| Prion diseases, also known as transmissible spongiform encephalopathies (TSEs), are a group of fatal neurodegenerative diseases affecting both humans and animals worldwide. A central event in these diseases is the conformational modification of the normal cellular prion protein (PrPc) from a soluble, predominant alpha-helical conformation to a disease-causing, infectious form (PrPSc) that isβ sheet-rich, insoluble, and protease resistant. The misfolding, aggregation and accumulation of PrPSc on the neuronal cell leads to amyloid deposition, eventually resulting in cytotoxicity and cell death. Common animal prion diseases include, bovine spongiform encephalopathy (BSE), scrapie in sheep and goats, chronic wasting disease (CWD) in deer and Elk, and Transmissible mink encephalopathy (TME) in Mink, while human disease include Kuru, Creutzfeldt-Jakob disease (CJD), Variant Creutzfeldt-Jakob disease (vCJD) and Fatal familial insomnia (FFI). Despite the low incidence of prion disease in humans, it remains in spotlight due to the unique biology of its transmissible agent and the zoonotic potential of some animal prion diseases which represents a constant threat to public health. Parkin is a 53-kDa cytosolic protein containing 465 amino acid residues and a unique RING/HECT hybrid that includes both major classes of E3 ligases in one protein. Parkin plays a vital role in the degradation of misfolded proteins and has been reported to mediate both K48-linked and K63-linked polyubiquitination to accomplish degradation of several putative substrates. Parkin mutation is linked with Autosomal recessive juvenile Parkinson’s disease (ARJPD). Parkin expression decrease β-amyloid load and down regulate amyloidogenic protein expression in Alzheimer’s disease (AD). Parkin is reported to reduce inflammation, reactive oxygen species (ROS) and reduces oxidative stress. Parkin also mediates the selective autophagy of dysfunctional mitochondria by initiating mitophagy in Parkinson’s disease (PD). Parkin ameliorates the autophagic defects and enhances autophagic clearance of autophagosomes and autophagic vacuoles leading to clearance of Aβ in AD models. Inshort parkin is such a multifunctional protein that has broad neuroprotective properties against a wide range of toxic insults and can be exploited as a therapeutic strategy in neurodegenerative pathologies. To date. most parkin-related research has been conducted in either PD or AD models, but almost no work has been performed to determine the role of parkin in prion diseases. The current study was designed to evaluate the possible neuroprotective role of parkin in prion disease and to elucidate the parkin-mediated cytoprotective mechanism(s).In current study we investigated the role of parkin in a prion disease cell model employing neuroblastoma2a (N2a) cells and a prion peptide fragment Prp106-126. Prp106-126 retains the ability of PrPSc to aggregate into amyloid-like fibrils. Therefore, PrP106-126 is often used is as a model for the in vitro study of prion-induced cell toxicity and death. Our results of PrP106-126 treated cells showed a gradual decrease in the soluble parkin level in a time-dependent manner. Furthermore, the Immunocytochemistry revealed the co-localization of parkin with FITC tagged prion fragment 106-126, suggesting the interaction between parkin and FITC-PrP106-126 leading to ubiquitination and clearance of PrP fragment. Overexpression of parkin in N2a cells via transfection with pCMV-HA-Parkin plasmid DNA alleviated PrP106-126-induced apoptosis evident by the Annexin V FITC assay showing significant reduction in Annexin V FITC positive cells in parkin expressing cells compared to pCMV-HA empty vector-transfected cells. While CCK-8 assay also showed significant difference in cell viability between pCMV-HA-Parkin expressing cells and empty vector cells on treatment with the PrP106-126. On treatment with PrP106-126 parkin overexpressed cells showed enhanced autophagy compared to empty vector cells. Parkin overexpression also showed reduction in apoptotic Bax translocation to the mitochondria and cytochrome c release to the cytosol, which ultimately inhibited activation of proapoptotic caspase 9,12 and 3. In conclusion, our results indicate that parkin protects against PrP106-126-induced neurotoxicity and cell death by enhancing autophagy, reducing ER stress, suppressing Bax translocation to mitochondria, and inhibiting cytochrome c release leading to abridged activation of proapoptotic caspases. Although further research is necessary to explore in depth the factors that enable autophagy to regulate distinct cell survival and death response mechanisms, our results suggest that parkin may have benefits for the therapy in prion diseases. |
PDF Full Text Request