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Summary: Background: Intestinal T cells are key in gut barrier function. Their role in early stages of alcohol‐associated liver disease (ALD) remain unknown. Aim: To explore the links between intestinal T cells, microbial translocation and ALD Methods: Patients with alcohol use disorder (AUD) following a rehabilitation programme were compared to subjects with non‐alcoholic fatty liver disease (NAFLD) and healthy controls. Clinical and laboratory data (liver stiffness, controlled attenuation parameter, AST, ALT, K18‐M65) served to identify AUD patients with isolated steatosis (minimal liver disease) or steatohepatitis/fibrosis (ALD). Serum microbial translocation markers were measured by ELISA, duodenal and plasma levels of sphingolipids by targeted LC–MS. T lymphocytes in duodenal biopsies were characterised by immunohistochemistry, flow cytometry and RNA sequencing on FACS‐sorted cells. Mechanisms for T‐cell alterations were assessed in vitro. Results: Patients with ALD, but not those with minimal liver disease, showed reduced numbers of duodenal CD8+ T resident memory (TRM) cells compared to controls or patients with NAFLD. TRM transcriptomic analysis, in vitro analyses and pharmacological inhibition of cathepsin B confirmed TRM apoptosis driven by lysosomal membrane permeabilisation and cathepsin B release into the cytosol. Altered lipid metabolism and increased duodenal and plasma sphingolipids correlated with apoptosis. Dihydroceramide dose‐dependently reduced viability of TRM. Duodenal TRM phenotypic changes, apoptosis and transcriptomic alterations correlated with increased levels of microbial translocation markers. Short‐term abstinence did not reverse TRM cell death in patients with ALD. Conclusions: Duodenal CD8+ TRM apoptosis related to functional changes in lysosomes and lipid metabolism points to impaired gut adaptive immunity specifically in patients with AUD who developed early ALD. [ABSTRACT FROM AUTHOR]

Tumor development involves alterations in DNA methylation patterns, which include both gains (hypermethylation) and losses (hypomethylation) in different genomic regions. The mechanisms underlying these two opposite, yet co-existing, alterations in tumors remain unclear. While studying the human MAGEA6/GABRA3 gene locus, we observed that DNA hypomethylation in tumor cells can lead to the activation of a long transcript (CT-GABRA3) that overlaps downstream promoters (GABRQ and GABRA3) and triggers their hypermethylation. Overlapped promoters displayed increases in H3K36me3, a histone mark deposited during transcriptional elongation and known to stimulate de novo DNA methylation. Consistent with such a processive mechanism, increases in H3K36me3 and DNA methylation were observed over the entire region covered by the CT-GABRA3 overlapping transcript. Importantly, experimental induction of CT-GABRA3 by depletion of DNMT1 DNA methyltransferase, resulted in a similar pattern of regional DNA hypermethylation. Bioinformatics analyses in lung cancer datasets identified other genomic loci displaying this process of coupled DNA hypo/hypermethylation, and some of these included tumor suppressor genes, e.g. RERG and PTPRO. Together, our work reveals that focal DNA hypomethylation in tumors can indirectly contribute to hypermethylation of nearby promoters through activation of overlapping transcription, and establishes therefore an unsuspected connection between these two opposite epigenetic alterations. [ABSTRACT FROM AUTHOR]

