Mapping Gene Variants Reveals New Neurodevelopmental Condition

How genetic variations cause disease, and the extent of their contributions to disease pathology, has historically been difficult to define. 

By mapping all the possible variations in a single gene, researchers have uncovered a previously hidden neurodevelopmental condition. 

RNU4-2 and ReNU syndrome  

ReNU syndrome is a rare, inherited neurodevelopmental disorder identified in 2024 that affects brain function, development, and motor skills and is predicted to affect tens of thousands of individuals worldwide.

 

Mutations to a non-protein-coding gene, RNU4-2, which codes for an RNA component of the spliceosome, were found to cause ReNU syndrome by Prof. Nicky Whiffin and colleagues. 

They identified an 18-base pair region of RNU4-2 that held several distinct mutations linked to ReNU syndrome, although UK Biobank data indicated that many other variants in RNU4-2 occur in people without ReNU syndrome. 

To understand the spectrum of deleterious mutations and potentially identify additional pathogenic variants, the researchers turned to functional genomics. 

Saturation genome editing 

A new method, called saturation genome editing (SGE), was developed by Dr. Greg Findlay and enables every single variant in a given gene to be mapped in order to identify its functional and clinical impact. 

“Saturation genome editing uses a molecular tool called CRISPR that lets us rewrite DNA in the human genome,” Findlay told Technology Networks. “This gives us a way to test genetic variants to see what effects they have on human cells in the lab.”

 

“What’s special about SGE is that we can test hundreds of variants per experiment, all at once,” he said. Human genes consist of hundreds to thousands of DNA bases, and SGE systematically tests variants at each base. By removing the need to select a few variants for study, SGE enables variant testing at scale and helps to interpret variants of uncertain significance. 

While SGE has been performed across numerous protein-coding genes, the work on RNU4-2 was the first time it had been used on a non-coding gene.

Understanding mutations to non-coding genes 

“Typically, a lot less is known about non-coding genes,” noted Findlay. “Because non-coding genes do not make proteins, it can be more difficult to predict how variants may impact their function.” 

Non-coding genes are also not frequently sequenced in diagnostic workflows like exome sequencing, meaning there is less information available about the mutations that occur in these genes in people. 

What made RNU4-2 a compelling case to test SGE on a non-coding gene? “A relatively large number of neurodevelopmental disease cases were attributable to a very small number of variants at the center of the gene,” explained Findlay. “We were curious whether variants elsewhere in the gene may also cause disease, especially as variants in RNU4-2 are relatively common in the general population.” 

Dr. Joachim De Jonghe, a former postdoc in Findlay’s team, installed over 500 different single-nucleotide RNU4-2 variants in human cells, using a slightly modified version of SGE. Variants were assigned an SGE function score based on how they impacted cell fitness.  

“We wanted to understand precisely which of the 539 variants we tested in RNU4-2 actually caused ReNU syndrome, and which were benign,” said Findlay. Understanding the genotype–phenotype relationship could also provide new insights about why certain variants cause more severe disease.

 

However, RNU4-2 presented some unique challenges for SGE. “There are many similar sequences to RNU4-2 in the human genome, so it was difficult for us to design CRISPR experiments to specifically manipulate only RNU4-2 and not any of the other genes,” explained Findlay. 

Promising but puzzling results 

The results of the SGE analysis of RNU4-2 “looked very promising but also a little puzzling,” said Findlay.  

The SGE function scores for variants in the 18-base pair region identified by Whiffin closely correlated with phenotypic severity, but Findlay and De Jonghe identified a further set of mutations outside of this region that also impaired gene function. 

“All variants known to cause ReNU syndrome that we tested had strong effects in the experiment, telling us the method was working, essentially,” explained Findlay. “But we also found many variants that had strong effects in new regions of RNU4-2 that hadn’t yet been linked to disease.”  

A recessive form of ReNU syndrome 

Whiffin coordinated a search of genetic databases of patients with unsolved neurodevelopmental disease cases across the globe to identify individuals with two variants to the RNU4-2 gene. 

“We quickly saw a pattern in individuals who had two variants with strong effects in the experiment, one on each copy of their RNU4-2 gene,” said Findlay. “All such individuals, about 20 in total, had an unexplained neurodevelopmental disorder.” 

“We knew we had discovered the recessive form of the disease.” — Dr. Greg Findlay 

Following the discovery of recessive ReNU syndrome, 38 individuals were diagnosed with the condition. 

While recessive ReNU syndrome shares some of the features of dominant ReNU syndrome, it impacts the white matter of the brain differently, which can be distinguished by magnetic resonance imaging. 

The researchers also found that RNU4-2 transcripts were reduced in recessive ReNU syndrome, suggesting a loss-of-function disease mechanism distinct from dominant ReNU syndrome. 

Impact on clinical genetics 

SGE data for RNU4-2 will be critical for diagnosing patients with currently unexplained neurodevelopmental disorders and potentially altering the course of their care. 

“For RNU4-2, like other genes, the SGE data will help clinical teams decide whether a rare variant in a patient may actually be causing disease,” explained Findlay. “Often, functional data is critical for tipping this assessment, providing enough certainty to make a formal diagnosis.” 

“Further, knowing the molecular cause of each patient’s disease provides a starting place for therapy development,” he concluded. 

 

Reference: De Jonghe J, Kim HC, Adedeji A, et al. Saturation editing of RNU4-2 reveals distinct dominant and recessive disorders. Nature. 2026. doi: 10.1038/s41586-026-10334-9 

 

About the interviewee: 

Dr. Greg Findlay leads the Genome Function Laboratory at the Francis Crick Institute. Findlay completed his PhD (2018) and MD (2020) at the University of Washington in Seattle. During this time, Findlay developed multiple CRISPR-based methods for functional genomics. In 2020, Findlay started his independent group at the Crick, where his lab leverages high-throughput methods to explore basic questions in human genetics.

The lab has systematically mapped variant effects across tumor suppressor genes such as BRCA1 and VHL using saturation genome editing, and more recently has developed prime editing strategies to investigate variants at scale. The lab’s overarching aim is to facilitate precision medicine approaches through improved understanding of human genetics.