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Proteasome inhibition can induce abnormal accumulation and phosphorylation of microtubule-associated protein tau. The major function of tau protein is to promote microtubules assembly and stabilization, and abnormal tau protein would disturb its microtubule-binding function. In this study, proteasome inhibitor MG132 was used to treat hippocampal slices to explore the role and mechanism of Akt/glycogen synthase kinase-3β (GSK-3β) in proteasome inhibition-induced tau abnormality. During the culture period, we measure the lactate dehydrogenase (LDH) content to assay the viability of hippocampal slices. Following 2.5 and 5 μM MG132 treatment for 6 h, we detected the expression, phosphorylation modification, and microtubule-binding function of tau protein of slices. We also analyzed the changed activities of glycogen synthase kinase-3β (GSK-3β) and protein kinase B (PKB/Akt) and the level of heat shock protein 90 (Hsp90) in the process. In addition, co-immunoprecipitation was used to investigate the interaction between Akt and Hsp90, Akt and protein phosphatase-2A (PP2A) in the MG132-treated organotypic hippocampal slices. Our results indicated that proteasome inhibition led to degradation obstacles and abnormal phosphorylation of tau protein. The downregulated Akt/GSK-3β signaling pathway might be responsible for the abnormal phosphorylation of tau protein at multiple sites which further reduced the microtubule-binding function of tau protein. Furthermore, proteasome inhibition decreased the binding capacity of Akt-Hsp90 while increased the Akt-PP2A binding ability which mediated Akt inactivity. This current study establishes a hippocampal slice model targeting Akt/GSK-3β signaling pathway to explore the pivotal role of proteasome inhibition in tau pathology.
Ubiquitin C-terminal hydrolase L1 (UCH-L1) is abundantly expressed in the brain and is critical for the normal function of synapses. cAMP response element binding protein (CREB) is a transcription factor which initiates the expression of proteins that related to the regulation of synaptic plasticity and memory function. Studies have shown that UCH-L1 can influence the expression and activity of CREB, but the underlying mechanisms remain unclear. In this study, we used UCH-L1 inhibitor LDN to treat mice hippocampal slices and found that UCH-L1 inhibition caused the dephosphorylation of CREB at Ser133 site. Meanwhile, hyperphosphorylation of microtubule-associated protein tau; increased expression of synaptic protein components of PSD-95 and synapsin-1, and decreased activity of tyrosine kinase Fyn were observed after UCH-L1 inhibition. Moreover, all these alternations have an influence on the normal function of N-methyl-D-aspartate (NMDA) receptor NR2B subunit which is likely to result in the dephosphorylation of CREB. We also found that LDN treatment mediated protein kinase A (PKA) deactivation was involved in the dephosphorylation of CREB. Thus, our study introduces a novel possible mechanism for elaborating the effects of UCH-L1 inhibition on the CREB activity and the implicated signaling pathways.