The tumor suppressor p53, which is sometimes called 'guardian of the genome' is a potent transcription factor that is activated in response to diverse stresses and environmental insults to restrain the emergence of transformed cells with genetic instabilities. But activation of p53 target genes is highly tissue specific. Elevated wild-type p53 activity is extremely deleterious to some tissues while others are seemingly unbothered by it.
In a paper published in the September 10, 2020, online issue of the Proceedings of the National Academy of Sciences (PNAS), Sydney Moyer and her colleagues from MD Anderson Cancer Center have examined why and how these vast differences exist among tissues exposed to p53 activation.
Speaking to BioWorld Science, the lead author of the study, Moyer, said that "we feel that one cannot combat these 'side effects' of elevated p53 without understanding their origin. We looked exclusively at normal tissues in this study and we believe that any aspect that makes a tissue unique could potentially play a role in shaping the physiological response to p53 pathway activation." Moyer is a PhD candidate in the laboratory of Prof. Guillermina Lozano at the MD Anderson Cancer Center. Lozano focuses on understanding the signals that regulate the p53 pathway and the consequences of expressing wild-type or mutant p53.
For their study, Moyer and her team used a genetically engineered mouse model of p53 activation, mediated by conditional deletion of Mdm2. In normal cells, p53 is kept at low levels by murine double minute 2 (Mdm2), a ubiquitin ligase. They then selected five tissues (pancreas, heart, kidney, ovary and intestine) and conducted RNA-sequencing analysis and a comparison with existing ChIP-sequencing data to understand the physiological response to p53 activation. "The biggest challenges we faced had to do with using an in vivo model --the same thing that makes our study so valuable. We were studying the tissue-specificity of p53 in the pancreas, kidney, ovary, heart and small intestine of our mice. However, all of these tissues are within the same animal and thus interacting with each other," said Moyer.
While the majority of p53 target genes were upregulated in the study, a small percentage of differentially expressed genes in the heart, ovary and pancreas were direct p53 targets and downregulated in the genetically modified mice, suggesting an in vivo role for p53 repression. The researchers were able to identify several tissue-specific and seven common p53 target genes by comparison of tissue-specific bulk RNA-sequencing data with existing sequencing data obtained from mouse embryonic fibroblasts. This led to the surprising result of a response in the pancreas that was the result of increased p53 expression in other organs.
After full-body activation of p53, the pancreas displayed a metaplastic phenotype, that is, they changed their cell fates in response to high levels of p53 and inflammation.
But "activation in the pancreas alone does not result in metaplasia or inflammation," Moyer said. "There is something unique about elevating p53 in an entire organism that led to this phenotype. It took us a long time to realize this pancreatic phenotype was non-cell-autonomous, yet these striking differences is why the struggles associated with animal models are so valuable -- everything acts together as a complicated and intricate system."
The most surprising result, Moyer said, was the list of genes that showed up in the p53 transcriptional signature. This list does not include p21, the gene historically used as the gold standard readout for p53 transcriptional activation. p53 was seen to bind to the promoter of seven genes (Ccng1, Eda2r, Gtse1, Mdm2, Polk, Psrc1 and QZfp365) which were transcriptionally upregulated in multiple tissues of two different mouse strains and in response to p53 activation genetically and exogenously. The authors propose that they serve as a universal signature for transcriptional activation by wild-type p53.
Moyer hopes that this work would be an important resource to all those studying p53, whether in normal or cancerous tissues. Understanding the tissue specificity of the p53 transcriptional response can lead to better drugs. p53 reactivation is a potentially promising concept in cancer as several cancers exhibit mutant or inactive forms of p53 that contribute to carcinogenesis, progression and poor prognosis. However, p53 reactivation can have lethal effects.
"Specifically in terms of practical applications, more and more people are exploring the usefulness of p53 pathway-reactivating drugs. While these drugs have promising effects in the tumors themselves, the side effects associated with these drugs have proved to be a problem for finding safe treatment regimens," Moyer said. "Now, for example, if one was studying intestinal toxicities in response to these drugs, they could use our sequencing data to find potential downstream targets of p53 that may be contributing to these phenotypes." This new knowledge may contribute to improvement of p53 as a viable drug target.
Moyer is now planning to explore the new downstream p53 target genes on a tissue-specific basis. The group has already generated some knockout mouse models of these target genes of interest. Their goal is to use these knockouts to determine which phenotype that gene was contributing to. Moyer explained, "For example, if we knock out gene X in a mouse and then p53 activation no longer results in the small intestinal defects that we observed previously, then gene X likely played a strong role in bringing that defect to fruition. Our study beautifully highlights how much is left to learn about wild-type or natural p53 activity. Even though p53 has been highly studied for decades, we still have only scratched the surface." (Moyer, S.M. et al. Proc Natl Acad Sci U S A 2020, Advanced publication).