Biology subjects

Robitaille, A.

Publications and source records attributed to Robitaille, A..

5 recordsLinked to original sources

bioRxiv2025

Polycomb group proteins protect latent Kaposi sarcoma-associated herpesvirus from episome clearance and HUSH-dependent chromatin silencing

Kaposi sarcoma-associated herpesvirus (KSHV) is the causative agent of a number of human neoplasms. KSHV causes lifelong chronic infections by establishing persistent reservoirs of latently infected cells in which the viral double-stranded DNA genome is present in the form of an extrachromosomal episome. Polycomb group protein (PcG) mediated gene repression is detected early after entry of the CpG-rich and epigenetically naive viral DNA into the host nucleus. Here we show that the establishment and maintenance of KSHV latency is strongly influenced by Polycomb Repressive Complexes 1 and 2 (PRC1, and PRC2, respectively). Using a series of genetic deletion models, we dissect the functional interdependence of PRC1 and PRC2 on KSHV and identify KSHV episomes as a prime target for independent as well as collaborative de novo PRC recruitment. Loss of PRC function leads to a severely altered chromatin state on KSHV and we show that PRC mediated gene repression is not only preventing transcription of viral genomes but also protects KSHV episomes from detection by the human silencing hub (HUSH). Furthermore, we demonstrate that PRC complexes play an important role in ensuring episomal maintenance in dividing cells. Taken together, our results show that absence of PRC complexes creates a metastable state of KSHV latency and suggest early chromatinization as an integral step for KSHV latency establishment.

microbiology↗

bioRxiv2022

The E2F4/p130 repressor complex cooperates with oncogenic ΔNp73α to promote cell survival in human papillomavirus 38 E6/E7-transformed keratinocytes and in cancer cells

Tumor suppressor p53 and its related proteins, p63 and p73, can be synthesized as multiple isoforms lacking part of the N- or C-terminal regions. Specifically, high expression of the {Delta}Np73 isoform is notoriously associated with various human malignancies characterized by poor prognosis. This isoform is also accumulated by oncogenic viruses such as Epstein-Barr virus (EBV), as well as genus beta human papillomaviruses (HPV) that appear to be involved in carcinogenesis. To gain additional insight into {Delta}Np73 mechanisms, we have performed proteomics analyses using human keratinocytes transformed by the E6 and E7 proteins of the beta-HPV type 38 virus as an experimental model (38HK). We find that {Delta}Np73 associates with the E2F4/p130 repressor complex through a direct interaction with E2F4. This interaction is favored by the N-terminal truncation of p73 characteristic of {Delta}Np73 isoforms. Moreover, it is independent of the C-terminal splicing status, suggesting that it could represent a general feature of {Delta}Np73 isoforms (, {beta}, {gamma}, {delta}, {varepsilon}, {zeta}, {theta}, {eta}, and {eta}1). We also show that the {Delta}Np73- E2F4/p130 complex inhibits the expression of specific genes, including genes encoding for negative regulators of proliferation, both in 38HK and in HPV-negative cancer-derived cell lines. Consistently, silencing of E2F4 in 38HK and in cancer cells results in induction of senescence. In conclusion, we have identified and characterized a novel transcriptional regulatory complex that exerts pro-survival functions in transformed cells. IMPORTANCEThe TP53 gene is mutated in about 50% of human cancers. In contrast, the TP63 and TP73 genes are rarely mutated but rather expressed as {Delta}Np63 and {Delta}Np73 isoforms in a wide range of malignancies, where they act as p53 antagonists. Accumulation of {Delta}Np63 and {Delta}Np73, which is associated with chemoresistance, can result from infection by oncogenic viruses such as EBV or HPV. Our study focuses on the highly carcinogenic {Delta}Np73 isoform and uses a viral model of cellular transformation. We unveil a physical interaction between {Delta}Np73 and the E2F4/p130 complex involved in cell cycle control, which rewires the E2F4/p130 transcriptional program. Consistently, we find that E2F4 gains pro-survival functions in transformed cells expressing {Delta}Np73. This report shows, for the first time, that {Delta}Np73 isoforms acquire novel protein-protein interactions with respect to the TAp73 tumor suppressor. This situation is analogous to the gain-of-function interactions of p53 mutants supporting cellular proliferation.

cancer biology↗

bioRxiv2018

Molecular features of premenopausal breast cancers in Latin American women: pilot results from the PRECAMA study