Background: Most studies on tumour progression from precursor lesion toward gallbladder adenocarcinoma investigate lesions sampled from distinct patients, providing an overarching view of pathogenic cascades. Whether this reflects the tumourigenic process in individual patients remains insufficiently explored. Genomic and epigenomic studies suggest that a subset of gallbladder cancers originate from biliary intraepithelial neoplasia (BilIN) precursor lesions, whereas others form independently from BilINs. Spatial transcriptomic data supporting these conclusions are missing. Moreover, multiple areas with precursor or adenocarcinoma lesions can be detected within the same pathological sample. Yet, knowledge about intra-patient variability of such lesions is lacking. Methods: To characterise the spatial transcriptomics of gallbladder cancer tumourigenesis in individual patients, we selected two patients with distinct cancer aetiology and whose samples simultaneously displayed multiple areas of normal epithelium, BilINs and adenocarcinoma. Using GeoMx digital spatial profiling, we characterised the whole transcriptome of a high number of regions of interest (ROIs) per sample in the two patients (24 and 32 ROIs respectively), with each ROI covering approximately 200 cells of normal epithelium, low-grade BilIN, high-grade BilIN or adenocarcinoma. Human gallbladder organoids and cell line-derived tumours were used to investigate the tumour-promoting role of genes. Results: Spatial transcriptomics revealed that each type of lesion displayed limited intra-patient transcriptomic variability. Our data further suggest that adenocarcinoma derived from high-grade BilIN in one patient and from low-grade BilIN in the other patient, with co-existing high-grade BilIN evolving via a distinct process in the latter case. The two patients displayed distinct sequences of signalling pathway activation during tumour progression, but Semaphorin 4 A (SEMA4A) expression was repressed in both patients. Using human gallbladder-derived organoids and cell line-derived tumours, we provide evidence that repression of SEMA4A promotes pseudostratification of the epithelium and enhances cell migration and survival. Conclusion: Gallbladder adenocarcinoma can develop according to patient-specific processes, and limited intra-patient variability of precursor and cancer lesions was noticed. Our data suggest that repression of SEMA4A can promote tumour progression. They also highlight the need to gain gene expression data in addition to histological information to avoid understimating the risk of low-grade preneoplastic lesions. [ABSTRACT FROM AUTHOR]

Global loss of DNA methylation is frequently observed in the genome of human tumors. Although this epigenetic alteration is clearly associated with cancer progression, the way it exerts its pro-tumoral effect remains incompletely understood. A remarkable consequence of DNA hypomethylation in tumors is the aberrant activation of "cancer-germline" genes (also known as "cancer-testis" genes), which comprise a diverse group of germline-specific genes that use DNA methylation as a primary mechanism for repression in normal somatic tissues. Here we review the evidence that such cancer-germline genes contribute to key processes of tumor development. Notably, several cancer-germline genes were found to stimulate oncogenic pathways involved in cell proliferation (SSX, DDX43, MAEL, PIWIL1), angiogenesis (DDX53), immortality (BORIS/CTCFL), and metastasis (CT-GABRA3). Others appear to inhibit tumor suppressor pathways, including those controlling growth inhibition signals (MAGEA11, MAGEB2), apoptosis (MAGEA2, MAGEC2), and genome integrity (HORMAD1, NXF2). Cancer-germline genes were also implicated in the regulation of tumor metabolism (MAGEA3/MAGEA6). Together, our survey substantiates the concept that DNA hypomethylation promotes tumorigenesis via transcriptional activation of oncogenes. Importantly, considering their highly restricted pattern of expression, cancer-germline genes may represent valuable targets for the development of anti-cancer therapies with limited side effects. [ABSTRACT FROM AUTHOR]

Gene MAGEA1 belongs to a group of human germline-specific genes that rely on DNA methylation for repression in somatic tissues. Many of these genes, termed cancer-germline (CG) genes, become demethylated and activated in a wide variety of tumors, where they encode tumor-specific antigens. The process leading to DNA demethylation of CG genes in tumors remains unclear. Previous data suggested that histone acetylation might be involved. Here, we investigated the relative contribution of DNA methylation and histone acetylation in the epigenetic regulation of gene MAGEA1. We show that MAGEA1 DNA hypomethylation in expressing melanoma cells is indeed correlated with local increases in histone H3 acetylation (H3ac). However, when MAGEA1-negative cells were exposed to a histone deacetylase inhibitor (TSA), we observed only short-term activation of the gene and detected no demethylation of its promoter. As a more sensitive assay, we used a cell clone harboring a methylated MAGEA1/hph construct, which confers resistance to hygromycin upon stable re-activation. TSA induced only transient de-repression of the transgene, and did not lead to the emergence of hygromycin-resistant cells. In striking contrast, transient depletion of DNA-methyltransferase-1 in the reporter cell clone gave rise to a hygromycin-resistant population, in which the re-activated MAGEA1/hph transgene displayed not only marked DNA hypomethylation, but also significant reversal of histone marks, including gains in H3ac and H3K4me2, and losses of H3K9me2. Collectively, our results indicate that DNA methylation has a dominant role in the epigenetic hierarchy governing MAGEA1 expression. [ABSTRACT FROM AUTHOR]

