High-resolution genomic architecture of a 1.71-Mb inverted SNCA triplication in a patient-derived PARK4 model

PARK4, caused by SNCA locus multiplications, is a severe, early-onset form of autosomal dominant parkinsonism. However, the precise structural topography of these massive genomic rearrangements remains inadequately resolved. In this study, we resolved the nucleotide-level architecture of a novel 1.71-Mb PARK4 triplication lineage (PARK4-4) and elucidated its early-stage biochemical consequences in a patient-derived cortical model. By integrating Optical Genome Mapping (OGM) and Oxford Nanopore Technologies (ONT) long-read sequencing, we mapped a complex 1.71-Mb inverted triplication spanning the HERC6 to CCSER1 loci. This rearrangement is organized in a Forward-Inverted-Forward (F-I-F) architecture, punctuated by precisely defined Head-to-Head (H2H) and Tail-to-Tail (T2T) junctions. Using patient-derived induced pluripotent stem cells (iPSCs) differentiated into forebrain cortical neurons, targeted RT-qPCR and automated capillary western blotting demonstrated SNCA gene-dosage-associated α-synuclein overexpression, with a robust ∼2.3-fold increase. Furthermore, exploratory transcriptomic profiling (RNA-seq) revealed candidate synchronized upregulation of co-triplicated neighboring genes (PIGY, FAM13A, and TIGD2), supporting a regional gene-dosage effect. To our knowledge, this is the first structural characterization of a 1.71-Mb F-I-F inverted SNCA triplication at nucleotide resolution, revealing a complex architecture distinct from previously reported tandem triplications. While future isogenic and functional studies are required to establish mature disease-relevant pathology, this precisely mapped model provides an invaluable platform for studying dosage-dependent early biochemical phenotypes and evaluating targeted therapeutics.