BackgroundIn Latin America (LA), there is a high incidence rate of breast cancer (BC) in premenopausal women, and the genomic features of these BC remain unknown. Here, we aim to characterize the molecular features of BC in young LA women within the framework of the PRECAMA study, a multicenter population-based case-control study on breast cancer in premenopausal women.\n\nMethodsPathological tumor tissues were collected from incident cases from four LA countries. Immunohistochemistry (IHC) was performed centrally for ER, PR, HER2, Ki67, EGFR, CK5/6 and p53 protein markers. Targeted deep sequencing was done on genomic DNA extracted from formalin-fixed paraffin-embedded (FFPE) tumour tissues and their paired blood samples to screen for somatic mutations in eight genes frequently mutated in BC. A subset of samples was analyzed by exome sequencing to identify somatic mutational signatures.\n\nResultsThe majority of cases were positive for ER or PR (168/233; 72%) and there were 21% triple negative (TN) cases, mainly of basal type. Most tumors were positive for Ki67 (189/233; 81%). In 126 sequenced cases, TP53 and PIK3CA were the most frequently mutated genes (32.5% and 21.4%, respectively), followed by AKT1 (9.5%). TP53 mutations were more frequent in HER2-enriched and TN IHC subtypes, while PIK3CA/AKT1 mutations were more frequent in ER positive tumors, as expected. Interestingly, a higher proportion of G:C>T:A mutations was observed in TP53 gene in PRECAMA cases compared to TCGA and METABRIC breast cancer series (27% vs 14%). Exome-wide mutational patterns in 10 TN cases revealed alterations in signaling transduction pathways and major contributions of mutational signatures caused by altered DNA repair pathways.\n\nConclusionsThis pilot results on PRECAMA tumors gives a preview of the molecular features of premenopausal BC in LA. Although, the overall mutation burden was as expected from data in other populations, mutational patterns observed in TP53 and exome-wide suggested possible differences in mutagenic processes giving rise to these tumors compared to other populations. Further omics analyses of a larger number of cases in the near future will allow investigating relationships between these molecular features and risk factors.

cancer biology↗

bioRxiv2018

Interplay between the epigenetic enzyme lysine (K)-specific demethylase 2B and Epstein-Barr virus infection.

Histone modifier lysine (K)-specific demethylase 2B (KDM2B) plays a role in hematopoietic cells differentiation and its expression appears to be deregulated in certain cancers of hematological and lymphoid origins. We have previously found that KDM2B gene is differentially methylated in cell lines derived from the Epstein-Barr virus (EBV) associated endemic Burkitts lymphomas (eBL) compared to EBV negative sporadic BL cells. However, whether KDM2B plays a role in eBL development has never been previously demonstrated. 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 would alter KDM2B levels to enable its life cycle and promote B-cells transformation. We show that infection of B-cells with EBV leads to down-regulation of KDM2B levels. We also show that LMP1, one of the main EBV transforming proteins, induces increased DNMT1 recruitment to KDM2B gene and augments its methylation. By altering KDM2B levels and performing chromatin immunoprecipitation in EBV infected B-cells, we were able to 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.\n\nIMPORTANCE. 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 to the sporadic Burkitt lymphoma. In a previous study, by analyzing the DNA methylation changes in endemic compared to sporadic Burkitt lymphomas cell lines, we identified several differential methylated genomic positions in proximity of genes with a potential role in cancer, among them 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 never 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-cells genes expression, thus highlighting a novel interaction between the virus and the cellular epigenome.

microbiology↗

bioRxiv2018

Experimental analysis of exome-scale mutational signature of glycidamide, the reactive metabolite of acrylamide

Acrylamide, a probable human carcinogen, is ubiquitously present in the human environment, with sources including heated starchy foods, coffee and cigarette smoke. Humans are also exposed to acrylamide occupationally. Acrylamide is genotoxic, inducing gene mutations and chromosomal aberrations in various experimental settings. Covalent haemoglobin adducts were reported in acrylamide-exposed humans and DNA adducts in experimental systems. The carcinogenicity of acrylamide has been attributed to the effects of glycidamide, its reactive and mutagenic metabolite capable of inducing rodent tumors at various anatomical sites. In order to characterize the pre-mutagenic DNA lesions and global mutation spectra induced by acrylamide and glycidamide, we combined DNA-adduct and whole-exome sequencing analyses in an established exposure-clonal immortalization system based on mouse embryonic fibroblasts. Sequencing and computational analysis revealed a unique mutational signature of glycidamide, characterized by predominant T:A>A:T transversions, followed by T:A>C:G and C:G>A:T mutations exhibiting specific trinucleotide contexts and significant transcription strand bias. Computational interrogation of human cancer genome sequencing data indicated that a combination of the glycidamide signature and an experimental benzo[a]pyrene signature are nearly equivalent to the COSMIC tobacco-smoking related signature 4 in lung adenocarcinomas and squamous cell carcinomas. We found a more variable relationship between the glycidamide- and benzo[a]pyrene-signatures and COSMIC signature 4 in liver cancer, indicating more complex exposures in the liver. Our study demonstrates that the controlled experimental characterization of specific genetic damage associated with glycidamide exposure facilitates identifying corresponding patterns in cancer genome data, thereby underscoring how mutation signature laboratory experimentation contributes to the elucidation of cancer causation.\n\nA 40-word summaryInnovative experimental approaches identify a novel mutational signature of glycidamide, a metabolite of the probable human carcinogen acrylamide. The results may elucidate the cancer risks associated with exposure to acrylamide, commonly found in tobacco smoke, thermally processed foods and beverages.

genomics↗