E4F1 and ZNF148 are transcriptional activators of the -57A > C and wild-type TERT promoter.

Point mutations within the TERT promoter are the most recurrent somatic noncoding mutations identified across different cancer types, including glioblastoma, melanoma, hepatocellular carcinoma, and bladder cancer. They are most abundant at -146C > T and -124C > T, and rarer at -57A > C, with the latter originally described as a familial case, but subsequently shown also to occur somatically. All three mutations create de novo E26-specific (ETS) binding sites and result in activation of the TERT gene, allowing cancer cells to achieve replicative immortality. Here, we used a systematic proteomics screen to identify transcription factors preferentially binding to the -146C > T, -124C > T, and -57A > C mutations. Although we confirmed binding of multiple ETS factors to the mutant -146C > T and -124C > T sequences, we identified E4F1 as a -57A > C-specific binder and ZNF148 as a TERT wild-type (WT) promoter binder that showed reduced interaction with the -124C > T allele. Both proteins are activating transcription factors that bind specifically to the -57A > C and WT (at position 124) TERT promoter sequence in corresponding cell lines, and up-regulate TERT transcription and telomerase activity. Our work describes new regulators of TERT gene expression with possible roles in cancer.

Lysine acetyltransferase Tip60 is required for hematopoietic stem cell maintenance.

Hematopoietic stem cells (HSCs) have the potential to replenish the blood system for the lifetime of the organism. Their 2 defining properties, self-renewal and differentiation, are tightly regulated by the epigenetic machineries. Using conditional gene-knockout models, we demonstrated a critical requirement of lysine acetyltransferase 5 (Kat5, also known as Tip60) for murine HSC maintenance in both the embryonic and adult stages, which depends on its acetyltransferase activity. Genome-wide chromatin and transcriptome profiling in murine hematopoietic stem and progenitor cells revealed that Tip60 colocalizes with c-Myc and that Tip60 deletion suppress the expression of Myc target genes, which are associated with critical biological processes for HSC maintenance, cell cycling, and DNA repair. Notably, acetylated H2A.Z (acH2A.Z) was enriched at the Tip60-bound active chromatin, and Tip60 deletion induced a robust reduction in the acH2A.Z/H2A.Z ratio. These results uncover a critical epigenetic regulatory layer for HSC maintenance, at least in part through Tip60-dependent H2A.Z acetylation to activate Myc target genes.

TFregulomeR reveals transcription factors' context-specific features and functions.

Transcription factors (TFs) are sequence-specific DNA binding proteins, fine-tuning spatiotemporal gene expression. Since genomic occupancy of a TF is highly dynamic, it is crucial to study TF binding sites (TFBSs) in a cell-specific context. To date, thousands of ChIP-seq datasets have portrayed the genomic binding landscapes of numerous TFs in different cell types. Although these datasets can be browsed via several platforms, tools that can operate on that data flow are still lacking. Here, we introduce TFregulomeR (https://github.com/benoukraflab/TFregulomeR), an R-library linked to an up-to-date compendium of cistrome and methylome datasets, implemented with functionalities that facilitate integrative analyses. In particular, TFregulomeR enables the characterization of TF binding partners and cell-specific TFBSs, along with the study of TF's functions in the context of different partnerships and DNA methylation levels. We demonstrated that TFs' target gene ontologies can differ notably depending on their partners and, by re-analyzing well characterized TFs, we brought to light that numerous leucine zipper TFBSs derived from ChIP-seq experiments documented in current databases were inadequately characterized, due to the fact that their position weight matrices were assembled using a mixture of homodimer and heterodimer binding sites. Altogether, analyses of context-specific transcription regulation with TFregulomeR foster our understanding of regulatory network-dependent TF functions.

An epi(c)genetic war: Pathogens, cancer and human genome.

Cancer is characterized by inter- and intra-tumor heterogeneity and this is also observed in the context of cancers caused by pathogens. Nearly 20% of all cancers are attributable to pathogenic organisms. Pathogenic infections result in deregulation of gene expression both by genetic and epigenetic mechanisms, thereby causing malignant transformation. Another characteristic of pathogen-induced cancers is the occurrence of chronic inflammation due to activation of the innate and adaptive arms of the immune system. This review focuses on the epigenetic changes induced by oncoviruses, parasites, cancer-causing bacteria and 'endogenous pathogens' to trigger host cell proliferation indefinitely as well as the inflammation associated with pathogen-induced cancers. The opportunity of targeting components of both pathogen and host epigenetic machinery to limit tumor progression is also discussed.

