醋酸棉酚通过磷脂酰肌醇 3-激酶（PI3K）家族诱导肿瘤细胞中 H2AX 的磷酸化。

Induction of H2AX phosphorylation in tumor cells by gossypol acetic acid is mediated by phosphatidylinositol 3-kinase (PI3K) family.

机构信息

Medical College of Northwest University for Nationalities, Lanzhou, 730030 PR China.

出版信息

Cancer Cell Int. 2014 Dec 16;14(1):141. doi: 10.1186/s12935-014-0141-5. eCollection 2014.

DOI:10.1186/s12935-014-0141-5

PMID:25530717

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC4272777/

Abstract

BACKGROUND

H2AX is phosphorylated (γH2AX) by members of the phosphatidylinositol 3-kinase (PI3K) family, including Ataxia telangiectasia-mutated (ATM), ATM- and Rad3-related (ATR) and DNA-PK in response to DNA damage. Our study shows that gossypol acetic acid (GAA) alone can induce γH2AX in Human mucoepidermoid carcinoma cell line (MEC-1) in vitro. Thus, we further examined the possible mechanisms of GAA to induce γH2AX in tumor cells.

MATERIALS AND METHODS

The PI3K inhibitors caffeine and wortmannin were used in an effort to identify the kinase(s) responsible for GAA -induced γH2AX in MEC-1 cells. DNA dependent protein kinase (DNA-PK) - proficient and -deficient cells, human glioma cell lines M059K and M059J, were also used to evaluate the kinases responsible for GAA induced H2AX phosphorylation. γH2AX expression was detected by immunofluorescent microscopy. Flow cytometry assay was used to assay γH2AX and cell cycle.

RESULTS

GAA induced H2AX phosphorylation in a cell cycle-dependent manner and a significant G0/G1 phase arrest in MEC-1 cells was shown. Caffeine and wortmannin significantly inhibited GAA-induced H2AX phosphorylation in MEC-1 cells. GAA induced H2AX phosphorylation in M059K, but not in M059J. Taken together, these data suggested that GAA treatment alone could induce H2AX phosphorylation in a cell cycle dependent manner in MEC-1 and M059K, but not in M059J cells. A significant G0/G1 phase arrest was shown in MEC-1.

CONCLUSIONS

The member of PI3K family, DNA-PK, ATM and ATR are involved in the H2AX phosphorylation of MEC-1 cells.

摘要

背景

磷脂酰肌醇 3-激酶（PI3K）家族成员，包括共济失调毛细血管扩张突变（ATM）、ATM 和 Rad3 相关（ATR）以及 DNA 依赖性蛋白激酶（DNA-PK），在 DNA 损伤时使组蛋白 H2AX 发生磷酸化（γH2AX）。我们的研究表明，棉酚乙酸（GAA）可单独诱导体外人黏液表皮样癌细胞系（MEC-1）中γH2AX 的产生。因此，我们进一步研究了 GAA 诱导肿瘤细胞中γH2AX 的可能机制。

材料与方法

使用 PI3K 抑制剂咖啡因和渥曼青霉素，以确定导致 GAA 诱导 MEC-1 细胞中γH2AX 的激酶。还使用 DNA 依赖性蛋白激酶（DNA-PK）有缺陷和无缺陷的细胞，即人神经胶质瘤细胞系 M059K 和 M059J，来评估导致 GAA 诱导 H2AX 磷酸化的激酶。免疫荧光显微镜检测γH2AX 的表达。流式细胞术检测γH2AX 和细胞周期。

结果

GAA 以细胞周期依赖性方式诱导 H2AX 磷酸化，并显示 MEC-1 细胞中出现明显的 G0/G1 期阻滞。咖啡因和渥曼青霉素显著抑制 MEC-1 细胞中 GAA 诱导的 H2AX 磷酸化。GAA 诱导 M059K 中 H2AX 磷酸化，但不诱导 M059J 中 H2AX 磷酸化。综上所述，这些数据表明 GAA 处理可单独诱导 MEC-1 和 M059K 细胞中 H2AX 的磷酸化，呈细胞周期依赖性，但不能诱导 M059J 细胞中 H2AX 的磷酸化。在 MEC-1 中观察到明显的 G0/G1 期阻滞。