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DJ-1/PARK7, the Parkinson's disease-related protein, plays an important role in mitochondrial function. However, the mechanisms by which DJ-1 affects mitochondrial function are not fully understood. Akt is a promoter of neuron survival and is partly involved in the neurodegenerative process. This research aimed at investigating a possible relationship between DJ-1 and Akt signalling in regulating mitochondrial function in the dopaminergic neuron-like cells SH-SY5Y and PC-12. Overexpression of DJ-1 was firstly validated at both the transcriptional and translational levels after transit transfection with plasmid pcDNA3-Flag-DJ-1. Confocal fluorescence microscopy demonstrated that overexpression of DJ-1 increased the mitochondrial mass, but did not disrupt the mitochondrial morphology. In addition, mitochondrial complex I activity was raised in DJ-1-overexpressing cells, and this rise occurred with an increase in cellular adenosine 50-triphosphate content. Moreover, immunoblotting demonstrated that the levels of phosphoinositide 3-kinase and the total Akt were not altered in DJ-1-overexpressing cells, and nor was the Akt phosphorylation on serine 473 changed. By contrast, Akt phosphorylation on threonine 308 was significantly augmented by overexpression of DJ-1, and the expression of glycogen synthase kinase-3beta, a downstream effector of Akt, was suppressed. In summary, these results suggest that overexpression of DJ-1 improves the mitochondrial function, at least in part, through a mechanism involving Akt phosphorylation on threonine 308. [ABSTRACT FROM AUTHOR]

Highlights • DJ-1 knockdown resulted in mitochondrial dysfunction. • loss of DJ-1 suppressed Akt pathway, and reduced nuclear translocation of Nrf2. • DJ-1 deficiency led to an obvious increase in the ROS level, and then activated JNK signaling. Abstract Deletions or some mutations in the gene encoding the multifunctional protein, DJ-1, have been considered to be linked with autosomal recessive early onset Parkinson's disease (PD). Current emerging evidence suggests that DJ-1 is involved in the protection against oxidative stress-induced mitochondrial damage. However, the exact molecular mechanisms underlying this are not completely clear. The aim of this study was to investigate the effects of DJ-1 on the Akt pathway, nuclear factor erythroid 2-related factor (Nrf2), and c-Jun N-terminal kinase (JNK) with regard to modulating mitochondrial function. Here we showed that knockdown of DJ-1 resulted in mitochondrial dysfunction, including a decrease in active mitochondrial mass, complex I deficits, and inhibition of cellular adenosine 5′-triphosphate (ATP) content in the dopaminergic neuron-like cells PC12 and SH-SY5Y. Additionally, loss of DJ-1 impaired Akt signaling, and reduced nuclear translocation of Nrf2, thereby inhibiting activity of Nrf2-regulated downstream antioxidant enzymes such as heme oxygenase-1 and NAD(P)H quinone oxidoreductase 1. Moreover, DJ-1 knockdown also led to a significant increase in the mitochondrial reactive oxygen species, and then promoted the activation of JNK pathways. Furthermore, oxidative stress and mitochondrial dysfunction induced by knockdown of DJ-1 were blocked by a JNK inhibitor, which confirmed the important role of JNK activation in mitochondrial dysfunction. In conclusion, the present study indicates that DJ-1 knockdown leads to mitochondrial dysfunction in dopaminergic neuron-like cells, at least in part, through suppressing the Akt/GSK3β pathway and impairing the oxidative stress response, as well as through the subsequent increased JNK activation in dopaminergic neuron-like cells. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Da-Bu-Yin-Wan (DBYW), a historically traditional Chinese medicine formula, was originally defined over 600 years ago. In recent decades, DBYW was clinically employed to treat Parkinson's disease (PD). Aim of the study To explore the underlying mechanism of DBYW on mitochondrial function, we investigated the effect of DBYW on mitochondrial function from the perspectives of DJ-1 and Akt signaling. Materials and methods Human derived neuroblastoma SH-SY5Y cells were transiently transfected with the plasmid pcDNA3-Flag-DJ-1 aimed to overexpress the DJ-1 protein. Transfected cells were treated with 1-methyl-4-phenylpyridinium (MPP + ), a PD-related mitochondrial complex I inhibitor, in the absence and presence of DBYW. The cell viability was assessed by Cell Counting Kit-8 assay. The protein expressions of DJ-1 and Akt signaling were examined by western blotting. The mitochondrial mass was evaluated by confocal fluorescence microscopy. The mitochondrial complex I activity and cellular ATP content were measured by commercial kits. Results Transfection with pcDNA3-Flag-DJ-1 decreased the MPP + -induced toxicity and overexpressed the DJ-1. In DJ-1 overexpressed cells, the mitochondrial mass was raised, mitochondrial complex I activity was improved, and cellular ATP content was increased. In addition, overexpression of DJ-1 augmented the Akt phosphorylation on threonine 308 and serine 473. Moreover, DBYW promoted the above effects in DJ-1 expressed cells. Conclusions These data suggest that DJ-1 protects the mitochondrial function by medicating Akt phosphorylation in MPP + -treated SH-SY5Y cells. Moreover, DBYW enhances the protective effect of DJ-1 medicated Akt phosphorylation on mitochondrial function. [ABSTRACT FROM AUTHOR]