The Inhibitory Effect of OGT Gene Expression on The Level of tau Phosphorylation / 生物化学与生物物理进展

ABSTRACT

To investigate the effect of OGT expression inhibited by RNAi on the alteration of tau phosphorylation and glycosylation level in HEK293T cells.The siRNAs targeting OGTgene(OGT-siRNA1～3) were designed and chemically synthesized.The OGT-siRNA1～3 were transfected into HEK293T cells via lipofectamine2000.The efficacy of RNA interference was detected by RT-PCR.The fluorescence of GFP/OGT was counted by fluorescence microscopy after pEGFP/OGT and OGT-siRNA1～3 cotransfected to HEK293T cells for 24 h.The level of tau phosphorylation and glycosylation were detected by Western blot after Plasmid pCI/tau441 and the siRNA cotransfected HEK293T cells for 48 h.OGT-siRNA3(100 nmol/L) could effectively downregulate the expression of OGT gene compared with Mock group(80.0% at mRNA level and 51.3% at protein level).The level of phosphorylation at various sites of the tau was significantly upregulated and the glycosylation was downregulated following the inhibition of OGT gene expression.There is an apparent negative correlation between the modification of phosphorylation and glycosylation of tau protein, the deficient in glucose uptaken/metabolism may be an important pathogenesis of AD.