ABSTRACT
The O-linked-ß-N-acetylglucosamine (O-GlcNAc) glycosylation (O-GlcNAcylation) is a critical post-translational modification that couples the external stimuli to intracellular signal transduction networks. However, the critical protein targets of O-GlcNAcylation in oxidative stress-induced apoptosis remain to be elucidated. Here, we show that treatment with H2O2 inhibited O-GlcNAcylation, impaired cell viability, increased the cleaved caspase 3 and accelerated apoptosis of neuroblastoma N2a cells. The O-GlcNAc transferase (OGT) inhibitor OSMI-1 or the O-GlcNAcase (OGA) inhibitor Thiamet-G enhanced or inhibited H2O2-induced apoptosis, respectively. The total and phosphorylated protein levels, as well as the promoter activities of signal transducer and activator of transcription factor 3 (STAT3) and Forkhead box protein O 1 (FOXO1) were suppressed by OSMI-1. In contrast, overexpressing OGT or treating with Thiamet-G increased the total protein levels of STAT3 and FOXO1. Overexpression of STAT3 or FOXO1 abolished OSMI-1-induced apoptosis. Whereas the anti-apoptotic effect of OGT and Thiamet-G in H2O2-treated cells was abolished by either downregulating the expression or activity of endogenous STAT3 or FOXO1. These results suggest that STAT3 or FOXO1 are the potential targets of O-GlcNAcylation involved in the H2O2-induced apoptosis of N2a cells.
Subject(s)
Apoptosis , Forkhead Box Protein O1 , Hydrogen Peroxide , STAT3 Transcription Factor , Hydrogen Peroxide/pharmacology , Hydrogen Peroxide/metabolism , N-Acetylglucosaminyltransferases/genetics , N-Acetylglucosaminyltransferases/metabolism , Protein Processing, Post-Translational , Signal Transduction , Glycosylation , Acylation , STAT3 Transcription Factor/metabolism , Forkhead Box Protein O1/metabolism , Animals , Mice , Cell Line, TumorABSTRACT
Protein O-GlcNAcylation is a post-translational modification that links environmental stimuli with changes in intracellular signal pathways, and its disturbance has been found in neurodegenerative diseases and metabolic disorders. However, its role in the mesolimbic dopamine (DA) system, especially in the ventral tegmental area (VTA), needs to be elucidated. Here, we found that injection of Thiamet G, an O-GlcNAcase (OGA) inhibitor, in the VTA and nucleus accumbens (NAc) of mice, facilitated neuronal O-GlcNAcylation and decreased the operant response to sucrose as well as the latency to fall in rotarod test. Mice with DAergic neuron-specific knockout of O-GlcNAc transferase (OGT) displayed severe metabolic abnormalities and died within 4-8 weeks after birth. Furthermore, mice specifically overexpressing OGT in DAergic neurons in the VTA had learning defects in the operant response to sucrose, and impaired motor learning in the rotarod test. Instead, overexpression of OGT in GABAergic neurons in the VTA had no effect on these behaviors. These results suggest that protein O-GlcNAcylation of DAergic neurons in the VTA plays an important role in regulating the response to natural reward and motor learning in mice.