Abstract: Several human germline-specific genes rely principally on DNA methylation for repression in somatic tissues. Many of these genes, including MAGEA1, were qualified as cancer-germline (CG), as they become activated in tumors, where losses of DNA methylation are common. The developmental stage at which CG genes acquire DNA methylation marks is unknown. Here, we show that in human preimplantation embryos, transcription of CG genes increases up to the morula stage, and then decreases dramatically in blastocysts, suggesting that CG gene silencing occurs in blastocyst stem cells. Consistently, transfection studies with MAGEA1 constructs in embryonal carcinoma (EC) cells, which represent a malignant surrogate of blastocyst-derived stem cells, revealed active repression and marked de novo methylation of MAGEA1 transgenes in these cells. Active repression of the endogenous MAGEA1 gene in human EC cells was evidenced by its rapid re-silencing following prior induction with a DNA methylation inhibitor. Moreover, de novo DNA methyltransferases DNMT3A and DNMT3B appeared to contribute to the silencing of MAGEA1 and other CG genes in EC cells. Altogether our data indicate that CG genes like MAGEA1 are programmed for repression in the blastocyst, and suggest that de novo DNA methylation is a key event in this process. [Copyright &y& Elsevier]

MAGE-A1 belongs to a group of germ line-specific genes that rely primarily on DNA methylation for repression in somatic tissues. In many types of tumors, the promoter of these genes becomes demethylated and transcription becomes activated. We showed previously that, although MZ2-MEL melanoma cells contain an active unmethylated MAGE-A1 gene, they lack the ability to induce demethylation of newly integrated MAGE-A1 transgenes that were methylated in vitro before transfection. In the same cells, unmethylated MAGE-A1 transgenes were protected against remethylation, and this appeared to depend on the level of transcriptional activity. We therefore proposed that hypomethylation of MAGE-A1 in tumors relies on a past demethylation event and on the presence of appropriate transcription factors that maintain the promoter unmethylated. Here, we tested this hypothesis further by examining whether induction of a transient demethylation phase in MZ2-MEL would suffice to convert a previously methylated MAGE-A1 transgene into a permanently hypomethylated and active one. For induction of the demethylation phase, we used antisense oligonucleotides targeting the three known human DNA methyltransferases. We found that clown-regulation of DNMT1, but not of DNMT3A and DNMT3B, induces activation of the MAGE-A1 transgene, suggesting that DNMT1 has a predominant role for methylation maintenance in MZ2-MEL cells. By using a selectable MA GE-A1 transgene construct, we were able to isolate a cell population in which DNMT1 depletion had resulted in transgene activation. The promoter region of the transgene was almost completely unmethylated in these cells, and this active and unmethylated state was maintained for over 60 days after restoration of normal DNMT1 expression. [ABSTRACT FROM AUTHOR]

Several male germ line-specific genes, including MAGE-A1, rely on DNA methylation for their repression in normal somatic tissues. These genes become activated in many types of tumors in the course of the genomewide demethylation process which often accompanies tumorigenesis. We show that in tumor cells expressing MAGE-A1, the 5' region is significantly less methylated than the other parts of the gene. The process leading to this site-specific hypomethylation does not appear to be permanent in these tumor cells, since in vitromethylated MAGE-A1 sequences do not undergo demethylation after being stably transfected. However, in these cells there is a process that inhibits de novo methylation within the 5' region of MAGE-A1, since unmethylated MAGE-A1 transgenes undergo remethylation at all CpGs except those located within the 5' region. This local inhibition of methylation appears to depend on promoter activity. We conclude that the site-specific hypomethylation of MAGE-A1 in tumor cells relies on a transient process of demethylation followed by a persistent local inhibition of remethylation due to the presence of transcription factors. [ABSTRACT FROM AUTHOR]

