PGC-1α coordinates with Bcl-2 to control the cell cycle in U251 cells through reducing ROS

Abstract

B-cell lymphoma 2 (Bcl-2) has a dual function, acting as both an oncogene and an anti-tumor gene. It is well known that Bcl-2 exerts its tumor promoting function through the mitochondrial pathway. However, the mechanism by which it suppresses tumor formation is not well understood. We have previously shown that Bcl-2 inhibits cell cycle progression from the G₀/G₁ to the S phase after serum starvation, and that quiescent Bcl-2 expressing cells maintain a significantly lower level of mitochondrial reactive oxygen species (ROS) than control cells. Based on the fact that ROS mediate cell cycle progression and are controlled by peroxisome proliferator-activated receptor-γ co-activator 1α (PGC-1α), a key molecule induced by prolonged starvation and involved in mitochondrial metabolism, we hypothesized that PGC-1α might be related to the cell cycle function of Bcl-2. In this paper, we show that PGC-1α is upregulated by Bcl-2 overexpression and downregulated following Bcl-2 knockdown or downregulation after serum starvation. However, Bcl-2 is negatively regulated by PGC-1α expression. Further, co-immunoprecipitation (co-IP) experiments showed that PGC-1α protein is co-precipitated with Bcl-2 at the G₀/G₁ phase. Taken together, our results suggest that PGC-1α interacts with Bcl-2 after serum depletion, and that Bcl-2 might recruit PGC-1α to reduce ROS, which in turn delays cell cycle progression in coordination with Bcl-2.

摘要

目的

探究过氧化物酶体增生激活受体γ 协同刺激因子 1α (PGC-1α)和B 细胞淋巴瘤-2(Bcl-2)在调 控细胞周期中的相互关系。

创新点

首次证明在血清饥饿时PGC-1α 负调控Bcl-2,并 且认为Bcl-2 可能通过招募PGC-1α 降低细胞中 的活性氧自由基(ROS)以调节细胞周期。

方法

用蛋白质印迹法(Western blotting)检测了接触 抑制和血清饥饿处理的NIH3T3 过表达Bcl-2 的 细胞中PGC-1α 的表达,并且分别检测了用Bcl-2 和PGC-1α 的小干扰RNA(siRNA)降低U251 细胞(内源性高表达Bcl-2 和PGC-1α)中的Bcl-2 和PGC-1α 的表达,最后用免疫共沉淀(co-IP) 检测了二者的关系。

结论

本实验中用两种细胞同步化的方法(接触抑制和 血清饥饿)处理了Bcl-2 过表达的NIH3T3 细胞 时发现PGC-1α 高表达,用Bcl-2 的siRNA 处理 了U251 细胞时发现PGC-1α 的表达降低,但是 血清饥饿处理了U251 后发现Bcl-2 升高而 PGC-1α 降低,而且PGC-1α 被siRNA 降低后 Bcl-2 反而上升,最后用Bcl-2 抗体免疫共沉淀了 PGC-1α 蛋白,这些结果说明在血清饥饿时 PGC-1α 负调控Bcl-2 行使调节细胞周期的功能。