| Uncontrolled proliferation and metabolic reprogramming are two hallmarks of cancer.Tumor cells proliferate via consecutive cell cycle progression.In order to meet the needs of proliferation,tumor cells reprogram metabolic pathways to provide intermediate metabolites and energy at different stages of the cell cycle.In addition,tumor cells maintain the redox homeostasis to avoid damages caused by dramatic redox changes.It was reported that the levels of redox-related metabolites fluctuate during cell cycle,and different cell cycle phases have different demands for building blocks and energy.Therefore,there might be a certain connection between the progress of tumor cell cycle and the redox changes mediated by metabolic reprogramming.However,how cell cycle inter-connects with redox metabolism is unclear.To explore the relationship between tumor cell cycle progression and redox state,I used a fluorescent probe to monitor the real-time dynamics of NADPH in living cells.The result showed that NADPH accumulated significantly in the mitosis of tumor cells.In order to explore the significance of the increased NADPH during tumor cell division,I first identified that G6PD is the main metabolic enzyme that promotes the accumulation of NADPH during mitosis by screening enzymes that generate NADPH.Biochemical experiments revealed that dimerization and enzyme activity of G6PD were up-regulated during mitosis.The results of functional experiments showed that inhibition of G6PD activity led to increase of intracellular ROS,decrease of Aurora kinase activity,and significant increase of chromosome mis-segregation.These results showed that by promoting the activity of G6PD,the tumor cells increased the level of NADPH to neutralize excessive ROS during mitosis,which maintained the activity of Aurora kinase and ensured the accurate separation of chromosomes.In order to find the factors that regulate the enzyme activity of G6PD during mitosis,we sought to explore the proteins that interact with G6PD at different stages of cell cycle by immunoprecipitation combined with mass spectrometry.The results showed that the co-chaperone protein BAG3,which can inhibit the activity of G6PD,can bind to G6PD during interphase,but not during mitosis.The results of subsequent biochemical experiments further revealed that BAG3 was highly phosphorylated during mitosis,and the phosphorylation modification of BAG3 was crucial for its interaction with G6PD.And we found that CDK1 and AMPK were the main upstream kinase of phosphorylation BAG3 during mitosis through screening.These results suggest that CDK1/AMPK-BAG3 axis regulates the dimerization and enzyme activity of G6PD during mitosis.The results of animal experiments showed that knockdown of G6PD or expression of BAG3T285A led to increase of chromosomal error segregation,production of micronucleus and tumor growth inhibition.To assess the clinical relevance of the BAG3-G6PD signaling,we analyzed the level of BAG3 phosphorylation in the tissue microarrays from colorectal cancer patients.Within 87 paired paracancer and cancer tissues BAG3 was highly phosphorylated in tumor cells during mitosis,and the phosphorylation level of BAG3 was closely related to the poor prognosis of patients.In summary,we found that CDK1/AMPK-BAG3-G6PD axis can promote the accumulation of NADPH during mitosis to maintain redox balance,promote Aurora kinase activation,ensure accurate chromosome separation and accelerate tumor progression.Our findings will contribute to an in-depth understanding of the role of G6PD mediated antioxidant pathway in tumor progression,and provide an important theoretical basis for the clinical diagnosis and treatment of tumors. |
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