Role of PADI4 as a key epigenetic regulator of the p53 pathway and tumor suppression

项目总结

PROJECT SUMMARY TP53 is the most frequently mutated gene in human cancer. While it is well understood that the ability of p53 to act as a transcription factor is required for tumor suppression, the key target genes downstream of p53 required for tumor suppression are still incompletely understood. Moreover, attempts at drugging p53 have proven ineffective, generating a need to understand downstream players in the tumor suppressive pathway to develop novel therapies. The Murphy lab has characterized three p53 variants termed “hypomorphs” that are minimally impaired for p53 transcriptional function but have increased cancer-risk. These three p53 hypomorphs are impaired for transactivation of only a few target genes, with the top target gene impaired being peptidyl-arginine deiminase 4 (PADI4). PADI4 is a regulator of histone modification via citrullination, which is the process of deiminating unmodified or mono-methylated arginine to the non-natural amino acid citrulline. Histone citrullination is predicted to regulate gene transcription and is known in some cases to decondense chromatin. PADI4 dependent citrullination is also involved in immune cell activation and recruitment. Previous reports have suggested PADI4 may act as an oncogene or a tumor suppressor, and the reasons underlying this controversy remain unclear. Additionally, mechanistic studies on PADI4 and histone citrullination in cancer are lacking. TCGA analysis reveals that PADI4 is downregulated or mutated in multiple human cancers, including mutations in 8% of melanoma, suggesting that this gene is a tumor suppressor. I have found that PADI4 levels increase in response to genotoxic injury in multiple tissues, but this is abolished in p53Y107H and PADI4 KO mice. PADI4 also suppresses the proliferation of both p53 wild type (WT) and p53-null cancer cells. RNA-seq analysis of cells treated with the p53 stabilizer Nutlin-3a, +/- si-PADI4, reveals PADI4 to be required for transcriptional activation of a subset of p53 target genes. Collectively, these data suggest that PADI4 as a novel epigenetic regulator in cancer and may play a key role in p53-mediated tumor suppression. I hypothesize that PADI4 contributes to tumor suppression through interaction with p53, and through regulating chromatin organization by citrullinating histones and opening chromatin at targeted loci. To test this hypothesis, I propose two aims. In the first aim, I will establish the direct transcriptional targets of PADI4 in p53 WT and p53 null melanoma cells. I will also formally test the hypothesis that PADI4 increases histone citrullination to induce changes in chromatin accessibility. In the second aim, I will assess the tumor suppressive ability of PADI4 and PADI4 target genes in xenograft tumor growth studies. Finally, whereas PADI4 has a clear transcriptional role, PADI4 is also known to enhance antigen recognition, but whether this contributes to tumor suppression is unknown. I will test this hypothesis. In sum, this proposal seeks to uncover new information about an exciting but poorly understood epigenetic modifier and its role in tumor suppression and the p53 pathway.