Abstract C46: DNA damage-induced autophagy is required for efficient DNA repair and cell cycle progression

Abstract

Abstract DNA damage is known to induce cell cycle arrest followed by DNA damage repair (DDR) or apoptosis depending upon the extent of the DNA damage. DNA damage has also been shown to induce autophagy, a catabolic process involving the degradation of cellular organelles and proteins, to provide the energy needs for critical cellular processes. Excessive or defective autophagy can mediate cell death as a consequence of the degradation of vital organelles and/or accumulation of toxic cellular content. However, in many tumor cells, autophagy can be an acquired mechanism for cell survival that has evolved to provide higher energy and to remove the non-functional mitochondria for more efficient cellular function. In this study we show that ionizing radiation-induced DNA damage can induce autophagy in prostate cancer cells. LC3 lipidation (LC3 II), a classical marker for autophagy signaling was observed following irradiation. The steady state levels of LC3 II were not affected, however there was increased accumulation of LC3 II following irradiation in the presence of chloroquine, an inhibitor of autolysosomal formation and acidification. This observation suggested that autophagy is induced by DNA damage. Cell death analysis showed that autophagy served as a protective mechanism since inhibiting autophagy by pharmacological inhibitors, such as 3-MA and chloroquine, as well as genetic knockdown of ATG7 and LAMP2, sensitized prostate cancer cells to irradiation induced cell death, as shown by increased sub-G1 DNA content and reduced clonogenic survival. In autophagy-deficient cells, there were increased γH2AX and 53BP1 foci compared to autophagy-proficient cells, an indication that DDR is inhibited upon inhibiting autophagy. Moreover, comet assay revealed increased tail length in autophagy-deficient cells indicating accumulation of damaged DNA. Collectively, these observations suggest that DNA damage-induced autophagy can facilitate DDR signaling and cell cycle progression while autophagy inhibition can inhibit DDR signaling resulting in increased apoptosis. Investigating cell cycle checkpoints and activation of apoptosis in autophagy-deficient cells could help to understand the role of autophagy in mediating DDR and cell cycle progression following irradiation. Autophagy, similar to DNA repair may play a critical function in the cellular response to ionizing radiation, an oxidizing agent that damages not only DNA but other cellular molecules and organelles. These findings could help to identify specific autophagy inhibitors for sensitizing tumor cells to irradiation leading to increased tumor cell death. Citation Format: Kamini Singh, Sayer R. Al-Harbi, Akwasi Agyeman, Janet A. Houghton, Warren D. Heston, Alex Almasan. DNA damage-induced autophagy is required for efficient DNA repair and cell cycle progression [abstract]. In: Proceedings of the AACR Special Conference on Advances in Prostate Cancer Research; 2012 Feb 6-9; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2012;72(4 Suppl):Abstract nr C46.