Simple Summary: Loss of DNA methylation is often observed in human tumors, but how this epigenetic alteration impacts the transcriptome of cancer cells remains largely undefined. So far, DNA hypomethylation in tumors has been associated with aberrant activation of a germline-specific gene expression program. Here, we exploited transcriptomic and methylomic datasets of lung adenocarcinoma to investigate the possibility that other gene expression programs also become ectopically activated in hypomethylated tumors. Remarkably, we found that DNA hypomethylation in lung adenocarcinoma is associated with ectopic activation of not only germline-specific genes, but also gene clusters displaying specific expression in the gastrointestinal tract, or in stratified epithelia. Interestingly, expression of genes in this latter group was of prognostic value. Together, our study brings novel insight into the transcriptomic changes associated with DNA hypomethylation in tumors, and is an incentive to explore the value of hypomethylated DNA sequences as cancer biomarkers. Genome-wide loss of DNA methylation is commonly observed in human cancers, but its impact on the tumor transcriptome remains ill-defined. Previous studies demonstrated that this epigenetic alteration causes aberrant activation of a germline-specific gene expression program. Here, we examined if DNA hypomethylation in tumors also leads to de-repression of gene clusters with other tissue specificities. To this end, we explored transcriptomic and methylomic datasets from human lung adenocarcinoma (LUAD) cell lines, normal lung, and lung alveolar type II cells, considered as the origin of LUAD. Interestingly, DNA demethylation in LUAD cell lines was associated with activation of not only germline-specific (CG) genes, but also gene clusters displaying specific expression in the gastrointestinal tract (GI), or in stratified epithelia (SE). Consistently, genes from all three clusters showed highly specific patterns of promoter methylation among normal tissues and cell types, and were generally sensitive to induction by a DNA demethylating agent. Analysis of TCGA datasets confirmed that demethylation and activation of CG, GI and SE genes also occurs in vivo in LUAD tumor tissues, in association with global genome hypomethylation. For genes of the GI cluster, we demonstrated that HNF4A is a necessary factor for transcriptional activation following promoter demethylation. Interestingly, expression of several SE genes, in particular FAM83A, correlated with both tumor grade and reduced patient survival. Together, our study uncovers novel cell-type specific gene clusters that become aberrantly activated in LUAD tumors in association with genome-wide hypomethylation. [ABSTRACT FROM AUTHOR]

Objective: The objective is to characterize transcriptomic profiles and immune cell composition and distribution in juvenile idiopathic arthritis (JIA) synovial biopsies, assess for associations of these features with clinical parameters, and compare JIA and rheumatoid arthritis (RA) synovial features. Methods: RNA sequencing (RNASeq) was performed on 24 samples, with pathway analysis and inference of relative abundance of immune cell subsets based on gene expression data. Two multiplex fluorescence immunohistochemistry (IHC) panels were performed on 28 samples (including 13 on which RNASeq was performed), staining for CD206− classical and CD206+ nonclassical macrophages, and CD8+ and CD4+ T and B lymphocytes. Data were compared to a published series of early RA synovial biopsies. Results: Pathway analysis of the most variably expressed genes (n = 339) identified a B and plasma cell signature as the main driver of heterogeneity in JIA synovia, with strong overlap between JIA and RA synovitis. Multiplex IHC confirmed heterogeneity of immune cell infiltration. M1‐like macrophage–rich synovial lining was associated with greater lining hypertrophy and higher (CD45+) pan–immune cell and CD8+ T cell infiltration. Conclusion: Our study indicates significant similarities between JIA and RA synovitis. Similar to RA, JIA synovia may be broadly categorized into two groups: (1) those with an inflammatory/adaptive immune transcriptomic signature, M1‐like macrophage and CD8+ T cell infiltration, and thicker, M1‐like macrophage–rich synovial lining, and (2) those with an M2‐like macrophage transcriptomic signature, greater M2/M1‐like macrophage ratios, and thinner, M2‐like macrophage–rich synovial lining. Synovial features were not significantly associated with clinical parameters, likely because of group size and heterogeneity. [ABSTRACT FROM AUTHOR]

