The Inhibition of DYRK1A Improves Parkinsons Disease by Inhibiting the Phosphorylation of α-Synuclein and Apoptosis of Dopaminergic Neurons

Abstract

Objective: Dual-specificity tyrosine-regulated kinase 1A (DYRK1A) is the main kinase which phosphorylates the residues of alpha-synuclein (α-syn). It has been reported that Single Nucleotide Polymorphisms (SNPs) distribution of the DYRK1A gene is associated with the risk of Parkinsons disease (PD) in different races. However, it is still unclear how α-syn phosphorylation modifications mediated by the DYRK1A gene and its mutations affect the progression of PD. The goal of this study is to uncover the molecular mechanisms involved in α-syn phosphorylation mediated by DYRK1A in dopaminergic (DA) neuronal apoptosis. Methods: SH-SY5Y cells were treated with 1-methyl-4-phenylpyridinium (MPP⁺) to establish PD model. Negative control small interfering RNA (siRNA) and DYRK1A siRNA reagents were administered to establish negative control (NC) siRNA and DYRK1A siRNA groups. Empty vector and lentivirus plasmids were employed to construct the Vector and DYRK1A overexpression (OE) groups. Cell apoptosis was then assessed via western blotting and flow cytometry. Western blotting was used to determine the α-syn phosphorylation and the related protein expression of the Akt (Akt kinase)/GSK3β (glycogen synthase kinase 3 beta)/NF-κB (nuclear factor kappa B)/Cyclin D1 pathway. Co-immunoprecipitation and western blotting were employed to investigate the interaction between α-syn and GSK3β. Upstream and downstream relative gene expressions in the Akt/GSK3β pathway, including PI3K (phosphatidylinositol 3-kinase), mTOR (mechanistic target of rapamycin kinase), c-Myc (Myc proto-oncogene), and HIF1α (hypoxia inducible factor 1 subunit alpha), were detected by quantitative Polymerase Chain Reaction (qPCR) analysis. Results: We found that DYRK1A knockdown resulted in lower phosphorylated-alpha-synuclein (P-α-syn, Ser129) levels (p < 0.01), whereas DYRK1A overexpression heightened levels of P-α-syn (p < 0.01). Furthermore, DYRK1A interacts with α-syn and GSK3β. DYRK1A knockdown resulted in activation of the Akt/GSK3β/NF-κB pathway to promote DA neuronal apoptosis. Finally, DYRK1A knockdown increased the messenger RNA (mRNA) levels of PI3K, mTOR, c-Myc, and HIF1α (p < 0.01), which are involved in the Akt/GSK3β/NF-κB signaling pathway. Conclusions: These findings demonstrate that DYRK1A is responsible for α-syn phosphorylation and regulates activation of the PI3K/Akt/GSK3β pathway in DA neuronal apoptosis.