Zhao X, Yang K, Song Z, He H, Zhang W
Department of Biochemistry, Jilin Medical College, Jilin 132013, China.
Department of Biochemistry, School of Basic Medicine, Yanbian University, Yanbian 133000, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2022 Jul 20;42(7):1026-1031. doi: 10.12122/j.issn.1673-4254.2022.07.09.
To observe the expression of c-Myc protein in cervical cancer HeLa cells and explore the effect of juglone on the proliferation and apoptosis of HeLa cells by affecting c-Myc ubiquitination.
HeLa cells treated with different concentrations (0, 10, 20, or 50 μmol/L) of juglone or with 20 μmol/L juglone for different time lengths were examined for expression of c-Myc protein with Western blotting. The half-life of c-Myc protein was determined using cycloheximide (CHX) and c-Myc protein degradation was detected using coimmunoprecipitation. We also assessed the effects of 20 μmol/L juglone combined with 0, 1.0 or 2.0 μmol/L MG132 (a proteasome inhibitor) on c-Myc expression. The effects of 20 μmol/L juglone on the proliferation and apoptosis of HeLa cells with RNA interference-mediated knockdown of c-Myc were evaluated with MTT assay and flow cytometry.
Treatment with juglone significantly lowered c-Myc protein expression in HeLa cells in a concentration-and time-dependent manner ( < 0.05). Juglone obviously shortened the half-life of c-Myc protein, and the addition of MG132 significantly up-regulated the expression level of c-Myc protein ( < 0.05). Juglone treatment also promoted ubiquitination of c-Myc protein in HeLa cells. Compared with the cells transfected with a negative control construct, the cells transfected with si-c-Myc showed significantly decreased proliferation inhibition and a lowered cell rate with early apoptosis after juglone treatment ( < 0.05).
Juglone inhibits proliferation and promotes apoptosis of HeLa cells by affecting the ubiquitination of c-Myc protein.
观察c-Myc蛋白在宫颈癌HeLa细胞中的表达情况，并探讨胡桃醌通过影响c-Myc泛素化对HeLa细胞增殖和凋亡的影响。
用不同浓度（0、10、20或50 μmol/L）的胡桃醌处理HeLa细胞，或用20 μmol/L胡桃醌处理不同时间，采用蛋白质免疫印迹法检测c-Myc蛋白的表达。使用放线菌酮（CHX）测定c-Myc蛋白的半衰期，并通过免疫共沉淀检测c-Myc蛋白的降解情况。我们还评估了20 μmol/L胡桃醌与0、1.0或2.0 μmol/L MG132（一种蛋白酶体抑制剂）联合使用对c-Myc表达的影响。采用MTT法和流式细胞术评估20 μmol/L胡桃醌对RNA干扰介导的c-Myc基因敲低的HeLa细胞增殖和凋亡的影响。
胡桃醌处理可显著降低HeLa细胞中c-Myc蛋白的表达，呈浓度和时间依赖性（<0.05）。胡桃醌明显缩短了c-Myc蛋白的半衰期，添加MG132可显著上调c-Myc蛋白的表达水平（<0.05）。胡桃醌处理还促进了HeLa细胞中c-Myc蛋白的泛素化。与转染阴性对照构建体的细胞相比，转染si-c-Myc的细胞在胡桃醌处理后增殖抑制明显降低，早期凋亡细胞率降低（<0.05）。
胡桃醌通过影响c-Myc蛋白的泛素化抑制HeLa细胞的增殖并促进其凋亡。