Biliary ducts collect bile from liver lobules, the smallest functional and anatomical units of liver, and carry it to the gallbladder. Disruptions in this process caused by defective embryonic development, or through ductal reaction in liver disease have a major impact on life quality and survival of patients. A deep understanding of the processes underlying bile duct lumen formation is crucial to identify intervention points to avoid or treat the appearance of defective bile ducts. Several hypotheses have been proposed to characterize the biophysical mechanisms driving initial bile duct lumen formation during embryogenesis. Here, guided by the quantification of morphological features and expression of genes in bile ducts from embryonic mouse liver, we sharpened these hypotheses and collected data to develop a high resolution individual cell-based computational model that enables to test alternative hypotheses in silico. This model permits realistic simulations of tissue and cell mechanics at sub-cellular scale. Our simulations suggest that successful bile duct lumen formation requires a simultaneous contribution of directed cell division of cholangiocytes, local osmotic effects generated by salt excretion in the lumen, and temporally-controlled differentiation of hepatoblasts to cholangiocytes, with apical constriction of cholangiocytes only moderately affecting luminal size. Author summary: The initial step in bile duct development is the formation of a biliary lumen, a process which involves several cellular mechanisms, such as cell division and polarization, and secretion of fluid. However, how these mechanisms are orchestrated in time and space is difficult to understand. Here, we built a computational model of biliary lumen formation which represents every cell and its function in detail. With the model we can simulate the effect of biophysical aspects that affect duct formation. We have tested the individual and combined effects of directed cell division, apical constriction, and osmotic effects on lumen expansion by varying the parameters that control their relative strength. Our simulations suggest that successful bile duct lumen formation requires the simultaneous contribution of directed cell division of cholangiocytes, local osmotic effects generated by salt excretion in the lumen, and temporally-controlled differentiation of hepatoblasts to cholangiocytes, with apical constriction of cholangiocytes only moderately affecting luminal size. [ABSTRACT FROM AUTHOR]

The human genome harbors many duplicated segments, which sometimes show very high sequence identity. This may complicate assignment during genome assembly. One such example is in Xq28, where the arrangement of 2 recently duplicated segments varies between genome assembly versions. The duplicated segments comprise highly similar genes, including MAGEA3 and MAGEA6, which display specific expression in testicular germline cells, and also become aberrantly activated in a variety of tumors. Recently, a new gene was identified, CT-GABRA3, the transcription of which initiates inside the segmental duplication but extends far outside. According to the latest genome annotation, CT- GABRA3 starts near MAGEA3, with which it shares a bidirectional promoter. In an earlier annotation, however, the duplicated segment was positioned in the opposite orientation, and CT-GABRA3 was instead coupled with MAGEA6. To resolve this discrepancy, and based on the contention that genes connected by a bidirectional promoter are almost always co-expressed, we decided to compare the expression profiles of CT-GABRA3, MAGEA3, and MAGEA6. We found that in tumor tissues and cell lines of different origins, the expression of CT-GABRA3 was better correlated with that of MAGEA6. Moreover, in a cellular model of experimental induction with a DNA demethylation agent, activation CT-GABRA3 was associated with that of MAGEA6, but not with that of MAGEA3. Together these results support a connection between CT-GABRA3 and MAGEA6 and illustrate how promoter-sharing genes can be exploited to resolve genome assembly uncertainties. [ABSTRACT FROM AUTHOR]

Earlier data on liver development demonstrated that morphogenesis of the bile duct, portal mesenchyme and hepatic artery is interdependent, yet how this interdependency is orchestrated remains unknown. Here, using 2D and 3D imaging, we first describe how portal mesenchymal cells become organised to form hepatic arteries. Next, we examined intercellular signalling active during portal area development and found that axon guidance genes are dynamically expressed in developing bile ducts and portal mesenchyme. Using tissue-specific gene inactivation in mice, we show that the repulsive guidance molecule BMP co-receptor A (RGMA)/neogenin (NEO1) receptor/ligand pair is dispensable for portal area development, but that deficient roundabout 2 (ROBO2)/ SLIT2 signalling in the portal mesenchyme causes reduced maturation of the vascular smooth muscle cells that form the tunica media of the hepatic artery. This arterial anomaly does not impact liver function in homeostatic conditions, but is associated with significant tissular damage following partial hepatectomy. In conclusion, our work identifies new players in development of the liver vasculature in health and liver regeneration. [ABSTRACT FROM AUTHOR]

