Elucidating Hexanucleotide Repeat Number and Methylation within the X-Linked Dystonia-Parkinsonism (XDP)-Related SVA Retrotransposon in TAF1 with Nanopore Sequencing

Author:

Lüth Theresa,Laβ Joshua,Schaake Susen,Wohlers InkenORCID,Pozojevic Jelena,Jamora Roland Dominic G.ORCID,Rosales Raymond L.ORCID,Brüggemann Norbert,Saranza Gerard,Diesta Cid Czarina E.,Schlüter Kathleen,Tse Ronnie,Reyes Charles Jourdan,Brand Max,Busch HaukeORCID,Klein Christine,Westenberger Ana,Trinh Joanne

Abstract

Background: X-linked dystonia-parkinsonism (XDP) is an adult-onset neurodegenerative disorder characterized by progressive dystonia and parkinsonism. It is caused by a SINE-VNTR-Alu (SVA) retrotransposon insertion in the TAF1 gene with a polymorphic (CCCTCT)n domain that acts as a genetic modifier of disease onset and expressivity. Methods: Herein, we used Nanopore sequencing to investigate SVA genetic variability and methylation. We used blood-derived DNA from 96 XDP patients for amplicon-based deep Nanopore sequencing and validated it with fragment analysis which was performed using fluorescence-based PCR. To detect methylation from blood- and brain-derived DNA, we used a Cas9-targeted approach. Results: High concordance was observed for hexanucleotide repeat numbers detected with Nanopore sequencing and fragment analysis. Within the SVA locus, there was no difference in genetic variability other than variations of the repeat motif between patients. We detected high CpG methylation frequency (MF) of the SVA and flanking regions (mean MF = 0.94, SD = ±0.12). Our preliminary results suggest only subtle differences between the XDP patient and the control in predicted enhancer sites directly flanking the SVA locus. Conclusions: Nanopore sequencing can reliably detect SVA hexanucleotide repeat numbers, methylation and, lastly, variation in the repeat motif.

Funder

Deutsche Forschungsgemeinschaft

Hermann and Lilly Schilling Foundation

European Community

Canadian Institutes of Health Research

Peter and Traudl Engelhorn Foundation

Publisher

MDPI AG

Subject

Genetics (clinical),Genetics

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