2020.06.12
Degeneration of dopaminergic neurons and impaired intracellular trafficking in Atp13a2 deficient zebrafish
(IBRO Rep. 2020 Jun 1;9:1-8. doi: 10.1016/j.ibror.2020.05.002. eCollection 2020 Dec.)
Nyuzuki Ha,c,d, Ito Sb, Nagasaki Ka, Nitta Yc, Matsui Nc,d, Saitoh Aa, Matsui Hc,d
aDepartment of Pediatrics, Niigata University Graduate School of Medical and Dental Sciences
bMedical Research Center, Graduate School of Medicine, Kyoto University
cDepartment of Neuroscience of Disease, Center for Transdisciplinary Research, Niigata University
dDepartment of Neuroscience of Disease, Brain Research Institute, Niigata University
Abstract
ATP13A2 is the autosomal recessive causative gene for juvenile-onset Parkinson's disease (PARK9, Parkinson's disease 9), also known as Kufor-Rakeb syndrome. The disease is characterized by levodopa-responsive Parkinsonism, supranuclear gaze palsy, spasticity, and dementia. Previously, we have reported that Atp13a2 deficient medaka fish showed dopaminergic neurodegeneration and lysosomal dysfunction, indicating that lysosome-autophagy impairment might be one of the key pathogeneses of Parkinson's disease. Here, we established Atp13a2 deficient zebrafish using CRISPR/Cas9 gene editing. We found that the number of TH+neurons in the posterior tuberculum and the locus coeruleus significantly reduced (dopaminergic neurons, 64 % at 4 months and 37 % at 12 months, p<0.001 and p<0.05, respectively; norepinephrine neurons, 52 % at 4 months and 40 % at 12 months, p<0.001 and p<0.05, respectively) in Atp13a2 deficient zebrafish, proving the degeneration of dopaminergic neurons. In addition, we found the reduction (60 %, p<0.05) of cathepsin D protein expression in Atp13a2 deficient zebrafish using immunoblot. Transmission electron microscopy analysis using middle diencephalon samples from Atp13a2 deficient zebrafish showed lysosome-like bodies with vesicle accumulation and fingerprint-like structures, suggesting lysosomal dysfunction. Furthermore, a significant reduction (p<0.001) in protein expression annotated with vesicle fusion with Golgi apparatus in Atp13a2 deficient zebrafish by liquid-chromatography tandem mass spectrometry suggested intracellular trafficking impairment. Therefore, we concluded that Atp13a2 deficient zebrafish exhibited degeneration of dopaminergic neurons, lysosomal dysfunction and the possibility of intracellular trafficking impairment, which would be the key pathogenic mechanism underlying Parkinson's disease.