MethMotif: an integrative cell specific database of transcription factor binding motifs coupled with DNA methylation profiles.

Several recent studies have portrayed DNA methylation as a new player in the recruitment of transcription factors (TF) within chromatin, highlighting a need to connect TF binding sites (TFBS) with their respective DNA methylation profiles. However, current TFBS databases are restricted to DNA binding motif sequences. Here, we present MethMotif, a two-dimensional TFBS database that records TFBS position weight matrices along with cell type specific CpG methylation information computed from a combination of ChIP-seq and whole genome bisulfite sequencing datasets. Integrating TFBS motifs with TFBS DNA methylation better portrays the features of DNA loci recognised by TFs. In particular, we found that DNA methylation patterns within TFBS can be cell specific (e.g. MAFF). Furthermore, for a given TF, different DNA methylation profiles are associated with different DNA binding motifs (e.g. REST). To date, MethMotif database records over 500 TFBSs computed from over 2000 ChIP-seq datasets in 11 different cell types. MethMotif portal is accessible through an open source web interface (https://bioinfo-csi.nus.edu.sg/methmotif) that allows users to intuitively explore the entire dataset and perform both single, and batch queries.

TIP60 represses activation of endogenous retroviral elements.

TIP60 is a lysine acetyltransferase and is known to be a haplo-insufficient tumor suppressor. TIP60 downregulation is an early event in tumorigenesis which has been observed in several cancer types including breast and colorectal cancers. However, the mechanism by which it regulates tumor progression is not well understood. In this study, we identified the role of TIP60 in the silencing of endogenous retroviral elements (ERVs). TIP60-mediated silencing of ERVs is dependent on BRD4. TIP60 and BRD4 positively regulate the expression of enzymes, SUV39H1 and SETDB1 and thereby, the global H3K9 trimethylation (H3K9me3) level. In colorectal cancer, we found that the loss of TIP60 de-represses retrotransposon elements genome-wide, which in turn activate the cellular response to pathogens, mediated by STING, culminating in an induction of Interferon Regulatory Factor 7 (IRF7) and associated inflammatory response. In summary, this study has identified a unique mechanism of ERV regulation in cancer cells mediated by TIP60 and BRD4 through regulation of histone H3 K9 trimethylation, and a new tumor suppressive role of TIP60 in vivo.

TIP60 represses telomerase expression by inhibiting Sp1 binding to the TERT promoter.

HIV1-TAT interactive protein (TIP60) is a haploinsufficient tumor suppressor. However, the potential mechanisms endowing its tumor suppressor ability remain incompletely understood. It plays a vital role in virus-induced cancers where TIP60 down-regulates the expression of human papillomavirus (HPV) oncoprotein E6 which in turn destabilizes TIP60. This intrigued us to identify the role of TIP60, in the context of a viral infection, where it is targeted by oncoproteins. Through an array of molecular biology techniques such as Chromatin immunoprecipitation, expression analysis and mass spectrometry, we establish the hitherto unknown role of TIP60 in repressing the expression of the catalytic subunit of the human telomerase complex, TERT, a key driver for immortalization. TIP60 acetylates Sp1 at K639, thus inhibiting Sp1 binding to the TERT promoter. We identified that TIP60-mediated growth suppression of HPV-induced cervical cancer is mediated in part due to TERT repression through Sp1 acetylation. In summary, our study has identified a novel substrate for TIP60 catalytic activity and a unique repressive mechanism acting at the TERT promoter in virus-induced malignancies.

CRL4(Cdt2) regulates cell proliferation and histone gene expression by targeting PR-Set7/Set8 for degradation.

PR-Set7/Set8 is a cell-cycle-regulated enzyme that monomethylates lysine 20 of histone H4 (H4K20). Set8 and monomethylated H4K20 are virtually undetectable during G1 and S phases of the cell cycle but increase in late S and in G2. We identify CRL4(Cdt2) as the principal E3 ubiquitin ligase responsible for Set8 proteolytic degradation in the S phase of the cell cycle, which requires Set8-PCNA interaction. Inactivation of the CRL4-Cdt2-PCNA-Set8 degradation axis results in (1) DNA damage and the induction of tumor suppressor p53 and p53-transactivated proapoptotic genes, (2) delayed progression through G2 phase of the cell cycle due to activation of the G2/M checkpoint, (3) specific repression of histone gene transcription and depletion of the histone proteins, and (4) repression of E2F1-dependent gene transcription. These results demonstrate a central role of CRL4(Cdt2)-dependent cell-cycle regulation of Set8 for the maintenance of a stable epigenetic state essential for cell viability.

