Abstract
Deoxycytidine kinase (dCK) is a key enzyme in deoxyribonucleoside salvage and the anti-tumor activity for many nucleoside analogs. dCK is activated in response to ionizing radiation (IR)-induced DNA damage and it is phosphorylated on Serine 74 by the Ataxia-Telangiectasia Mutated (ATM) kinase in order to activate the cell cycle G2/M checkpoint. However, whether dCK plays a role in radiation-induced cell death is less clear. In this study, we genetically modified dCK expression by knocking down or expressing a WT (wild-type), S74A (abrogates phosphorylation) and S74E (mimics phosphorylation) of dCK. We found that dCK could decrease IR-induced total cell death and apoptosis. Moreover, dCK increased IR-induced autophagy and dCK-S74 is required for it. Western blotting showed that the ratio of phospho-Akt/Akt, phospho-mTOR/mTOR, phospho-P70S6K/P70S6K significantly decreased in dCK-WT and dCK-S74E cells than that in dCK-S74A cells following IR treatment. Reciprocal experiment by co-immunoprecipitation showed that mTOR can interact with wild-type dCK. IR increased polyploidy and decreased G2/M arrest in dCK knock-down cells as compared with control cells. Taken together, phosphorylated and activated dCK can inhibit IR-induced cell death including apoptosis and mitotic catastrophe, and promote IR-induced autophagy through PI3K/Akt/mTOR pathway.
Highlights
Cell death is a fundamental biological process that has been mediated via intracellular program of biological systems [1,2]
We found that ionizing radiation (IR)-induced cell death can be reversed by reintroduction of Deoxycytidine kinase (dCK), especially by dCK S74E, which improved survival by 84%, suggesting that dCK has positive effects on radioresistance
Necrostatin-1 and Ferrostatin-1 had no effect on IR-induced cell death, while Rapamycin and ZVAD-FMK decreased the rate of cell death by 19% and 36%, and 3-MA increased the rate of cell death by 25% in dCK knock-down cells, indicating apoptosis contributed to the majority of cell death (Figure 1E)
Summary
Cell death is a fundamental biological process that has been mediated via intracellular program of biological systems [1,2]. The execution of cell death needs an orchestrated interplay between some important processes: apoptosis, autophagy, necrosis [3] and mitotic catastrophe [4]. Apoptosis refers to a constellation of characteristic changes leading directly to cell death [5]. Over the past few decades, apoptosis has been widely studied, and radiotherapy strategies targeting apoptosis has become one of the most important cancer treatments [6]. Autophagy is a kind of catabolic process for maintaining cellular homeostasis and supplying substrates for energy generation [7]. Autophagy may protect cells or lead to cell death [8]. Autophagy is considered a double-edged sword in the process of tumor development
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