Abstract
B cell lymphoma 2 (Bcl-2) is an important antiapoptotic gene that plays a dual role in the maintenance of the dynamic balance between the survival and death of cancer cells. In our previous study, Bcl-2 was shown to delay the G0/G1 to S phase entry by regulating the mitochondrial metabolic pathways to produce lower levels of adenosine triphosphate (ATP) and reactive oxygen species (ROS). However, the detailed molecular mechanisms or pathways by which Bcl-2 regulates the cell cycle remain unknown. Here, we compared the effects of Bcl-2 overexpression with an empty vector control in the NIH3T3 cell line synchronized by serum starvation, and evaluated the effects using proteomic analysis. The effect of Bcl-2 on cell cycle regulation was detected by monitoring Bcl-2 and p27 expression. The result of subsequent proteomic analysis of Bcl-2 overexpressing cells identified 169 upregulated and 120 downregulated proteins with a 1.5-fold change. These differentially expressed proteins were enriched in a number of signaling pathways predominantly involving the ribosome and oxidative phosphorylation, according to the data of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. These results indicated that Bcl-2 potentially acts at the translation level to influence proteins or enzymes of the respiratory chain or in the ribosome, and thereby regulates the cell cycle. Additionally, differentially expressed proteins involved in oxidative phosphorylation were determined to account for most of the effects of Bcl-2 on the cell cycle mediated by the mitochondrial pathway investigated in our previous study. These results can provide assistance for additional in-depth studies on the regulation of the cell cycle by Bcl-2. The results of the proteomic analysis determined the mechanism of Bcl-2-dependent delay of the cell cycle progression. In summary, the results of this study provide a novel mechanistic basis for identifying the key proteins or pathways for designing and develo** precisely targeted cancer drugs.
Abstract
目的
B细胞淋巴瘤-2(Bcl-2)基因除了广为人知的抗凋亡功能之外, 还具有调控细胞周期的非凋亡功能, 但是机制却不清楚. 作者前期研究发现Bcl-2可以通过阻滞G0/G1期进入S期的进程调控细胞周期, 可能与低水**的三磷酸腺苷(ATP)和活性氧自由基(ROS)有关. 因此, 本研究旨在探究其潜在调控机制.
创新点
基于Bcl-2通过ATP和ROS调控细胞周期的前期发现, 本研究首次利用蛋白组学方法系统研究了Bcl-2调控细胞周期的潜在机制.
方法
联合利用蛋白质印迹(western blotting)和蛋白质组学方法研究血清饥饿同步化处理的Bcl-2过表达和对照组细胞株, 并结合蛋白组学中差异蛋白的基因本体(Gene Ontology, GO)和Kyoto Encyclopedia of Genes and Genomes(KEGG)分析, 进一步明确Bcl-2调控细胞周期的潜在机制.
结论
蛋白组学结果显示, 在1.5倍差异下共有169个蛋白发生了上调, 120个蛋白发生了下调. 通过GO和KEGG分析, 这些差异蛋白富集到多个通路, 主要集中在呼吸链和核糖体相关信号通路. 这些结果表明Bcl-2可能在翻译水**影响核糖体和氧化磷酸化进而调控细胞周期. 本研究为进一步靶向Bcl-2调控细胞周期抗癌药物研究了提供重要的理论基础.
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Acknowledgments
This work was supported by the Ningxia Higher Education Scientific Research Project (No. NGY2018-69), the National Natural Science Foundation of China (No. 81960480), the Key Research and Development Program of Ningxia (No. 2018BEB04008), and the Ningxia Medical University Scientific Research Project (No. XT2017015), China.
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**ng DU, **g**g XIAO, Bo XU, and Hang HAN performed the experimental research and data analysis. **ng DU and Xufeng FU wrote and edited the manuscript. Yin WANG and **uying PEI contributed to the study design, data analysis, and writing and editing of the manuscript. All authors have read and approved the final manuscript and, therefore, have full access to all the data in the study and take responsibility for the integrity and security of the data.
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**ng DU, **g**g XIAO, Xufeng FU, Bo XU, Hang HAN, Yin WANG and **uying PEI declare that they have no conflict of interest.
This article does not contain any studies with human or animal subjects performed by any of the authors.
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Du, X., **ao, J., Fu, X. et al. A proteomic analysis of Bcl-2 regulation of cell cycle arrest: insight into the mechanisms. J. Zhejiang Univ. Sci. B 22, 839–855 (2021). https://doi.org/10.1631/jzus.B2000802
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DOI: https://doi.org/10.1631/jzus.B2000802