College of Life Sciences, Lamprey Research Center, Liaoning Provincial Key Laboratory of Biotechnology and Drug discovery, Liaoning Normal University, Dalian, 116081, China.
210th Hospital of PLA, Dalian, 116011, China.
Sci Rep. 2018 Mar 2;8(1):3932. doi: 10.1038/s41598-018-22212-0.
Prohibitin 2(PHB2) is a member of the SFPH trans-membrane family proteins. It is a highly conserved and functionally diverse protein that plays an important role in preserving the structure and function of the mitochondria. In this study, the lamprey PHB2 gene was expressed in HeLa cells to investigate its effect on cell proliferation. The effect of Lm-PHB2 on the proliferation of HeLa cells was determined by treating the cells with pure Lm-PHB2 protein followed by MTT assay. Using the synchronization method with APC-BrdU and PI double staining revealed rLm-PHB2 treatment induced the decrease of both S phase and G0/G1 phase and then increase of G2/M phase. Similarly, cells transfected with pEGFP-N1-Lm-PHB2 also exhibited remarkable reduction in proliferation. Western blot and quantitative real-time PCR(qRT-PCR) assays suggested that Lm-PHB2 caused cell cycle arrest in HeLa cells through inhibition of CDC25C and CCNB1 expression. According to our western blot analysis, Lm-PHB2 was also found to reduce the expression level of Wee1 and PLK1 and the phosphorylation level of CCNB1, CDC25C and CDK1 in HeLa cells. Lamprey prohibitin 2 could arrest G2/M phase transition of HeLa cells through down-regulating expression and phosphorylation level of cell cycle proteins.
PHB2 是 SFPH 跨膜家族蛋白的成员。它是一种高度保守且功能多样的蛋白质，在维持线粒体的结构和功能方面发挥着重要作用。在这项研究中，我们在 HeLa 细胞中表达了七鳃鳗 PHB2 基因，以研究其对细胞增殖的影响。通过用纯 Lm-PHB2 蛋白处理细胞，然后通过 MTT 测定来确定 Lm-PHB2 对 HeLa 细胞增殖的影响。使用 APC-BrdU 和 PI 双重染色的同步方法表明 rLm-PHB2 处理诱导 S 期和 G0/G1 期减少，然后 G2/M 期增加。同样，转染 pEGFP-N1-Lm-PHB2 的细胞也表现出明显的增殖减少。Western blot 和定量实时 PCR(qRT-PCR) 分析表明，Lm-PHB2 通过抑制 CDC25C 和 CCNB1 的表达导致 HeLa 细胞中的细胞周期停滞。根据我们的 Western blot 分析，Lm-PHB2 还被发现降低了 HeLa 细胞中 Wee1 和 PLK1 的表达水平以及 CCNB1、CDC25C 和 CDK1 的磷酸化水平。七鳃鳗 prohibitin 2 可以通过下调细胞周期蛋白的表达和磷酸化水平来阻滞 HeLa 细胞的 G2/M 期转换。
