptoms like auditory hypersensitivity in fragile X. Funding from the Schmitt Program in Integrative Neuroscience (SPIN) through the Del Monte Institute for Neuroscience Pilot Program and a NIH Research Career Development Award for clinician-scientists are supporting her research, which involves investigating the gene expression abnormalities in the auditory brainstem of the fragile X mouse model that might explain the auditory hypersensitivity in these mice. To date, she has found some interesting RNAs that encode synaptic proteins. These findings open up the possibility of targeting these genes for the treatment of hyperacusis.
In 2020, the National Institute of Child Health and Human Development (NICHD) designated the University of Rochester as an Intellectual and Developmental Disabilities Research Center. This recognition acknowledged the Medical Center’s national leadership in research
for conditions such as autism, Batten disease, Rett syndrome, and most recently fragile X syndrome. The Center’s researchers, including Maquat, Pröschel, Sakano, Telias, and Brima, work to translate scientific insights into new ways to diagnose and treat these conditions.
She co-authored a study with the Maquat and Pröschel labs in Genome Biology. The research used the mouse model whose FMR1 gene is knocked-out. These findings build upon Maquat’s previous research that showed NMD hyperactivation in neuronally induced stem cells from fragile X patients. This hyperactivity negatively impacts many neuronal mRNAs important to brain development. The Genome Biology paper showed NMD goes into overdrive in the brain during early development in a mouse with fragile X. These researchers are now testing various therapeutics to inhibit NMD.
“Being able to collaborate to gain meaningful results to move this science forward is the value of being at an academic medical center like Rochester,” said Sakano. “These steps are what will ultimately lead to treatments and therapies that I use in the clinic someday to help my patients.”
ON THE HORIZON
Forthcoming research aims to broaden the scope of the fragile X work at the Medical Center. One of the world’s largest clinics for fragile X is in Israel, where an estimated 80 percent of women are screened for the inherited disease. Michael Telias, Ph.D., assistant professor of Ophthalmology, Neuroscience, and Center for Visual Science, began studying fragile X as a graduate student in Israel. He uses human embryonic stem cells that carry the mutation for fragile X to look inside neurons at the molecular and cellular levels to shed light on the human-specific mechanisms affected by this syndrome.
From left: Christoph Pröschel, Ph.D., and Michael Telias, Ph.D.
“Human neurons have shown us that these cells have a problem receiving information and communicating information to the next cell,” said Telias. “We cannot do this work in humans, so using human cells enables us to know what to target in the cell. That is the only way we will be able to develop treatments that work.”
In the Frederick J. and Marion A. Schindler Cognitive Neurophysiology Laboratory, research assistant professor Tufikameni Brima, Ph.D., is aiming to use electroencephalography (EEG) and event-related potentials (ERP) to better understand how the brains of patients with fragile X respond to various stimuli. This work has the potential to build upon the ongoing molecular research being conducted by Telias and others.
“Ultimately, what we are figuring out is what happens when FMRP is absent. We don’t know the whole story,” Maquat said. “However, FMRP is an RNA-binding protein, and in work soon to be published in Molecular Cell, Kurosaki and I have now defined those messenger RNAs that are normally bound by FMRP and how the absence of FMRP binding results in those mRNAs making too much protein. These results have allowed us to identify which genes are affected and how. Our work will pave the way for better therapeutics for those living with fragile X.”