Impaired dephosphorylation renders G6PD-knockdown HepG2 cells more susceptible to H(2)O(2)-induced apoptosis.
Glucose-6-phosphate dehydrogenase (G6PD) plays a key role in the regeneration of NADPH and maintenance of cellular redox balance. In the present study, we investigate the effect of G6PD deficiency on H(2)O(2)-elicited signaling in HepG2 cells. H(2)O(2) was found to inhibit cellular protein tyrosine phosphatase (PTP) activity, resulting in activation of MAPKs. MKP-1 expression increased in the late phase of H(2)O(2) signaling. Using RNAi technology, we found that G6PD knockdown enhanced the inhibitory effect of H(2)O(2) on PTPs and led to sustained MAPK activation. This was accompanied by delayed expression and inhibition of MKP-1. Using a pharmacological inhibitor and siRNA, we demonstrate that MKP-1 acts as a regulator of MAPK activation in H(2)O(2) signaling. The prolonged MAPK activation in G6PD-knockdown cells was associated with an increased susceptibility to H(2)O(2)-induced apoptosis and growth retardation. Treatment with p38 and JNK inhibitors or N-acetylcysteine ameliorated such cellular effect, while triptolide and MKP-1-siRNA did the opposite. Glucose oxidase treatment had similar effects as addition of H(2)O(2). Taken together, these findings suggest that G6PD knockdown enhances the magnitude and duration of H(2)O(2)-induced MAPK signaling through inhibition of cellular PTPs, and the resultant anomalous signaling may lead to cell demise.
Lin
Chang-Jun
CJ
Graduate Institute of Medical Biotechnology, Chang Gung University, Tao-Yuan, Taiwan.
Ho
Hung-Yao
HY
Cheng
Mei-Ling
ML
You
Tsai-Hong
TH
Cheng
Tsai-Hong
TH
Yu
Jau-Song
JS
Chiu
Daniel Tsun-Yee
DT
D000111:Acetylcysteine / Q000494:pharmacology; D000873:Anthracenes / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects*; D054638:Dual Specificity Phosphatase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D005419:Flavonoids / Q000494:pharmacology; D055785:Gene Knockdown Techniques; D005954:Glucosephosphate Dehydrogenase / Q000235:genetics*; D056945:Hep G2 Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007093:Imidazoles / Q000494:pharmacology; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors; D011725:Pyridines / Q000494:pharmacology
D000873:Anthracenes; D005419:Flavonoids; D007093:Imidazoles; D011725:Pyridines; C432165:pyrazolanthrone; D006861:Hydrogen Peroxide; D005954:Glucosephosphate Dehydrogenase; D020928:Mitogen-Activated Protein Kinases; C517019:DUSP1 protein, human; D054638:Dual Specificity Phosphatase 1; D017027:Protein Tyrosine Phosphatases; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D000111:Acetylcysteine