Although p53-mediated cell-cycle arrest, senescence, and apoptosis remain critical barriers to cancer development, the emerging role of p53 in cell metabolism, oxidative responses, and ferroptotic cell death has been a topic of great interest. Nevertheless, it is unclear how p53 orchestrates its activities in multiple metabolic pathways into tumor suppressive effects. Here, we identified the SAT1 (spermidine/spermine N 1 -acetyltransferase 1) gene as a transcription target of p53. SAT1 is a rate-limiting enzyme in polyamine catabolism critically involved in the conversion of spermidine and spermine back to putrescine. Surprisingly, we found that activation of SAT1 expression induces lipid peroxidation and sensitizes cells to undergo ferroptosis upon reactive oxygen species (ROS)-induced stress, which also leads to suppression of tumor growth in xenograft tumor models. Notably, SAT1 expression is down-regulated in human tumors, and CRISPRcas9-mediated knockout of SAT1 expression partially abrogates p53-mediated ferroptosis. Moreover, SAT1 induction is correlated with the expression levels of arachidonate 15-lipoxygenase (ALOX15), and SAT1-induced ferroptosis is significantly abrogated in the presence of PD146176, a specific inhibitor of ALOX15. Thus, our findings uncover a metabolic target of p53 involved in ferroptotic cell death and provide insight into the regulation of polyamine metabolism and ferroptosis-mediated tumor suppression.SAT1 | p53 | polyamine metabolism | ferroptosis | tumor suppression

Summary Although previous studies indicate that loss of p53-mediated cell cycle arrest, apoptosis, and senescence, does not completely abrogate its tumor suppression function, it is unclear how the remaining activities of p53 are regulated. Here we identified an acetylation site at lysine K98 in mouse p53 (or K101 for human p53). While the loss of K98 acetylation (p53K98R) alone has very modest effects on p53-mediated transactivation, simultaneous mutations at all four acetylation sites (p534KR: K98R+3KR(K117R+K161R+K162R)) completely abolish its ability to regulate metabolic targets such as TIGAR and SLC7A11. Notably, in contrast to p533KR, p534KR is severely defective in suppressing tumor growth in mouse xenograft models. Moreover, p534KR is still capable of inducing the p53-Mdm2 feedback loop but p53-dependent ferroptotic responses are markedly abrogated. Together, these data indicate the critical role of p53 acetylation in ferroptotic responses and its remaining tumor suppression activity.

Mdm2 and Mdmx, both major repressors of p53 in human cancers, are predominantly localized to the nucleus and cytoplasm, respectively. The mechanism by which subcellular localization of Mdmx is regulated remains unclear. In this study, we identify the E3 ligase Peli1 as a major binding partner and regulator of Mdmx in human cells. Peli1 bound Mdmx and and promoted high levels of ubiquitination of Mdmx. Peli1-mediated ubiquitination was degradation-independent, promoting cytoplasmic localization of Mdmx, which in turn resulted in p53 activation. Consistent with this, knockdown or knockout Peli1 in human cancer cells induced nuclear localization of Mdmx and suppressed p53 activity. Myc-induced tumorigenesis was accelerated in Peli1-null mice and associated with downregulation of p53 function. Clinical samples of human cutaneous melanoma had decreased Peli1 expression, which was associated with poor overall survival. Together, these results demonstrate that Peli1 acts as a critical factor for the Mdmx-p53 axis by modulating the subcellular localization and activity of Mdmx, thus revealing a novel mechanism of Mdmx deregulation in human cancers. Peli1-mediated regulation of Mdmx, a major inhibitor of p53, provides critical insight into activation of p53 function in human cancers. .