Aflatoxin B1 and Epstein-Barr virus-induced CCL22 expression stimulates B cell infection.

Epstein-Barr Virus (EBV) infects more than 90% of the adult population worldwide. EBV infection is associated with Burkitt lymphoma (BL) though alone is not sufficient to induce carcinogenesis implying the involvement of co-factors. BL is endemic in African regions faced with mycotoxins exposure. Exposure to mycotoxins and oncogenic viruses has been shown to increase cancer risks partly through the deregulation of the immune response. A recent transcriptome profiling of B cells exposed to aflatoxin B1 (AFB1) revealed an upregulation of the Chemokine ligand 22 (CCL22) expression although the underlying mechanisms were not investigated. Here, we tested whether mycotoxins and EBV exposure may together contribute to endemic BL (eBL) carcinogenesis via immunomodulatory mechanisms involving CCL22. Our results revealed that B cells exposure to AFB1 and EBV synergistically stimulated CCL22 secretion via the activation of Nuclear Factor-kappa B pathway. By expressing EBV latent genes in B cells, we revealed that elevated levels of CCL22 result not only from the expression of the latent membrane protein LMP1 as previously reported but also from the expression of other viral latent genes. Importantly, CCL22 overexpression resulting from AFB1-exposure in vitro increased EBV infection through the activation of phosphoinositide-3-kinase pathway. Moreover, inhibiting CCL22 in vitro and in humanized mice in vivo limited EBV infection and decreased viral genes expression, supporting the notion that CCL22 overexpression plays an important role in B cell infection. These findings unravel new mechanisms that may underpin eBL development and identify novel pathways that can be targeted in drug development.

Interplay between the Epigenetic Enzyme Lysine (K)-Specific Demethylase 2B and Epstein-Barr Virus Infection.

The histone modifier lysine (K)-specific demethylase 2B (KDM2B) plays a role in the differentiation of hematopoietic cells, and its expression appears to be deregulated in certain cancers of hematological and lymphoid origins. We have previously found that the KDM2B gene is differentially methylated in cell lines derived from Epstein-Barr virus (EBV)-associated endemic Burkitt lymphoma (eBL) compared with that in EBV-negative sporadic Burkitt lymphoma-derived cells. However, whether KDM2B plays a role in eBL development has not been previously investigated. Oncogenic viruses have been shown to hijack the host cell epigenome to complete their life cycle and to promote the transformation process by perturbing cell chromatin organization. Here, we investigated whether EBV alters KDM2B levels to enable its life cycle and promote B-cell transformation. We show that infection of B cells with EBV leads to downregulation of KDM2B levels. We also show that LMP1, one of the main EBV transforming proteins, induces increased DNMT1 recruitment to the KDM2B gene and augments its methylation. By altering KDM2B levels and performing chromatin immunoprecipitation in EBV-infected B cells, we show that KDM2B is recruited to the EBV gene promoters and inhibits their expression. Furthermore, forced KDM2B expression in immortalized B cells led to altered mRNA levels of some differentiation-related genes. Our data show that EBV deregulates KDM2B levels through an epigenetic mechanism and provide evidence for a role of KDM2B in regulating virus and host cell gene expression, warranting further investigations to assess the role of KDM2B in the process of EBV-mediated lymphomagenesis.IMPORTANCE In Africa, Epstein-Barr virus infection is associated with endemic Burkitt lymphoma, a pediatric cancer. The molecular events leading to its development are poorly understood compared with those leading to sporadic Burkitt lymphoma. In a previous study, by analyzing the DNA methylation changes in endemic compared with sporadic Burkitt lymphoma cell lines, we identified several differential methylated genomic positions in the proximity of genes with a potential role in cancer, and among them was the KDM2B gene. KDM2B encodes a histone H3 demethylase already shown to be involved in some hematological disorders. However, whether KDM2B plays a role in the development of Epstein-Barr virus-mediated lymphoma has not been investigated before. In this study, we show that Epstein-Barr virus deregulates KDM2B expression and describe the underlying mechanisms. We also reveal a role of the demethylase in controlling viral and B-cell gene expression, thus highlighting a novel interaction between the virus and the cellular epigenome.

IL-21 Shapes the B Cell Response in a Context-Dependent Manner.

The cytokine interleukin-21 (IL-21) is a pivotal T cell-derived signal crucial for germinal center (GC) responses, but the precise mechanisms by which IL-21 influences B cell function remain elusive. Here, we investigated the B cell-intrinsic role of IL-21 signaling by employing a novel IL-21 receptor ( Il21r ) conditional knock-out mouse model and ex vivo culture systems and uncovered a surprising duality of IL-21 signaling in B cells. While IL-21 stimulation of naïve B cells led to Bim-dependent apoptosis, it promoted robust proliferation of pre-activated B cells, particularly class-switched IgG1 + B cells ex vivo . Consistent with this, B cell-specific deletion of Il21r led to a severe defect in IgG1 responses in vivo following immunization. Intriguingly, Il21r -deleted B cells are significantly impaired in their ability to transition from a pre-GC to a GC state following immunization. Although Il21r -deficiency did not affect the proportion of IgG1 + B cells among GC B cells, it greatly diminished the proportion of IgG1 + B cells among the plasmablast/plasma cell population. Collectively, our data suggest that IL-21 serves as a critical regulator of B cell fates, influencing B cell apoptosis and proliferation in a context-dependent manner.

