Curation SCA4 ZFHX3

Five Swedish SCA4 probands/families with cerebellar ataxia, sensory neuropathy, and uninterrupted exonic GGC repeat expansions in ZFHX3; expansions co-segregated in the families studied and were absent from large in-house/SweGen control datasets.

Expanded ZFHX3 GGC allele size correlated inversely with age at onset and supported anticipation; pathogenic alleles in the German cohort ranged from 44 to 68 repeats, with longer expansions associated with earlier onset and more severe phenotype.

Locus-level segregation evidence: a five-generation northern German SCA4 family showed linkage to chromosome 16q22, with affected members sharing the disease haplotype and a maximum two-point LOD score of 4.48 at D16S3018. This predates ZFHX3 GGC discovery and supports the SCA4 interval rather than directly genotyping the repeat expansion.

Postmortem SCA4 brain tissue showed neuronal intranuclear inclusions positive for ZFHX3, p62, and ubiquitin in basis pontis neurons, supporting aggregation/association with proteostasis markers; direct biochemical protein interaction was not separately demonstrated.

Regulatory/epigenetic effects are supported across available sources: ZFHX3 expression in nervous-system tissues was evaluated; SCA4 patient fibroblasts/iPSCs showed increased ZFHX3 protein without increased mRNA; and long-read methylation analyses in Swedish/Utah/Iowa and Chilean carriers showed allele-specific hypermethylation of the repeat-containing haplotype.

Patient-derived fibroblasts and induced pluripotent stem cells carrying the ZFHX3 GGC expansion showed increased ZFHX3 protein abundance and increased autophagy markers including p-mTOR, mTOR, p62, and LC3-II, consistent with impaired autophagy.

Cultured patient-derived fibroblast and iPSC models showed abnormal autophagy that was normalized after siRNA-mediated ZFHX3 knockdown, supporting a cell-culture disease model and a ZFHX3-dependent effect.