The critical cancer-fighting protein p53 plays a much broader role in human biology than previously thought, according to a new study by NIEHS researchers. The findings, published August 10 in the journal Nucleic Acids Research, provide unprecedented insights into how the so-called guardian of the genome regulates gene activity across diverse cell types and experimental conditions.

The researchers made these revelations by applying a uniform, rigorous analytical workflow to a large number of previously obtained p53 data sets. The results, which are publicly available through several web resources (see sidebar), may lead to the discovery of novel therapeutic targets for cancer and other diseases.

“The research community currently generates nearly 5,000 p53-related papers per year,” said study author Michael Resnick, Ph.D., head of the NIEHS Chromosome Stability Group. “We reasoned that developing a common, statistically meaningful pipeline of analysis for all studies of all p53-binding sites across the genome, and their associations with gene expression, would be an invaluable tool. Additionally, we expected the analysis to provide direct mechanistic evidence for how p53 functions at its targets in human cells.”

Back row, from left, Leping Li, Ph.D., Biostatistics and Computational Biology (BCB); Resnick; Kevin Gerrish, Ph.D., Molecular Genomics Core Lab; Pierre Bushel, Ph.D., BCB; James Ward, Ph.D., Integrative Bioinformatics Support Group (IBSG); David Fargo, Ph.D., Office of Scientific Computing. Front row, from left, Carl Anderson, Ph.D., Menendez, and Nguyen, all from the Chromosome Stability Group; and Chris Lavender, Ph.D., IBSG. Not pictured: Sara Grimm, Ph.D., and Brian Bennett, Ph.D., both from IBSG, and Yuanyuan Li, Ph.D., BCB. (Photo courtesy of Steve McCaw)

Rigorous workflow

As a transcription factor, p53 binds to DNA to turn genes on and off. To prevent tumor formation, the protein tightly controls cellular responses to various stress signals, such as exposure to carcinogens and ultraviolet radiation. Because mutations that inactivate the p53 gene are found in most human cancers, it has been extensively studied for decades.

Yet significant gaps remain regarding p53’s functions and the genes under its control. One major hurdle is that multiple studies have reported conflicting results, partly due to differences in methodology.

“When we were finishing our first p53 genome-wide binding study in cancer cells, we realized that we were not able to compare our results with other studies,” said study author Daniel Menendez, Ph.D., staff scientist in the Chromosome Stability Group. “It was like comparing apples to many types of fruits.”

To address this challenge, the researchers used a single analysis pipeline to reanalyze 41 data sets from genome-wide human p53 ChIP-seq studies. Chromatin immunoprecipitation followed by high-throughput sequencing, or ChIP-seq is a technique for studying protein-DNA interactions. Sixteen of the data sets also contained messenger RNA expression data, which can be used to assess the activity of genes.

“We are fortunate at NIEHS to have exceptional core laboratories and a large and wonderfully supportive bioinformatics team that could handle the huge analysis,” Menendez said.

Expanding the p53 universe

This comprehensive analysis tripled the number of known cistrome genes, or those directly targeted by p53, to a total of 943. The researchers also identified a core signature that consisted of 28 cistrome genes that are activated by p53, regardless of the cell type or experimental conditions. According to the authors, these core cistrome signature genes could be poised to provide a rapid, coordinated response across biological processes.

A p53 BAER of the 943 cistrome genes. The size of each word indicates the frequency of its overlap across data sets. (Image courtesy of Thuy-Ai Nguyen)

“From a clinical standpoint, the p53 core cistrome signature genes might be used as biomarkers to assess if certain chemotherapeutics are activating p53 in an individual to kill cancer cells,” said lead author Thuy-Ai Nguyen, Ph.D., a research fellow in the Chromosome Stability Group.

Moreover, the study revealed, for the first time, p53 ChIP-seq and gene expression data in normal human lymphocytes, which are immune cells that fight pathogens. The researchers identified 50 potential cistrome genes directly involved in cellular immune processes, providing a new dimension to the broad role that p53 plays in human biology.

Additional findings suggest that p53 is also involved in nervous system function and cardiovascular signaling. “Identifying the universe of genes directly targeted by p53 is clinically relevant, especially since many of these may have therapeutic value,” said Nguyen.

Citation: Nguyen TT, Grimm SA, Bushel PR, Li J, Li Y, Bennett BD, Lavender CA, Ward JM, Fargo DC, Anderson CW, Li L, Resnick MA, Menendez D. 2018. Revealing a human p53 universe. Nucleic Acids Res; doi: 10.1093/nar/gky720 [online 10 August 2018].

(Janelle Weaver, Ph.D., is a contract writer for the NIEHS Office of Communications and Public Liaison.)