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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 KrasG12D-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.

Epstein-Barr virus Latent Membrane Protein 2A (LMP2A) is expressed in EBV-infected B cells in the germinal center, a site of significant apoptosis induced by engagement of Fas on activated B cells. Signals from the B cell receptor (BCR) protect germinal center B cells from Fas-mediated apoptosis, and since LMP2A is a BCR mimic, we hypothesized that LMP2A would also protect B cells from Fas-mediated apoptosis. Surprisingly, latently-infected human and murine B cell lines expressing LMP2A were more sensitive to Fas-mediated apoptosis, as determined by increases in Annexin-V staining, and cleavage of caspase-8, −3 and PARP. Additional studies show that LMP2A-expressing B cell lines demonstrate a Lyn- and Syk-dependent increase in sensitivity to Fas-mediated apoptosis, due to an LMP2A-dependent enhancement in Fas expression. These findings demonstrate the ability for LMP2A to directly increase a pro-apoptotic molecule and have implications for EBV latency as well as the treatment of EBV-associated malignancies.

p53 is a crucial tumour suppressor that responds to diverse stress signals by orchestrating specific cellular responses, including transient cell cycle arrest, cellular senescence and apoptosis, which are all processes associated with tumour suppression. However, recent studies have challenged the relative importance of these canonical cellular responses for p53-mediated tumour suppression and have highlighted roles for p53 in modulating other cellular processes, including metabolism, stem cell maintenance, invasion and metastasis, as well as communication within the tumour microenvironment. In this Opinion article, we discuss the roles of classical p53 functions, as well as emerging p53-regulated processes, in tumour suppression.

summary Background: Oral cavity and in particular oral tongue cancers occur with a rising incidence in younger patients often lacking the typical risk factors of tobacco use, alcohol use, and human papilloma virus (HPV) infection. Their prognosis when treated with chemoradiation has not been well studied and responsible risk factors remain elusive. A viral etiology (other than HPV) has been hypothesized. Methods: First we analyzed outcomes from 748 head and neck cancer patients with locoregionally advanced stage tumors treated with curative-intent chemoradiation by anatomic site. Second, we analyzed seven oral tongue (OT) tumors from young, non-smokers/non-drinkers for the presence of viral mRNA using short-read massively-parallel sequencing (RNA-Seq) in combination with a newly-developed digital subtraction method followed by viral screening and discovery algorithms. For positive controls we used an HPV16-positive HNC cell line, a cervical cancer, and an EBV-LMP2A transgene lymphoma. Results: Younger patients with oral cavity tumors had worse outcomes compared to non-oral cavity patients. Surprisingly none of the seven oral tongue cancers showed significant presence of viral transcripts. In positive controls the expected viral material was identified. Conclusion: Oral cavity tumors in younger patients have a poor prognosis and do not appear to be caused by a transcriptionally active oncovirus.

The mechanisms by which the p53 tumor suppressor acts remain incompletely understood. To gain new insights into p53 biology we used high throughput sequencing to analyze global p53 transcriptional networks in primary mouse embryo fibroblasts in response to DNA damage. Chromatin immunoprecipitation sequencing reveals 4785 p53 bound sites in the genome located near 3193 genes involved in diverse biological processes. RNA sequencing analysis shows that only a subset of p53 bound genes is transcriptionally regulated yielding a list of 432 p53 bound and regulated genes. Interestingly we identify a host of autophagy genes as direct p53 target genes. While the autophagy program is regulated predominantly by p53 the p53 family members p63 and p73 contribute to activation of this autophagy gene network. Induction of autophagy genes in response to p53 activation is associated with enhanced autophagy in diverse settings and depends on p53 transcriptional activity. While p53 induced autophagy does not affect cell cycle arrest in response to DNA damage it is important for both robust p53 dependent apoptosis triggered by DNA damage and transformation suppression by p53. Together our data highlight an intimate connection between p53 and autophagy through a vast transcriptional network and indicate that autophagy contributes to p53 dependent apoptosis and cancer suppression. © 2013 by Cold Spring Harbor Laboratory Press.

The link between EBV infection and Burkitt lymphoma (BL) is strong, but the mechanism underlying that link has been elusive. We have developed a mouse model for EBV-associated BL in which LMP2A, an EBV latency protein, and MYC are expressed in B cells. Our model has demonstrated the ability of LMP2A to accelerate tumor onset, increase spleen size, and bypass p53 inactivation. Here we describe the results of total gene expression analysis of tumor and pretumor B cells from our transgenic mouse model. Although we see many phenotypic differences and changes in gene expression in pretumor B cells, the transcriptional profiles of tumor cells from LMP2A/λ-MYC and λ-MYC mice are strikingly similar, with fewer than 20 genes differentially expressed. We evaluated the functional significance of one of the most interesting differentially expressed genes, Egr1, and found that it was not required for acceleration of tumor onset by LMP2A. Our studies demonstrate the remarkable ability of LMP2A to affect the pretumor B-cell phenotype and tumorigenesis without substantially altering gene expression in tumor cells.

We measured anti-Haemophilus influenzae type a capsular polysaccharide serum immunoglobulin G antibodies in cord blood sera from Mexican (n = 68) and Chilean mothers (n = 72) by enzyme-linked immunosorbent assay. Measurable antibodies were found in 79.3% of samples. Immunoglobulin G antibodies correlated with serum bactericidal activity (r = 0.66). This enzyme-linked immunosorbent assay can be used for the evaluation of adaptive immune responses to Haemophilus influenzae type a and serosurveillance studies in populations at risk.