Abstract
Cell fate of mitotic cell is controlled by spindle assembly. Deficient spindle assembly results in mitotic catastrophe leading to cell death to maintain cellular homeostasis. Therefore, inducing mitotic catastrophe provides a strategy for tumor therapy. Nucleolar acetyltransferase NAT10 has been found to regulate various cellular processes to maintain cell homeostasis. Here we report that NAT10 regulates mitotic cell fate by acetylating Eg5. NAT10 depletion results in multinuclear giant cells, which is the hallmark of mitotic catastrophe. Live-cell imaging showed that knockdown of NAT10 dramatically prolongs the mitotic time and induces defective chromosome segregation including chromosome misalignment, bridge and lagging. NAT10 binds and co-localizes with Eg5 in the centrosome during mitosis. Depletion of NAT10 reduces the centrosome loading of Eg5 and impairs the poleward movement of centrosome, leading to monopolar and asymmetrical spindle formation. Furthermore, NAT10 stabilizes Eg5 through its acetyltransferase function. NAT10 acetylates Eg5 at K771 to control Eg5 stabilization. We generated K771-Ac specific antibody and showed that Eg5 K771-Ac specifically localizes in the centrosome during mitosis. Additionally, K771 acetylation is required for the motor function of Eg5. The hyper-acetylation mimic Flag-Eg5 K771Q but not Flag-Eg5 rescued the NAT10 depletion-induced defective spindle formation and mitotic catastrophe, demonstrating that NAT10 controls mitosis through acetylating Eg5 K771. Collectively, we identify Eg5 as an important substrate of NAT10 in the control of mitosis and provide K771 as an essential acetylation site in the stabilization and motor function of Eg5. Our findings reveal that targeting the NAT10-mediated Eg5 K771 acetylation provides a potential strategy for tumor therapy.
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The datasets used and/or analyzed during the current study are available from the corresponding author on reasonable request.
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Acknowledgements
We thank Dr. Qihua He for assistance with confocal microscopy, Dr. Xiajuan Zou for assistance in the mass spectrometric analysis. We thank Prof. Bo Zhang for providing us the anti-NAT10 antibody and Prof. Chenghao Xuan for providing us the GFP-RNF20 and GFP-RNF40 plasmids.
Funding
This work was supported by the grants from the National Natural Science Foundation of China (Grant No. 82173024 and 81874143 to XD), the Beijing Natural Science Foundation (Grant No.7212061 to XD) and the National Natural Science Foundation of China (Grant No. 81802305 to XL).
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JZ designed and performed experiments, analyzed the data and wrote the manuscript. YT performed the experiments and analyzed the data. XL helped with the mass spectrometry analysis. CZ helped with cell culture and purchased the reagents. KS helped with protein purification. YJ helped with live-cell imaging. JL provided experiment help and revised the manuscript. LL designed and supervised the study. XD designed and acquired the funding for the study, analyzed the data and revised the manuscript.
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Zheng, J., Tan, Y., Liu, X. et al. NAT10 regulates mitotic cell fate by acetylating Eg5 to control bipolar spindle assembly and chromosome segregation. Cell Death Differ 29, 846–860 (2022). https://doi.org/10.1038/s41418-021-00899-5
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DOI: https://doi.org/10.1038/s41418-021-00899-5
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