Background: Many human tumors show aberrant activation of a group of germline-specific genes, termed cancergermline (CG) genes, several of which appear to exert oncogenic functions. Although activation of CG genes in tumors has been linked to promoter DNA demethylation, the mechanisms underlying this epigenetic alteration remain unclear. Two main processes have been proposed: awaking of a gametogenic program directing demethylation of target DNA sequences via specific regulators, or general deficiency of DNA methylation activities resulting from mis-targeting or down-regulation of the DNMT1 methyltransferase. Results: By the analysis of transcriptomic data, we searched to identify gene expression changes associated with CG gene activation in melanoma cells. We found no evidence linking CG gene activation with differential expression of gametogenic regulators. Instead, CG gene activation correlated with decreased expression of a set of mitosis/divisionrelated genes (ICCG genes). Interestingly, a similar gene expression signature was previously associated with depletion of DNMT1. Consistently, analysis of a large set of melanoma tissues revealed that DNMT1 expression levels were often lower in samples showing activation of multiple CG genes. Moreover, by using immortalized melanocytes and fibroblasts carrying an inducible anti-DNMT1 small hairpin RNA (shRNA), we demonstrate that transient depletion of DNMT1 can lead to long-term activation of CG genes and repression of ICCG genes at the same time. For one of the ICCG genes (CDCA7L), we found that its down-regulation in melanoma cells was associated with deposition of repressive chromatin marks, including H3K27me3. Conclusions: Together, our observations point towards transient DNMT1 depletion as a causal factor of CG gene activation in vivo in melanoma. [ABSTRACT FROM AUTHOR]

Simple Summary: Incidence of thyroid cancer, including papillary thyroid cancer, is rapidly increasing. Oncogenes, such as the BRAFV600E, have been identified, and their effect on thyroid cancer cells have been studied in vitro and in mouse models. What is less understood is the impact of these mutations on thyroid cancer microenvironment and, in turn, the effect of changes in the microenvironment on tumor progression. We investigated the modifications in the cellular composition of thyroid cancer microenvironment using an inducible mouse model. We focused on a subpopulation of macrophages, expressing the STABILIN-1 protein, recruited in the thyroid tumor microenvironment following BRAFV600E expression. CRISPR/Cas9 genetic inactivation of Stablin-1 did not change macrophage recruitment but highlighted the immunosuppressive role of STABILIN-1-expressing macrophages. The identification of a similar subpopulation of STABILIN-1 macrophages in human thyroid diseases supports a conserved role for these macrophages and offers an opportunity for intervention. Papillary thyroid carcinoma (PTC) is the most frequent histological subtype of thyroid cancers (TC), and BRAFV600E genetic alteration is found in 60% of this endocrine cancer. This oncogene is associated with poor prognosis, resistance to radioiodine therapy, and tumor progression. Histological follow-up by anatomo-pathologists revealed that two-thirds of surgically-removed thyroids do not present malignant lesions. Thus, continued fundamental research into the molecular mechanisms of TC downstream of BRAFV600E remains central to better understanding the clinical behavior of these tumors. To study PTC, we used a mouse model in which expression of BRAFV600E was specifically switched on in thyrocytes by doxycycline administration. Upon daily intraperitoneal doxycycline injection, thyroid tissue rapidly acquired histological features mimicking human PTC. Transcriptomic analysis revealed major changes in immune signaling pathways upon BRAFV600E induction. Multiplex immunofluorescence confirmed the abundant recruitment of macrophages, among which a population of LYVE-1+/CD206+/STABILIN-1+ was dramatically increased. By genetically inactivating the gene coding for the scavenger receptor STABILIN-1, we showed an increase of CD8+ T cells in this in situ BRAFV600E-dependent TC. Lastly, we demonstrated the presence of CD206+/STABILIN-1+ macrophages in human thyroid pathologies. Altogether, we revealed the recruitment of immunosuppressive STABILIN-1 macrophages in a PTC mouse model and the interest to further study this macrophage subpopulation in human thyroid tissues. [ABSTRACT FROM AUTHOR]

