Transcript-Specific Loss-of-Function Variants in VPS16 Are Enriched in Patients With Dystonia

Abstract

Background and objectives: Our objective was to improve rare variant interpretation using statistical measures as well as publicly accessible annotation of expression levels and tissue specificity of different splice isoforms. We describe rare VPS16 variants observed in patients with dystonia and patients without dystonia, elaborate on our interpretation of VPS16 variants affecting different transcripts, and provide detailed clinical description of the movement disorder caused by VPS16 variants.

Methods: In-house exome and genome data sets (n = 11,539) were screened for rare heterozygous missense and putative loss-of-function (pLoF) variants in VPS16. Using pext (proportion expressed across transcripts) values from the Genome Aggregation Database (gnomAD), we differentiated variants affecting weakly and highly expressed exons/transcripts and applied statistical measures to systematically identify disease-associated genetic variation among patients with dystonia (n = 280).

Results: Six different heterozygous pLoFs in VPS16 transcripts were identified in 13 individuals. Three of these pLoFs occurred in 9 individuals with different phenotypes, and 3 pLoFs were identified in 4 unrelated individuals with early-onset dystonia. Although pLoFs were enriched in the dystonia cohort (n = 280; p = 2.04 × 10^-4; 4/280 cases vs 9/11,259 controls; Fisher exact test), it was not exome-wide significant. According to the pext values in gnomAD, all 3 pLoFs observed in the patients with dystonia were located in the highly expressed canonical transcript ENST00000380445.3, whereas 2 of 3 pLoFs detected in 8 individuals without dystonia were located in the first exon of the noncanonical transcript ENST00000380443.3 that is weakly expressed across all tissues. Taking these biological implications into account, pLoFs involving the canonical transcript were exome-wide significantly enriched in patients with dystonia (p = 1.67 × 10^-6; 4/280 cases vs 1/11,259 controls; Fisher exact test). All VPS16 patients showed mild progressive dystonia with writer's cramp as the presenting symptom between age 7 and 34 years (mean 20 years) that often progressed to generalized dystonia and was even accompanied by hyperkinetic movements and myoclonus in 1 patient.

Discussion: Our data provide strong evidence for VPS16 pLoFs to be implicated in dystonia and knowledge on exon resolution expression levels as well as statistical measures proved to be useful for variant interpretation.

Figure 1. Graphical Illustration of VPS16 Gene Structure

(A) Graphical illustration of VPS16 demonstrates the positions of the identified variants (red lines) and previously published variants in VPS16 (black lines). (B) Different transcripts of VPS16. The Ensembl canonical transcript is marked with an asterisk. (C) Approximate visualization of mean pext (proportion expressed across transcripts) value of VPS16 using GTEx v7²² transcriptomic sequencing data sets incorporated in gnomAD browser v2.1.1 illustrates normalized value of exon expression levels of all transcripts across all tissues. All exons of the canonical transcript ENST00000380445.3 show similar expression values, whereas the exon 1 of transcript ENST00000380443.3 shows a notably weaker expression. This exon partly overlaps with the exons 12 and 13 of the canonical transcript. Eight pLoFs were identified in the control group in this exon region that did not overlap with the exons 12 and 13 of the canonical transcript (red arrows).

Figure 2. Clinical Images of the Patients With Dystonia

Patient 1 exhibited myoclonic jerks and hyperkinetic movements of the right arm during upper limb posturing (A.a-c) and writer's cramp (A.d). Patient 2 had cervical dystonia (B.a), mild truncal dystonia, and scoliosis (B.b). She also showed hand dystonia and writer's cramp that were more prominent on the right hand (B.c) than on the left (B.d). Patient 3 had cervical dystonia with prominent retro- and torticollis (C.a-b). Patient 4 had cervical dystonia and dystonic postural tremor of the upper limb (D.a-b). All images are snapshots from the supplementary videos (Videos 1–4).