Destabilization of TIP60 by human papillomavirus E6 results in attenuation of TIP60-dependent transcriptional regulation and apoptotic pathway.

The TIP60 tumor suppressor is a histone acetyltransferase involved in transcriptional regulation, checkpoint activation, and p53-directed proapoptotic pathways. We report that human papillomavirus (HPV) E6 destabilizes TIP60 both in vivo and in vitro. TIP60 binds to the HPV major early promoter and acetylates histone H4 to recruit Brd4, a cellular repressor of HPV E6 expression. Both low- and high-risk HPV E6 destabilize TIP60, thereby derepressing their own promoter. Destabilization of TIP60 by HPV E6 also relieves cellular promoters from TIP60-initiated repression and abrogates p53-dependent activation of apoptotic pathway. Degradation of TIP60, therefore, allows low- and high-risk HPV to promote cell proliferation and cell survival.

High-Risk Human Papillomaviral Oncogenes E6 and E7 Target Key Cellular Pathways to Achieve Oncogenesis.

Infection with high-risk human papillomavirus (HPV) has been linked to several human cancers, the most prominent of which is cervical cancer. The integration of the viral genome into the host genome is one of the manners in which the viral oncogenes E6 and E7 achieve persistent expression. The most well-studied cellular targets of the viral oncogenes E6 and E7 are p53 and pRb, respectively. However, recent research has demonstrated the ability of these two viral factors to target many more cellular factors, including proteins which regulate epigenetic marks and splicing changes in the cell. These have the ability to exert a global change, which eventually culminates to uncontrolled proliferation and carcinogenesis.

RVB1/RVB2: running rings around molecular biology.

RVB1/RVB2 (also known as Pontin/Reptin, TIP49/TIP48, RuvbL1/RuvbL2, ECP54/ECP51, INO80H/INO80J, TIH1/TIH2, and TIP49A/TIP49B) are two highly conserved members of the AAA+ family that are present in different protein and nucleoprotein complexes. Recent studies implicate the RVB-containing complexes in many cellular processes such as transcription, DNA damage response, snoRNP assembly, cellular transformation, and cancer metastasis. In this review, we discuss recent advances in our understanding of RVB-containing complexes and their role in these pathways.

Oncoviruses: How do they hijack their host and current treatment regimes.

Viruses have the ability to modulate the cellular machinery of their host to ensure their survival. While humans encounter numerous viruses daily, only a select few can lead to disease progression. Some of these viruses can amplify cancer-related traits, particularly when coupled with factors like immunosuppression and co-carcinogens. The global burden of cancer development resulting from viral infections is approximately 12%, and it arises as an unfortunate consequence of persistent infections that cause chronic inflammation, genomic instability from viral genome integration, and dysregulation of tumor suppressor genes and host oncogenes involved in normal cell growth. This review provides an in-depth discussion of oncoviruses and their strategies for hijacking the host's cellular machinery to induce cancer. It delves into how viral oncogenes drive tumorigenesis by targeting key cell signaling pathways. Additionally, the review discusses current therapeutic approaches that have been approved or are undergoing clinical trials to combat malignancies induced by oncoviruses. Understanding the intricate interactions between viruses and host cells can lead to the development of more effective treatments for virus-induced cancers.

Targeting RNA Exonuclease XRN1 Potentiates Efficacy of Cancer Immunotherapy.

Despite the remarkable clinical responses achieved with immune checkpoint blockade therapy, the response rate is relatively low and only a subset of patients can benefit from the treatment. Aberrant RNA accumulation can mediate IFN signaling and stimulate an immune response, suggesting that targeting RNA decay machinery might sensitize tumor cells to immunotherapy. With this in mind, we identified an RNA exoribonuclease, XRN1, as a potential therapeutic target to suppress RNA decay and stimulate antitumor immunity. Silencing of XRN1 suppressed tumor growth in syngeneic immunocompetent mice and potentiated immunotherapy efficacy, while silencing of XRN1 alone did not affect tumor growth in immunodeficient mice. Mechanistically, XRN1 depletion activated IFN signaling and the viral defense pathway; both pathways play determinant roles in regulating immune evasion. Aberrant RNA-sensing signaling proteins (RIG-I/MAVS) mediated the expression of IFN genes, as depletion of each of them blunted the elevation of antiviral/IFN signaling in XRN1-silenced cells. Analysis of pan-cancer CRISPR-screening data indicated that IFN signaling triggered by XRN1 silencing is a common phenomenon, suggesting that the effect of XRN1 silencing may be extended to multiple types of cancers. Overall, XRN1 depletion triggers aberrant RNA-mediated IFN signaling, highlighting the importance of the aberrant RNA-sensing pathway in regulating immune responses. These findings provide the molecular rationale for developing XRN1 inhibitors and exploring their potential clinical application in combination with cancer immunotherapy. SIGNIFICANCE: Targeting XRN1 activates an intracellular innate immune response mediated by RNA-sensing signaling and potentiates cancer immunotherapy efficacy, suggesting inhibition of RNA decay machinery as a novel strategy for cancer treatment.

