HPV16 E6 蛋白对 C33A 细胞中 Daxx 诱导的凋亡的影响。

Effects of HPV16 E6 protein on Daxx-induced apoptosis in C33A cells.

机构信息

Pathogenic Biology Institute, University of South China, Hengyang, 421001 P. R. China.

出版信息

Cell Mol Biol Lett. 2020 Aug 8;25:38. doi: 10.1186/s11658-020-00230-z. eCollection 2020.

AIMS

Daxx is a highly conserved nuclear protein with an important role in transcription, apoptosis and other cell processes. We investigated the role of HPV16 E6 in Daxx-induced apoptosis through their interactions in C33A cells.

METHODS

The binding of HPV16 E6 and Daxx was confirmed in C33A cells using co-immunoprecipitation and indirect immunofluorescence assays. Quantitative PCR and western blotting were performed to determine the RNA and protein expressions of Daxx, respectively. Automatic cell count and MTT assays were performed to investigate the proliferation of C33A cells. The apoptosis rate of C33A cells was determined via flow cytometry using Annexin V-FITC/PI staining. The relative activity of caspase-8 was tested using ELISA.

RESULTS

HPV16 E6 can bind with Daxx and cause its translocation in C33A cells. The transfected HPV16 E6 can cause a decrease in relative quantification for Daxx in Daxx-overexpressing cells. After Daxx transfection, cell proliferation was found to decrease sharply and cell apoptosis to increase sharply. However, when HPV16 E6 was co-transfected with Daxx, this decrease and increase both became gentle. Similarly, HPV16 E6 made the Daxx-induced increase in caspase-8 activity milder.

CONCLUSIONS

HPV16 E6 is involved in inhibiting apoptosis through deregulation of Daxx-induced caspase-8 activities.

目的

Daxx 是一种高度保守的核蛋白，在转录、凋亡和其他细胞过程中具有重要作用。我们通过 HPV16 E6 与 Daxx 在 C33A 细胞中的相互作用，研究了 HPV16 E6 在 Daxx 诱导的细胞凋亡中的作用。

方法

通过共免疫沉淀和间接免疫荧光测定，在 C33A 细胞中证实了 HPV16 E6 与 Daxx 的结合。通过定量 PCR 和 Western blot 分别测定 Daxx 的 RNA 和蛋白质表达。自动细胞计数和 MTT 测定用于研究 C33A 细胞的增殖。通过 Annexin V-FITC/PI 染色，用流式细胞术测定 C33A 细胞的凋亡率。用 ELISA 测定 caspase-8 的相对活性。

结果

HPV16 E6 可以与 Daxx 结合，并导致其在 C33A 细胞中的易位。转染的 HPV16 E6 可导致 Daxx 过表达细胞中 Daxx 的相对定量减少。Daxx 转染后，细胞增殖明显下降，细胞凋亡明显增加。然而，当 HPV16 E6 与 Daxx 共转染时，这种减少和增加都变得温和。同样，HPV16 E6 使 Daxx 诱导的 caspase-8 活性增加变得温和。