Proline oxidase activates both intrinsic and extrinsic pathways for apoptosis: the role of ROS/superoxides, NFAT and MEK/ERK signaling

Abstract

Proline oxidase (POX), often considered a 'housekeeping enzyme' might play an important role in apoptosis. We have shown that POX generated proline-dependent reactive oxygen species (ROS), specifically superoxide radicals, and induced apoptosis through the mitochondrial (intrinsic) pathway. In our current report, we used DLD-1 colorectal cancer cells stably transfected with the POX gene under the control of a tetracycline-inducible promoter and found POX-stimulated expression of tumor necrosis factor-related apoptosis inducing ligand (TRAIL), DR5 and cleavage of caspase-8. Importantly, apoptosis measured by flow cytometry was partially inhibited by Z-IETD-FMK, a specific inhibitor of caspase-8. These findings suggest that the extrinsic (death receptor) pathway also is activated by POX. Furthermore, the mechanism of this effect on the extrinsic pathway, specifically, the induction of TRAIL by POX, may be mediated by NFAT transcription factors. Additionally, POX expression also dramatically decreased phosphorylation of MEK and ERK, and the decrease was partially reversed by expression of manganese superoxide dismutase (MnSOD). Overexpression of constitutively active form of MEK, acMEK, partially blocked POX-induced apoptosis. These findings suggest the involvement of MEK/ERK signaling and further confirm the role of ROS/superoxides in POXinduced apoptosis. Combined with previously published data, we conclude that POX may induce apoptosis through both intrinsic and extrinsic pathways and is involved in nuclear factor of activated T cells (NFAT) signaling and regulation of the MEK/ERK pathway. It is suggested that, as a nutrition factor, POX may modulate apoptosis signals induced by p53 or other anti-cancer agents and enhance apoptosis in stress situations.

2-Deoxyglucose combined with wild-type p53 overexpression enhances cytotoxicity in human prostate cancer cells via oxidative stress

“…These results also suggested that both O 2 •-and hydroperoxides were capable of contributing to the oxidative stress and cytotoxicity seen during 2DG and Adp53 treatment. These results are also consistent with the previous observations that p53-dependent apoptosis was preceded by the induction of the enzyme proline oxidase and increased respiration presumably leading to the increased generation of O 2 •-and hydroperoxides [9,10,20,36].…”

“…Recent reports also documented that the combination of ionizing radiation and adenoviral p53 gene therapy increased radiosensitivity of both p53-mutant and wild type cancer cells [7;8;35]. In addition several reports have suggested that p53 over expression can induce oxidative stress and alterations in respiration in cancer cells [1,2,9,10,20,36] and that 2DG can enhance cell killing in malignant cells by agents that induce metabolic oxidative stress [3,4,37]. Finally, several investigators have reported that androgenindependent prostate cancer cells have a high frequency of p53 mutations [38;39].…”

“…The biochemical rationale for this combination to enhance cancer cell killing was based on previous results in other human cancer cells suggesting that 2DG would inhibit glucose metabolism leading to a reduction in intracellular pyruvate and NADPH [3,4,37], limiting the capacity of the tumor cells to metabolize hydroperoxides [4,15,25], and p53 would increase metabolic hydroperoxide production [1,2,20,36] leading to enhanced oxidative stress. As shown in our results the combination of 2DG and Adp53 lead to over expression of p53 (Figure 1), increased clonogenic cell killing (Figure 2 and Figure 3), and increased oxidative stress that appeared to be mediated by increased steady-state levels of hydroperoxides including H 2 O 2 (Fig 4-Fig 6) in prostate cancer cells.…”

“…Recently p53-induced apoptosis in cancer cells has been shown to occur through induction of p53-induced genes (PIGs) and the tumor suppressive effects of some PIGs are thought to be the result of increased formation of reactive oxygen species and mitochondrial respiration (ROS) (i.e. O 2 •-and H 2 O 2 ), which in turn lead to oxidative stress [1,2,9,10,20]. Since 2DG would be expected to inhibit the formation of pyruvate and NADPH, which function in the detoxification pathways of H 2 O 2 [3,4,15], we hypothesized that the combination of 2DG and wt p53 over expression would lead to increased steady-state levels of ROS and enhanced cell killing by oxidative stress.…”

Proline modulates the intracellular redox environment and protects mammalian cells against oxidative stress

“…P5C is then converted to glutamate in a NAD + -dependent reaction catalyzed by P5CDH [2]. Phang and others have shown that in mammalian cells PRODH expression is activated by p53 with proline oxidation leading to increases in intracellular reactive oxygen species (ROS) and induction of cellular apoptosis in various cancer cell lines [3][4][5][6][7][8][9]. Thus, proline serves as a prooxidant in apoptotic signaling via PRODH activity which may be an important mechanism for reducing carcinogenesis [6].…”