SUMMARY How cancer cells respond to nutrient deprivation remains poorly understood. In certain cancer cells, deprivation of cystine induces a non-apoptotic, iron-dependent form of cell death termed ferroptosis. Recent evidence suggests that ferroptosis sensitivity may be modulated by the stress-responsive transcription factor and canonical tumor suppressor protein p53. Using CRISPR/Cas9 genome editing, small-molecule probes, and high-resolution, time-lapse imaging, we find that stabilization of wild-type p53 delays the onset of ferroptosis in response to cystine deprivation. This delay requires the p53 transcriptional target CDKN1A (encoding p21) and is associated with both slower depletion of intracellular glutathione and a reduced accumulation of toxic lipid-reactive oxygen species (ROS). Thus, the p53-p21 axis may help cancer cells cope with metabolic stress induced by cystine deprivation by delaying the onset of non-apoptotic cell death.

The p53 gene is mutated in over half of all cancers, reflecting its critical role as a tumor suppressor. Although p53 is a transcriptional activator that induces myriad target genes, those p53-inducible genes most critical for tumor suppression remain elusive. Here, we leveraged p53 ChIP-seq (chromatin immunoprecipitation [ChIP] combined with high-throughput sequencing) and RNA-seq (RNA sequencing) data sets to identify new p53 target genes, focusing on the noncoding genome. We identify Neat1, a noncoding RNA (ncRNA) constituent of paraspeckles, as a p53 target gene broadly induced by mouse and human p53 in different cell types and by diverse stress signals. Using fibroblasts derived from Neat1 −/− mice, we examined the functional role of Neat1 in the p53 pathway. We found that Neat1 is dispensable for cell cycle arrest and apoptosis in response to genotoxic stress. In sharp contrast, Neat1 plays a crucial role in suppressing transformation in response to oncogenic signals. Neat1 deficiency enhances transformation in oncogene-expressing fibroblasts and promotes the development of premalignant pancreatic intraepithelial neoplasias (PanINs) and cystic lesions in Kras G12D -expressing mice. Neat1 loss provokes global changes in gene expression, suggesting a mechanism by which its deficiency promotes neoplasia. Collectively, these findings identify Neat1 as a p53-regulated large intergenic ncRNA (lincRNA) with a key role in suppressing transformation and cancer initiation, providing fundamental new insight into p53-mediated tumor suppression.

SUMMARY The p53 transcription factor is a critical barrier to pancreatic cancer progression. To unravel mechanisms of p53-mediated tumor suppression, which have remained elusive, we analyzed pancreatic cancer development in mice expressing p53 transcriptional activation domain (TAD) mutants. Surprisingly, the p5353,54 TAD2 mutant behaves as a “super-tumor suppressor”, with an enhanced capacity to both suppress pancreatic cancer and transactivate select p53 target genes, including Ptpn14. Ptpn14 encodes a negative regulator of the Yap oncoprotein and is necessary and sufficient for pancreatic cancer suppression, like p53. We show that p53 deficiency promotes Yap signaling and that PTPN14 and TP53 mutations are mutually exclusive in human cancers. These studies uncover a p53-Ptpn14-Yap pathway that is integral to p53-mediated tumor suppression.

Although TP53 is the most commonly mutated gene in human cancers, the p53-dependent transcriptional programs mediating tumor suppression remain incompletely understood.Here, to uncover critical components downstream of p53 in tumor suppression, we perform unbiased RNAi and CRISPR/Cas9-based genetic screens in vivo. These screens converge upon the p53-inducible gene Zmat3, encoding an RNA-binding-protein, and we demonstrate that ZMAT3 is an important tumor suppressor downstream of p53 in mouse Kras G12D -driven lung and liver cancers and human carcinomas. Integrative analysis of the ZMAT3 RNA-binding landscape and transcriptomic profiling reveals that ZMAT3 directly modulates exon inclusion in transcripts encoding proteins of diverse functions, including the p53 inhibitors MDM4 and MDM2, splicing regulators, and components of varied cellular processes. Interestingly, these exons are enriched in NMD signals, and, accordingly, ZMAT3 broadly affects target transcript stability. Collectively, these studies reveal ZMAT3 as a novel RNA-splicing and homeostasis regulator and key component of p53-mediated tumor suppression.We thank Laurakay Bruhn, Steven Altschuler, Ben Borgo, Peter Sheffield and Carsten Carstens of Agilent Inc. for oligonucleotide synthesis and helpful discussions. We thank Lin He for the Eμ-Myc lymphoma cells, Andreas Strasser and Ana Janic for the Zmat3 null MEFs, and Julien Sage and Aaron Gitler for critical reading of the manuscript.

The mechanisms by which TP53, the most frequently mutated gene in human cancer, suppresses tumorigenesis remain unclear. p53 modulates various cellular processes, such as apoptosis and proliferation, which has led to distinct cellular mechanisms being proposed for p53-mediated tumor suppression in different contexts. Here, we asked whether during tumor suppression p53 might instead regulate a wide range of cellular processes. Analysis of mouse and human oncogene-expressing wild-type and p53-deficient cells in physiological oxygen conditions revealed that p53 loss concurrently impacts numerous distinct cellular processes, including apoptosis, genome stabilization, DNA repair, metabolism, migration, and invasion. Notably, some phenotypes were uncovered only in physiological oxygen. Transcriptomic analysis in this setting highlighted underappreciated functions modulated by p53, including actin dynamics. Collectively, these results suggest that p53 simultaneously governs diverse cellular processes during transformation suppression, an aspect of p53 function that would provide a clear rationale for its frequent inactivation in human cancer.