Particle-mediated delivery of frataxin plasmid to a human sensory neuronal model of Friedreich's ataxia.

Increasing frataxin protein levels through gene therapy is envisaged to improve therapeutic outcomes for patients with Friedreich's ataxia (FRDA). A non-viral strategy that uses submicrometer-sized multilayered particles to deliver frataxin-encoding plasmid DNA affords up to 27 000-fold increase in frataxin gene expression within 2 days in vitro in a stem cell-derived neuronal model of FRDA

ALS-linked CCNF variant disrupts motor neuron ubiquitin homeostasis

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are fatal neurodegenerative disorders that share pathological features, including the aberrant accumulation of ubiquitinated protein inclusions within motor neurons. Previously, we have shown that the sequestration of ubiquitin (Ub) into inclusions disrupts Ub homeostasis in cells expressing ALS-associated variants superoxide dismutase 1 (SOD1), fused in sarcoma (FUS) and TAR DNA-binding protein 43 (TDP-43). Here, we investigated whether an ALS/FTD-linked pathogenic variant in the CCNF gene, encoding the E3 Ub ligase Cyclin F (CCNF), also perturbs Ub homeostasis. The presence of a pathogenic CCNF variant was shown to cause ubiquitin-proteasome system (UPS) dysfunction in induced pluripotent stem cell-derived motor neurons harboring the CCNF S621G mutation. The expression of the CCNFS621G variant was associated with an increased abundance of ubiquitinated proteins and significant changes in the ubiquitination of key UPS components. To further investigate the mechanisms responsible for this UPS dysfunction, we overexpressed CCNF in NSC-34 cells and found that the overexpression of both wild-type (WT) and the pathogenic variant of CCNF (CCNFS621G) altered free Ub levels. Furthermore, double mutants designed to decrease the ability of CCNF to form an active E3 Ub ligase complex significantly improved UPS function in cells expressing both CCNFWT and the CCNFS621G variant and were associated with increased levels of free monomeric Ub. Collectively, these results suggest that alterations to the ligase activity of the CCNF complex and the subsequent disruption to Ub homeostasis play an important role in the pathogenesis of CCNF-associated ALS/FTD