Background and Objective: Friedreich ataxia (FRDA) is a life-shortening, neurodegenerative disorder caused by frataxin deficiency. FRDA patients’ neurological features include progressive ataxia and dysarthria, which are controlled to a large degree by the cerebellum. Cerebellar neuropathology in FRDA patients includes loss of large principal neurons and synaptic terminals in the cerebellar dentate nucleus as well as Purkinje neuronal injury; however, the mechanism by which frataxin deficiency impacts the cerebellum remains to be fully elucidated. In the present work, we utilized an inducible mouse model of FRDA (FRDAkd) to examine biochemical results of frataxin deficiency and a possible mechanism that leads to those changes.
Methods: 2-month-old FRDAkd mice were treated with inducing agent for 2 weeks, and behavior and weight were measured. Western blot and immunohistochemistry were conducted to measure the expression and localization of frataxin and AMPA(GluR2) receptors in the cerebellum. Cerebellar slides were also stained with Purkinje cell marker calbindin, and cells were counted. To determine if GluR2 was transcriptionally affected, RNA levels were measured using PCR. Analysis of GluR2 post-translational modifications (phosphorylation and palmitoylation) were conducted using antibodies and acyl biotin exchange assays.
Results: Frataxin and GluR2 expression are decreased to 50% by 2 weeks of induction, while Purkinje cell number is not changed. Immunostaining in wildtype cerebellar slides shows that GluR2 is enriched in Purkinje cell somatodendritic region while it is decreased in FRDAkd PCs. GluR2 RNA levels are not changed. GluR2 phosphorylation decreased, yet when normalized, phosphorylation is unchanged. Palmitoylation is disrupted in the FRDAkd cerebellum.
Discussion: These results indicate that frataxin knockdown leads to GluR2 palmitoylation disruption, suggesting the importance of frataxin for AMPA receptors trafficking in the cerebellum. Interestingly, at 2 weeks of induction FRDAkd mice do not present a behavioral phenotype, thus indicating early biochemical effects caused by frataxin knock down.