De novo variants in the non-coding spliceosomal snRNA gene RNU4-2 are a frequent cause of syndromic neurodevelopmental disorders.

Around 60% of individuals with neurodevelopmental disorders (NDD) remain undiagnosed after comprehensive genetic testing, primarily of protein-coding genes1. Increasingly, large genome-sequenced cohorts are improving our ability to discover new diagnoses in the non-coding genome. Here, we identify the non-coding RNA RNU4-2 as a novel syndromic NDD gene. RNU4-2 encodes the U4 small nuclear RNA (snRNA), which is a critical component of the U4/U6.U5 tri-snRNP complex of the major spliceosome2. We identify an 18 bp region of RNU4-2 mapping to two structural elements in the U4/U6 snRNA duplex (the T-loop and Stem III) that is severely depleted of variation in the general population, but in which we identify heterozygous variants in 119 individuals with NDD. The vast majority of individuals (77.3%) have the same highly recurrent single base-pair insertion (n.64_65insT). We estimate that variants in this region explain 0.41% of individuals with NDD. We demonstrate that RNU4-2 is highly expressed in the developing human brain, in contrast to its contiguous counterpart RNU4-1 and other U4 homologs, supporting RNU4-2's role as the primary U4 transcript in the brain. Overall, this work underscores the importance of non-coding genes in rare disorders. It will provide a diagnosis to thousands of individuals with NDD worldwide and pave the way for the development of effective treatments for these individuals.

Differences in 5'untranslated regions highlight the importance of translational regulation of dosage sensitive genes.

BACKGROUND: Untranslated regions (UTRs) are important mediators of post-transcriptional regulation. The length of UTRs and the composition of regulatory elements within them are known to vary substantially across genes, but little is known about the reasons for this variation in humans. Here, we set out to determine whether this variation, specifically in 5'UTRs, correlates with gene dosage sensitivity. RESULTS: We investigate 5'UTR length, the number of alternative transcription start sites, the potential for alternative splicing, the number and type of upstream open reading frames (uORFs) and the propensity of 5'UTRs to form secondary structures. We explore how these elements vary by gene tolerance to loss-of-function (LoF; using the LOEUF metric), and in genes where changes in dosage are known to cause disease. We show that LOEUF correlates with 5'UTR length and complexity. Genes that are most intolerant to LoF have longer 5'UTRs, greater TSS diversity, and more upstream regulatory elements than their LoF tolerant counterparts. We show that these differences are evident in disease gene-sets, but not in recessive developmental disorder genes where LoF of a single allele is tolerated. CONCLUSIONS: Our results confirm the importance of post-transcriptional regulation through 5'UTRs in tight regulation of mRNA and protein levels, particularly for genes where changes in dosage are deleterious and lead to disease. Finally, to support gene-based investigation we release a web-based browser tool, VuTR, that supports exploration of the composition of individual 5'UTRs and the impact of genetic variation within them.

Systematic identification of disease-causing promoter and untranslated region variants in 8,040 undiagnosed individuals with rare disease.

Background: Both promoters and untranslated regions (UTRs) have critical regulatory roles, yet variants in these regions are largely excluded from clinical genetic testing due to difficulty in interpreting pathogenicity. The extent to which these regions may harbour diagnoses for individuals with rare disease is currently unknown. Methods: We present a framework for the identification and annotation of potentially deleterious proximal promoter and UTR variants in known dominant disease genes. We use this framework to annotate de novo variants (DNVs) in 8,040 undiagnosed individuals in the Genomics England 100,000 genomes project, which were subject to strict region-based filtering, clinical review, and validation studies where possible. In addition, we performed region and variant annotation-based burden testing in 7,862 unrelated probands against matched unaffected controls. Results: We prioritised eleven DNVs and identified an additional variant overlapping one of the eleven. Ten of these twelve variants (82%) are in genes that are a strong match to the individual's phenotype and six had not previously been identified. Through burden testing, we did not observe a significant enrichment of potentially deleterious promoter and/or UTR variants in individuals with rare disease collectively across any of our region or variant annotations. Conclusions: Overall, we demonstrate the value of screening promoters and UTRs to uncover additional diagnoses for previously undiagnosed individuals with rare disease and provide a framework for doing so without dramatically increasing interpretation burden.