Combined genomics and proteomics unveils elusive variants and vast aetiologic heterogeneity in dystonia
Authors: Zech Michael , Dzinovic Ivana , Skorvanek Matej , Harrer Philip , Necpal Jan , Kopajtich Robert , Kittke Volker , Tilch Erik , Zhao Chen , Tsoma Eugenia , Sorrentino Ugo , Indelicato Elisabetta , Stehr Antonia , Saparov Alice , Abela Lucia , Adamovicova Miriam , Afenjar Alexandra , Assmann Birgit , Baloghova Janette , Baumann Matthias , Berutti Riccardo , Brezna Zuzana , Brugger Melanie , Brunet Theresa , Cogne Benjamin , Colangelo Isabel , Conboy Erin , Distelmaier Felix , Eckenweiler Matthias , Garavaglia Barbara , Geerlof Arie , Graf Elisabeth , Hackenberg Annette , Harvanova Denisa , Haslinger Bernhard , Havrankova Petra , Hoffmann Georg F , Janzarik Wibke G , Keren Boris , Kolnikova Miriam , Kolokotronis Konstantinos , Kosutzka Zuzana , Koy Anne , Krenn Martin , Krygier Magdalena , Kusikova Katarina , Maier Oliver , Meitinger Thomas , Mertes Christian , Milenkovic Ivan , Monfrini Edoardo , Mourao Andre Santos Dias , Musacchio Thomas , Nizon Mathilde , Ostrozovicova Miriam , Pavlov Martin , Prihodova Iva , Rektorova Irena , Romito Luigi M , Rybanska Barbora , Sadr-Nabavi Ariane , Schwenger Susanne , Shoeibi Ali , Sitzberger Alexandra , Smirnov Dmitrii , Svantnerova Jana , Tautanova Raushana , Toelle Sandra P , Ulmanova Olga , Vetrini Francesco , Vill Katharina , Wagner Matias , Weise David , Zorzi Giovanna , Di Fonzo Alessio , Oexle Konrad , Berweck Steffen , Mall Volker , Boesch Sylvia , Schormair Barbara , Prokisch Holger , Jech Robert , Winkelmann Juliane 
Published: February 2025

Dystonia is a rare-disease trait for which large-scale genomic investigations are still underrepresented. Genetic heterogeneity among patients with unexplained dystonia warrants interrogation of entire genome sequences, but this has not yet been systematically evaluated.

To significantly enhance our understanding of the genetic contribution to dystonia, we (re)analyzed 2,874 whole-exome sequencing (WES), 564 whole-genome sequencing (WGS), as well as 80 fibroblast-derived proteomics datasets, representing the output of high-throughput analyses in 1,990 patients and 973 unaffected relatives from 1,877 families. Recruitment and precision-phenotyping procedures were driven by long-term collaborations of international experts with access to overlooked populations.

By exploring WES data, we found that continuous scaling of sample sizes resulted in steady gains in the number of associated disease genes without plateauing. On average, every second diagnosis involved a gene not previously implicated in our cohort. Second-line WGS focused on a subcohort of undiagnosed individuals with high likelihood of having monogenic forms of dystonia, comprising large proportions of patients with early onset (81.3%), generalized symptom distribution (50.8%) and/or coexisting features (68.9%). We undertook extensive searches for variants in nuclear and mitochondrial genomes to uncover 38 (ultra)rare diagnostic-grade findings in 37 of 305 index patients (12.1%), many of which had remained undetected due to methodological inferiority of WES or pipeline limitations. WGS-identified elusive variations included alterations in exons poorly covered by WES, RNA-gene variants, mitochondrial-DNA mutations, small copy-number variants, complex rearranged genome structure, and short tandem repeats. For improved variant interpretation in WGS-inconclusive cases, we employed systematic integration of quantitative proteomics. This aided in verifying diagnoses related to technically challenging variants and in upgrading a variant of uncertain significance (3 of 70 WGS-inconclusive index patients, 4.3%). Further, unsupervised proteomic outlier-analysis supplemented with transcriptome sequencing revealed pathological gene underexpression induced by transcript disruptions in three more index patients with underlying (deep) intronic variants (3/70, 4.3%), highlighting the potential for targeted antisense-oligonucleotide therapy development. Finally, trio-WGS prioritized a de-novo missense change in the candidate PRMT1, encoding a histone-methyltransferase. Data-sharing strategies supported the discovery of three distinct PRMT1 de-novo variants in four phenotypically similar patients, associated with loss-of-function effects in in-vitro assays.

This work underscores the importance of continually expanding sequencing cohorts to characterize the extensive spectrum of gene aberrations in dystonia. We show that a pool of unresolved cases is amenable to WGS and complementary multi-omic studies, directing advanced etiopathological concepts and future diagnostic-practice workflows for dystonia.