Abstract
Chaperone-mediated autophagy (CMA) is involved in wild-type α-synuclein degradation in Parkinson’s disease (PD), and LAMP2A and Hsc 70 have recently been indicated to be deregulated by microRNAs. To recognize the regularory role of miR-320a in CMA and the possible role in α-synuclein degradation, in the present study, we examined the targeting and regulating role of miR-320 in Hsc 70 expression. We first constructed an α-synuclein-overexpressed human neuroblastoma cell line, SH-SY5Y-Syn(+), stably over-expressing wild-type α-synuclein and sensitive to an autophagy inhibitor, which exerted no effect on the expression of LAMP2A and Hsc 70. Then we evaluated the influence on the CMA by miR-320a in the SH-SY5Y-Syn(+) cells. It was shown that miR-320a mimics transfection of specifically targeted Hsc 70 and reduced its expression at both mRNA and protein levels, however, the other key CMA molecule, LAMP2A was not regulated by miR-320a. Further, the reduced Hsc 70 attenuated the α-synuclein degradation in the SH-SY5Y-Syn(+) cells, and induced a significantly high level of α-synuclein accumulation. In conclusion, we demonstrate that miR-320a specifically targeted the 3' UTR of Hsc 70, decreased Hsc 70 expression at both protein and mRNA levels in α-synuclein-over-expressed SH-SY5Y cells, and resulted in significant α-synuclein intracellular accumulation. These results imply that miR-320a might be implicated in the α-synuclein aggravation in PD.
Highlights
Parkinson’s disease (PD) is the second most common disorder of the central nervous system in humans [1], with a prominent characteristic of dopaminergic cells degeneration within the substantia nigra pars compacta (SNpc) [2] and the cytoplasmic accumulation of proteinaceous material within aggregates called Lewy bodies (LBs) [3]
To confirm whether there was an influence on the expression of Chaperone-mediated autophagy (CMA)-associated molecules, we examined the expression of Hsc 70 and LAMP2A in protein levels by western blot assay; it was shown in Figure 1B,D that there was no significant difference in the expression of the two molecules between
Compared to familial PD, which is believed to be caused by the multiplication of the α-synuclein locus and increased α-synuclein synthesis [8], sporadic PD is normal in α-synuclein synthesis, whereas it is deficient in α-synuclein degradation, which leads to α-synuclein accumulation and aggregation [45,46]
Summary
Parkinson’s disease (PD) is the second most common disorder of the central nervous system in humans [1], with a prominent characteristic of dopaminergic cells degeneration within the substantia nigra pars compacta (SNpc) [2] and the cytoplasmic accumulation of proteinaceous material within aggregates called Lewy bodies (LBs) [3]. A strong association of α-synuclein (α-Syn) aggregation with PD is suggested by elevated synthesis and/or reduced degradation. Increased α-synuclein gene copy number [8] promotes the α-synuclein synthesis and plays a role in PD. On the other side, impaired degradation pathways responsible for α-synuclein may be compromised in PD [9,10]. Almost all major known degradation pathways have been implicated in the degradation of α-synuclein [11,12], autophagy and proteasomal pathways are considered to play key roles in the process [13,14,15], in which both ubiquitin-proteasome system [16,17] and the autophagy-lysosomal pathway were confirmed to degrade α-synuclein [9,18]
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