Stress-induced activation of the p53 tumor suppressor in leukemia cells and normal lymphocytes requires mitochondrial activity and reactive oxygen species

Abstract

The p53 system is highly stress sensitive and integrates diverse intracellular signals in a complex and poorly defined manner. We report on the high dependence of stress-induced p53 activation on mitochondrial activity. Down-regulation of mitochondrial transmembrane potential (MTMP) by inhibitors of electron transport (rotenone, thenoyltrifluoroacetone (TTFA)) and adenosine triphosphate (ATP) synthesis (oligomycin) prevented stress-induced p53 protein accumulation and abrogated p53-dependent apoptosis in a wild-type p53 leukemia cell line MOLT-3, in primary leukemia cells and in normal T lymphocytes. Using genomewide gene expression analysis, stressinduced up-regulation of the p53 transcriptional targets and their specific inhibition by oligomycin has been demonstrated. Oligomycin did not impair p53-independent apoptosis and caused only a slight reduction of intracellular ATP levels. Reactive oxygen species (ROS) localized to mitochondria decreased in the presence of oligomycin, and stress-induced p53 activation showed strong ROS sensitivity both in leukemic and normal cells. These observations identify mitochondrial activity, described by MTMP and ROS levels, as a critical intracellular determinant of the p53 stress sensitivity and suggest potential implications of this linkage in the mechanisms of chemoresistance of acute leukemia cells. IntroductionSublethal damage or impairment of cellular functions result in cellular responses, which include activation of repair mechanisms, cell-cycle checkpoints, and induction of endogenous apoptotic programs. Cellular stress response often implicates activation of the proapoptotic p53-dependent machinery, which determines the threshold level of cellular stress able to trigger apoptosis. 1,2 p53 is regulated mainly at the posttranscriptional level and its protein level in the absence of stress is consistently low due to a high rate of p53 protein degradation via the proteasome pathway. Stressinduced signaling results in modification of the p53 protein either by phosphorylation or acetylation and in the resistance of p53 to proteasome degradation. 3,4 A further basic regulatory mechanism of the p53 system is the interaction of p53 with its negative regulator Mdm2 protein, which acts as a p53-specific E3 ligase and is, in turn, transcriptionally regulated by p53. 5 The importance of p53/Mdm2 interactions is underscored by the variety of mechanisms used by a cell to disrupt these interactions in response to stress, including posttranslational modifications, subcellular redistribution, inhibition of Mdm2 activity, and its transcription. 5 More recently, nicotinamide adenine dinucleotide phosphate (NADH) quinone oxidoreductase 1 (NQO1) was demonstrated to regulate p53 stability. 6 NQO1 was suggested to interact physically with p53 and to stabilize p53 through Mdm2-independent mechanisms. 7,8 Intracellular p53 accumulation is the hallmark of the p53 activation pathway and results in p53 protein complexation, nuclear localization of p53 tetramers, and induction of their transcri...

Pyrimidine biosynthesis links mitochondrial respiration to the p53 pathway

“…Previous evidence suggests that mitochondrial ETC could play a role in stress-induced activation of p53 (13)(14)(15)(16). However, in the present study we found that the inhibition of mitochondrial ETC does not necessarily generate p53-activating signals.…”

“…However, the role of mitochondrial ETC activity in the induction of p53 response remains ambiguous. It was suggested that mitochondrial activity could be required for the stress-induced activation of p53, as inhibitors of complexes I and V mitigate the response to etoposide treatment (14) and inhibitors of complex III interfere with the activation of p53 after treatment with cisplatin (15). On the other hand, it was noticed that certain ETC inhibitors produce a cell senescence phenotype associated with a modest activation of p53, leading to the suggestion that the reduced mitochondrial membrane potential (MMP) could initiate the p53 response (16).…”

Dual inhibition of topoisomerases enhances apoptosis in melanoma cells

“…Also, generation of reactive oxygen species (ROS) and resulting oxidative stress often accompanies disturbed DNA integrity in cells exposed to topoisomerase inhibitors [28,29] and to this end our data indicating significantly elevated levels of oxidative stress in treated cells agree with this scenario. ROS may activate an entire array of stress signaling mediators including stress kinases p38 and SAPK/JNK [30,31].…”

Feedback Control of p53 Translation by REDD1 and mTORC1 Limits the p53-Dependent DNA Damage Response

“…Ϫ/Ϫ cells, as expected (16); however, this treatment did not normalize p53 levels in Redd1 Ϫ/Ϫ cells and instead may have further increased the relative difference in p53 levels between Redd1 Ϫ/Ϫ and wild-type cells (Fig. 4C).…”