Histone-lysine N-methyltransferase EHMT2 (G9a) inhibition mitigates tumorigenicity in Myc-driven liver cancer.

Hepatocellular carcinoma (HCC) is the third deadliest and sixth most common cancer in the world. Histone-lysine N-methyltransferase EHMT2 (also known as G9a) is a histone methyltransferase frequently overexpressed in many cancer types, including HCC. We showed that Myc-driven liver tumours have a unique H3K9 methylation pattern with corresponding G9a overexpression. This phenomenon of increased G9a was further observed in our c-Myc-positive HCC patient-derived xenografts. More importantly, we showed that HCC patients with higher c-Myc and G9a expression levels portend a poorer survival with lower median survival months. We demonstrated that c-Myc interacts with G9a in HCC and cooperates to regulate c-Myc-dependent gene repression. In addition, G9a stabilises c-Myc to promote cancer development, contributing to the growth and invasive capacity in HCC. Furthermore, combination therapy between G9a and synthetic-lethal target of c-Myc, CDK9, demonstrates strong efficacy in patient-derived avatars of Myc-driven HCC. Our work suggests that targeting G9a could prove to be a potential therapeutic avenue for Myc-driven liver cancer. This will increase our understanding of the underlying epigenetic mechanisms of aggressive tumour initiation and lead to improved therapeutic and diagnostic options for Myc-driven hepatic tumours.

Epimutations and Their Effect on Chromatin Organization: Exciting Avenues for Cancer Treatment.

The three-dimensional architecture of genomes is complex. It is organized as fibers, loops, and domains that form high-order structures. By using different chromosome conformation techniques, the complex relationship between transcription and genome organization in the three-dimensional organization of genomes has been deciphered. Epigenetic changes, such as DNA methylation and histone modification, are the hallmark of cancers. Tumor initiation, progression, and metastasis are linked to these epigenetic modifications. Epigenetic inhibitors can reverse these altered modifications. A number of epigenetic inhibitors have been approved by FDA that target DNA methylation and histone modification. This review discusses the techniques involved in studying the three-dimensional organization of genomes, DNA methylation and histone modification, epigenetic deregulation in cancer, and epigenetic therapies targeting the tumor.

Hypertension among Young Adults with Ischemic Stroke of a Tertiary Care Centre in Nepal: A Descriptive Cross-sectional Study.

INTRODUCTION: Ischemic strokes in young patients have been increasing. Younger patients with ischemic stroke tend to have a different long-term prognosis than older patients. Young patients who have residual neurological deficits following ischemic stroke affect their quality of life. This study was carried out to find out the prevalence of hypertension among patients with ischemic stroke in a tertiary care hospital. METHODS: This descriptive cross-sectional study was conducted in all the patients who were admitted with young ischemic stroke in the Department of Neurology, Nobel Medical College Biratnagar from December 2019 to December 2020 after taking ethical clearance from the Institutional Review Committee (reference number: 332/2019). Convenience sampling was done and data was collected, entered in Microsoft Excel, and analysis was done using Statistical Package for the Social Science software version 16. Point estimate at 95% Confidence Interval was calculated along with frequency and proportion for binary data. RESULTS: Out of the total patients with ischemic stroke in the young adults 30 (40%) (95% Confidence Interval= 28.91-51.08) had hypertension. During follow-up at 6 months, 20 (66.7%) of the patients with hypertension had favorable outcomes with a Modified Rankin Score of 0 or 1. Death during a hospital stay or during follow-up was observed in 2 (7%) of patients with hypertension. Mean age of the patients was 40±4.87 years. Most patients were in the age range of 36-45, 24 (80%). CONCLUSIONS: Prevalence of hypertension among young patients with ischemic stroke was high compared to other studies.

