Cancer Letters

Cancer Letters

Volume 292, Issue 2, 28 June 2010, Pages 228-236
Cancer Letters

Induction of apoptosis by quercetin is mediated through AMPKα1/ASK1/p38 pathway

https://doi.org/10.1016/j.canlet.2009.12.005Get rights and content

Abstract

Effective strategies for cancer prevention and treatment can be identified by understanding the mechanism of apoptotic pathways. In this study, we investigated the regulatory mechanism of quercetin-induced apoptosis through apoptosis signal-regulating kinase (ASK)-1 and mitogen-activated protein kinase pathways. Our results showed that quercetin increased apoptotic cell death through reactive oxygen species (ROS) generation and was responsible for ASK1 activation. Increasing ASK1 activity was accompanied by p38 activation. Interestingly, AMP-activated protein kinase (AMPK) seemed to be a critical controller of quercetin-regulated ASK1/p38 activation. Blocking AMPKα1 activity using Compound C, a synthetic inhibitor or siRNA showed that quercetin-activated ASK1 could not stimulate p38 activity. Thus, we suggested that quercetin-exerted apoptotic effects involve ROS/AMPKα1/ASK1/p38 signaling pathway, and AMPKα1 is a necessary element for apoptotic event induced by ASK1.

Introduction

Apoptosis is a programmed cell death process that controls normal development and homeostasis in organisms [1], [2]. The loss of apoptotic control contributes to the survival of tumor cells, and the enhancement of cancer cell apoptosis is one approach of controlling cancer by anticancer agents. Although there seems to be remarkable progresses of the detailed mechanism of the mitochondrial cytochrome c-induced apoptosis, the overall regulatory signaling pathways whereby apoptosis is targeted by anticancer agents have not been clearly presented [3], [4], [5]. Providing the detailed pathways leading to apoptosis is an important goal for establishing effective tumor prevention and treatment strategies using chemopreventive materials such as quercetin. One of the well-known intracellular signaling pathways for apoptosis is the kinase cascade, which has been identified as a transducing pathway of apoptotic signals initiated by outside stimuli, and the mitogen-activated protein (MAP) kinases and their upstream kinases such as MAP kinase kinases [6], [7]. Apoptosis signal-regulating kinase (ASK)-1 is a member of the ROS-sensitive MAP kinase kinases and it acts as a redox sensor of cells [6], [8], [9], [10]. ASK1 is commonly regulated through protein–protein interactions, and numerous proteins have been shown to bind ASK1. The binding of ASK1 to TNF receptor-associated factor (TRAF) or death domain-associated protein (Daxx) stimulates ASK1 function, whereas the ASK1 inhibitory proteins such as thioredoxin (Trx) downregulate pro-apoptotic activity of ASK1 [11], [12], [13], [14], [15].

In this study, we investigated the involvement of ASK1 in the apoptotic process of cancer cells treated with a chemopreventive agent, quercetin. We previously reported that quercetin-induced apoptosis through the generation of ROS, which was responsible for the activation of AMPKα1, and was found to be an indispensable element of apoptosis [16]. Here, we demonstrated that quercetin strongly generated ROS and that this oxidative stress induced apoptosis through activation of AMPKα1 and ASK1, which are critical upstream signals for p38 MAP kinase activation in MCF-7 breast cancer cells. Thus, we suggest that AMPKα1 is a master regulator of ASK1-mediated apoptotic pathway.

Section snippets

Cell culture and reagents

MCF-7 cells were obtained from the American Type Culture Collection (Manassas, VA, USA) and grown in RPMI 1640 medium containing 10% fetal bovine serum (Gibco, Grand Island, NY, USA) at 37 °C in a 5% CO2 atmosphere. Quercetin, hydrogen peroxide, and N-acetyl-cystein were purchased from Sigma (St. Louis, MO, USA). Specific antibodies that recognize the phosphorylated forms of AMPKα1Thr172 and ACC were obtained from Cell Signaling Technology (Danvers, MA, USA) and p-ASK1Ser83, ASK1 and p-ERK were

Quercetin-generated ROS in MCF-7 breast cancer cells

To examine whether quercetin stimulates ROS generation in MCF-7 breast cancer cells, we measured the intracellular ROS levels after treatment of quercetin (0–400 μM) at the indicated time series. As shown in Fig. 1A, quercetin continuously increased ROS levels from 30 min to 9 h after treatment, and the levels were maintained at concentrations above 200 μM. These effects were completely blocked by combined treatment with NAC, a ROS scavenger (Fig. 1B). We also observed intracellular ROS by

Discussion

Our results demonstrate that activation of AMPKα1 is required for ASK1 to induce apoptosis. Additionally, a signaling pathway of AMPKα1/ASK1/p38 is responsible for MCF-7 cells to undergo apoptosis initiated by ROS when the cancer cells were treated with quercetin. Previously, we have shown that quercetin regulates cancer cell proliferation and apoptosis through AMPKα1/cyclooxygenase-2 (COX-2) pathway [16]. AMPK is known to phosphorylate several downstream proteins that control cell growth and

Conflicts of interest

None declared.

Acknowledgments

This work was supported by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (No. R01-2008-000-20131-0).

References (27)

  • M.S. Ricci et al.

    Chemotherapeutic approaches for targeting cell death pathways

    Oncologist

    (2006)
  • M.L. Tan et al.

    Programmed cell death pathways and current antitumor targets

    Pharm. Res.

    (2009)
  • A. Matsuzawa et al.

    Molecular mechanisms of the decision between life and death: regulation of apoptosis by apoptosis signal-regulating kinase 1

    J. Biochem.

    (2001)
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