Epigenetic Alteration of the Cancer-Related Gene TGFBI in B Cells Infected with Epstein-Barr Virus and Exposed to Aflatoxin B1: Potential Role in Burkitt Lymphoma Development.

Burkitt lymphoma (BL) is a malignant B cell neoplasm that accounts for almost half of pediatric cancers in sub-Saharan African countries. Although the BL endemic prevalence is attributable to the combination of Epstein-Barr virus (EBV) infection with malaria and environmental carcinogens exposure, such as the food contaminant aflatoxin B1 (AFB1), the molecular determinants underlying the pathogenesis are not fully understood. Consistent with the role of epigenetic mechanisms at the interface between the genome and environment, AFB1 and EBV impact the methylome of respectively leukocytes and B cells specifically. Here, we conducted a thorough investigation of common epigenomic changes following EBV or AFB1 exposure in B cells. Genome-wide DNA methylation profiling identified an EBV-AFB1 common signature within the TGFBI locus, which encodes for a putative tumor suppressor often altered in cancer. Subsequent mechanistic analyses confirmed a DNA-methylation-dependent transcriptional silencing of TGFBI involving the recruitment of DNMT1 methyltransferase that is associated with an activation of the NF-κB pathway. Our results reveal a potential common mechanism of B cell transformation shared by the main risk factors of endemic BL (EBV and AFB1), suggesting a key determinant of disease that could allow the development of more efficient targeted therapeutic strategies.

HPV Infection Leaves a DNA Methylation Signature in Oropharyngeal Cancer Affecting Both Coding Genes and Transposable Elements.

HPV oncoproteins can modulate DNMT1 expression and activity, and previous studies have reported both gene-specific and global DNA methylation alterations according to HPV status in head and neck cancer. However, validation of these findings and a more detailed analysis of the transposable elements (TEs) are still missing. Here we performed pyrosequencing to evaluate a 5-CpG methylation signature and Line1 methylation in an oropharyngeal squamous cell carcinoma (OPSCC) cohort. We further evaluated the methylation levels of the TEs, their correlation with gene expression and their impact on overall survival (OS) using the TCGA cohort. In our dataset, the 5-CpG signature distinguished HPV-positive and HPV-negative OPSCC with 66.67% sensitivity and 84.33% specificity. Line1 methylation levels were higher in HPV-positive cases. In the TCGA cohort, Line1, Alu and long terminal repeats (LTRs) showed hypermethylation in a frequency of 60.5%, 58.9% and 92.3%, respectively. ZNF541 and CCNL1 higher expression was observed in HPV-positive OPSCC, correlated with lower methylation levels of promoter-associated Alu and LTR, respectively, and independently associated with better OS. Based on our findings, we may conclude that a 5-CpG methylation signature can discriminate OPSCC according to HPV status with high accuracy and TEs are differentially methylated and may regulate gene expression in HPV-positive OPSCC.

Comparison between coronary angioplasty and coronary artery bypass surgery for the treatment of unprotected left main coronary artery stenosis (the Bologna Registry).

Although great interest exists in the relative efficacy of coronary artery bypass grafting (CABG) versus percutaneous coronary intervention (PCI) for the treatment of unprotected left main coronary artery stenosis, data comparing the 2 strategies are scant. Furthermore, no comparison has ever been performed between CABG and drug-eluting stents in this setting. From January 2002 to June 2005, 154 patients with unprotected left main coronary artery stenosis underwent CABG and 157 underwent PCI. Ninety-four patients received a drug-eluting stent in the left main artery. After a median follow-up of 430 days, the rate of mortality, acute myocardial infarction, and target lesion revascularization was 12.3%, 4.5%, and 2.6%, respectively, in the CABG group and 13.4%, 8.3%, and 25.5%, respectively, in the PCI group (death and myocardial infarction p = NS, target lesion revascularization p = 0.0001). Although patients treated with drug-eluting stents had a 25% relative risk reduction in the rate of death, myocardial infarction, and target lesion revascularization compared with patients treated with bare stents, event-free survival was still better for patients treated with CABG. In the multivariate analysis, age >or=70 years, New York Heart Association classes III and IV, acute coronary syndromes, and peripheral vascular disease were the only independent predictors of mortality. In conclusion, our results have indicated that at long-term follow-up no difference exists in the rate of mortality and myocardial infarction between PCI and CABG for the treatment of unprotected left main coronary artery stenosis. However, the rate of target lesion revascularization was higher in the PCI group.