Integrating Long‐Read Nanopore Sequencing for Precision Resolution of Genomic Variants in Dystonia

Ugo Sorrentino, Martin Pavlov, Nazanin Mirza‐Schreiber, Melanie Brügger, Theresa Brunet, Eugenia Tsoma, Alice Saparov, Ivana Dzinovic, Philip Harrer, Antonia Margarethe Stehr

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Matias Wagner, Erik Tilch, Barbara Wallacher, Shiraz Alhasan, Anne Koy, Alessio Di Fonzo, Miriam Kolníková, Katarína Kušíková, Petra Havránková, Raushana Tautanova, Sandy Lösecke, Sebastian H. Eck, Sylvia Boesch, Ján Necpál, Matěj Škorvánek, Robert Jech, Holger Prokisch, Juliane Winkelmann, Konrad Oexle, Elisabeth Graf, Michael Zech

2025

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DOI 10.1002/mds.70072
Data confidence 70 medium
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Abstract

BACKGROUND: Although many individuals with dystonia present with features indicative of single-gene etiologies, obtaining definitive genetic diagnoses can be challenging. OBJECTIVE: We assessed the value of nanopore-based long-read sequencing (LRS) in achieving molecular clarification of dystonic syndromes. METHODS: From a large dystonia cohort with short-read sequencing (SRS) data, 14 cases with unclear, difficult-to-evaluate, or missing causative variants were recruited. Long-read whole-genome sequencing was performed according to Oxford Nanopore Technologies (ONT) protocols. RESULTS: ONT sequencing produced long-range haplotypes, variant calls inaccessible to short-read technology, as well as methylation data. Phase inference allowed for changes in variant classification, establishing compound heterozygosity of causative variants in four cases. We illustrate an important advantage of LRS compared with SRS in (re)defining the identity of dystonia-causing structural variants and repeat expansions for seven individuals. One patient was found to harbor a novel exonic LINE-1 insertion in SGCE, expanding the genetic mechanism in myoclonus-dystonia. ONT data also provided unexpected insights into apparent mosaic expanded repeats in FMR1 in a subject with isolated focal dystonia. We further showed that LRS outperformed SRS in avoiding erroneous calls resulting from confounding pseudogene sequences and in discovering pathogenic alterations missed by conventional pipeline utilization (three cases). Moreover, simultaneous methylome analysis aided in directing the interpretation of three variants, including a KMT2B variant of uncertain significance that was reclassified as causal by LRS-based episignature profiling. CONCLUSIONS: ONT-based LRS uniquely improves analysis of dystonia-associated variations that had not previously been resolved by SRS, implying broad utility for future exploration of the molecular origins of the condition. © 2025 The Author(s). Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.