Understanding the disease mechanisms of cerebellar ataxia through induced pluripotent stem cell models

The cerebellar ataxias are a heterogeneous group of incurable disorders that are characterised by progressive dysfunction and degeneration of Purkinje cells. Although numerous causative genes have been identified, the underlying pathogenic mechanisms remain incompletely understood, due to the limited availability of human brain tissue and the lack of reliable disease models. Accumulating evidence has suggested that subtypes of autosomal dominant ataxias (SCAs) are likely to be caused by shared pathogenic pathways. Focused on spinocerebellar ataxia type 14 (SCA14), a subtype of SCAs, this work aimed to understand the aetiology of SCA14 and related SCAs using novel disease models derived from human induced pluripotent stem cells (iPSCs). SCA14 is caused by mutations in protein kinase C gamma (PKCγ), a central molecule in signalling pathways that are important for Purkinje cell development and function. Using iPSCs derived from patients carrying two distinct SCA14 mutations (H36R and H101Q), this work has revealed three novel interconnected disease mechanisms. (1) SCA14 mutations promoted formation of PKCγ foci, which accumulate in the cytoplasm. (2) Mutant PKCγ showed increased kinase activity resulting in increased phosphorylation of substrates. (3) The mislocalisation of hyperactive mutant PKCγ is likely to affect the phosphorylation of its targets at the membrane and the cytoplasm. Importantly, results obtained in SCA14 iPSCs faithfully recapitulated the pathological findings observed in the patient post-mortem cerebellum. Finally, to determine the developmental changes in Purkinje cells, human Purkinje cells were generated from control and SCA14 iPSCs using a novel simplified protocol. Notably, the dendritic development and survival of human SCA14 Purkinje cells were significantly compromised in long-term culture. CRISPR/Cas9-mediated genome editing was also employed to correct the mutations and produce isogenic controls of SCA14 iPSCs. Together, this work highlighted the importance of a faithful disease model and the value of a patient material-driven approach for understanding the aetiology of neurological disorders. In the future, these patient iPSCs would be an ideal platform for further investigation of molecular pathways and therapeutic development for SCA14 and other cerebellar ataxias.