The oncogenic E3 ligase TRIP12 suppresses epithelial-mesenchymal transition (EMT) and mesenchymal traits through ZEB1/2.

Thyroid hormone receptor interactor 12 (TRIP12) is an E3 ligase most notably involved in the proteolytic degradation of the tumor suppressor p14ARF. Through this process, it is proposed that TRIP12 plays an oncogenic role in tumor initiation and growth. However, its role in other cancer processes is unknown. In this study, using publicly available cancer patient datasets, we found TRIP12 to be associated with distant metastasis-free survival in breast cancer, suggesting an inhibitory role in metastasis. Following TRIP12 depletion, an epithelial-mesenchymal transition (EMT) shift occurred with concomitant changes in EMT cell adhesion markers identified through RNA-seq. In line with EMT changes, TRIP12-depleted cells gained mesenchymal traits such as loss of cell polarity, dislodgement from bulk cells at a higher frequency, and increased cellular motility. Furthermore, ectopic TRIP12 expression sensitized cells to anoikis. Mechanistically, TRIP12 suppresses EMT through inhibiting ZEB1/2 gene expression, and ZEB1/2 depletion rescues EMT markers and mesenchymal behavior. Overall, our study delineates TRIP12's role in inhibition of EMT and implies a potential suppressive role in breast cancer metastasis.

GAGE mediates radio resistance in cervical cancers via the regulation of chromatin accessibility.

Radiotherapy (RT) resistance is a major cause of treatment failure in cancers that use definitive RT as their primary treatment modality. This study identifies the cancer/testis (CT) antigen G antigen (GAGE) as a mediator of radio resistance in cervical cancers. Elevated GAGE expression positively associates with de novo RT resistance in clinical samples. GAGE, specifically the GAGE12 protein variant, confers RT resistance through synemin-dependent chromatin localization, promoting the association of histone deacetylase 1/2 (HDAC1/2) to its inhibitor actin. This cumulates to elevated histone 3 lysine 56 acetylation (H3K56Ac) levels, increased chromatin accessibility, and improved DNA repair efficiency. Molecular or pharmacological disruption of the GAGE-associated complex restores radiosensitivity. Molecularly, this study demonstrates the role of GAGE in the regulation of chromatin dynamics. Clinically, this study puts forward the utility of GAGE as a pre-screening biomarker to identify poor responders at initial diagnosis and the therapeutic potential of agents that target GAGE and its associated complex in combination with radiotherapy to improve outcomes.

Integrative epigenomic and high-throughput functional enhancer profiling reveals determinants of enhancer heterogeneity in gastric cancer.

BACKGROUND: Enhancers are distal cis-regulatory elements required for cell-specific gene expression and cell fate determination. In cancer, enhancer variation has been proposed as a major cause of inter-patient heterogeneity-however, most predicted enhancer regions remain to be functionally tested. METHODS: We analyzed 132 epigenomic histone modification profiles of 18 primary gastric cancer (GC) samples, 18 normal gastric tissues, and 28 GC cell lines using Nano-ChIP-seq technology. We applied Capture-based Self-Transcribing Active Regulatory Region sequencing (CapSTARR-seq) to assess functional enhancer activity. An Activity-by-contact (ABC) model was employed to explore the effects of histone acetylation and CapSTARR-seq levels on enhancer-promoter interactions. RESULTS: We report a comprehensive catalog of 75,730 recurrent predicted enhancers, the majority of which are GC-associated in vivo (> 50,000) and associated with lower somatic mutation rates inferred by whole-genome sequencing. Applying CapSTARR-seq to the enhancer catalog, we observed significant correlations between CapSTARR-seq functional activity and H3K27ac/H3K4me1 levels. Super-enhancer regions exhibited increased CapSTARR-seq signals compared to regular enhancers, even when decoupled from native chromatin contexture. We show that combining histone modification and CapSTARR-seq functional enhancer data improves the prediction of enhancer-promoter interactions and pinpointing of germline single nucleotide polymorphisms (SNPs), somatic copy number alterations (SCNAs), and trans-acting TFs involved in GC expression. We identified cancer-relevant genes (ING1, ARL4C) whose expression between patients is influenced by enhancer differences in genomic copy number and germline SNPs, and HNF4α as a master trans-acting factor associated with GC enhancer heterogeneity. CONCLUSIONS: Our results indicate that combining histone modification and functional assay data may provide a more accurate metric to assess enhancer activity than either platform individually, providing insights into the relative contribution of genetic (cis) and regulatory (trans) mechanisms to GC enhancer functional heterogeneity.