• We identified a gene regulatory network (GRN) involved in a subset of hepatocellular carcinomas. • Expression of GRN members and targets correlates with proliferation and prognosis. • A quantitative mathematical model of the GRN revealed the network dynamics. • This model is a potential tool to assess the impact of pharmacological inhibitors. Alterations of individual genes variably affect the development of hepatocellular carcinoma (HCC). Thus, we aimed to characterize the function of tumor-promoting genes in the context of gene regulatory networks (GRNs). Using data from The Cancer Genome Atlas, from the LIRI-JP (Liver Cancer – RIKEN, JP project), and from our transcriptomic, transfection and mouse transgenic experiments, we identify a GRN which functionally links LIN28B-dependent dedifferentiation with dysfunction of β-catenin (CTNNB1). We further generated and validated a quantitative mathematical model of the GRN using human cell lines and in vivo expression data. We found that LIN28B and CTNNB1 form a GRN with SMARCA4, Let-7b (MIRLET7B), SOX9, TP53 and MYC. GRN functionality is detected in HCC and gastrointestinal cancers, but not in other cancer types. GRN status negatively correlates with HCC prognosis, and positively correlates with hyperproliferation, dedifferentiation and HGF/MET pathway activation, suggesting that it contributes to a transcriptomic profile typical of the proliferative class of HCC. The mathematical model predicts how the expression of GRN components changes when the expression of another GRN member varies or is inhibited by a pharmacological drug. The dynamics of GRN component expression reveal distinct cell states that can switch reversibly in normal conditions, and irreversibly in HCC. The mathematical model is available via a web-based tool which can evaluate the GRN status of HCC samples and predict the impact of therapeutic agents on the GRN. We conclude that identification and modelling of the GRN provide insights into the prognosis of HCC and the mechanisms by which tumor-promoting genes impact on HCC development. Hepatocellular carcinoma (HCC) is a heterogeneous disease driven by the concomitant deregulation of several genes functionally organized as networks. Here, we identified a gene regulatory network involved in a subset of HCCs. This subset is characterized by increased proliferation and poor prognosis. We developed a mathematical model which uncovers the dynamics of the network and allows us to predict the impact of a therapeutic agent, not only on its specific target but on all the genes belonging to the network. [ABSTRACT FROM AUTHOR]

The Myc transcription factor is an essential mediator of cell growth and proliferation through its ability to both positively and negatively regulate transcription. The mechanisms by which Myc silences gene expression are not well understood. The current model is that Myc represses transcription through functional interference with transcriptional activators. Here we show that Myc binds the corepressor Dnmt3a and associates with DNA methyltransferase activity in vivo. In cells with reduced Dnmt3a levels, we observe specific reactivation of the Myc-repressed p21Cip1 gene, whereas the expression of Myc-activated E-boxes genes is unchanged. In addition, we find that Myc can target Dnmt3a selectively to the promoter of p21Cip1. Myc is known to be recruited to the p21Cip1 promoter by the DNA-binding factor Miz-1. Consistent with this, we observe that Myc and Dnmt3a form a ternary complex with Miz-1 and that this complex can corepress the p21Cip1 promoter. Finally, we show that DNA methylation is required for Myc-mediated repression of p21Cip1. Our data identify a new mechanism by which Myc can silence gene expression not only by passive functional interference but also by active recruitment of corepressor proteins. Furthermore, these findings suggest that targeting of DNA methyltransferases by transcription factors is a wide and general mechanism for the generation of specific DNA methylation patterns within a cell. [ABSTRACT FROM AUTHOR]

An inappropriate response to progestogens in the human endometrium can result in fertility issues and jeopardize progestin-based treatments against pathologies such as endometriosis. PGRMC1 can mediate progesterone response in the breast and ovaries but its endometrial functions remain unknown. AG-205 is an alleged PGRMC1 inhibitor but its specificity was recently questioned. We added AG-205 in the cultures of two endometrial cell lines and performed a transcriptomic comparison. AG-205 significantly increased expression of genes coding enzymes of the cholesterol biosynthetic pathway or of steroidogenesis. However, these observations were not reproduced with cells transfected with siRNA against PGRMC1 or its related proteins (MAPRs). Furthermore, AG-205 retained its ability to increase expression of selected target genes even when expression of PGRMC1 or all MAPRs was concomitantly downregulated, indicating that neither PGRMC1 nor any MAPR is required to mediate AG-205 effect. In conclusion, although AG-205 has attractive effects encouraging its use to develop therapeutic strategies, for instance against breast cancer, our study delivers two important warning messages. First, AG-205 is not specific for PGRMC1 or other MAPRs and its mechanisms of action remain unclear. Second, due to its effects on genes involved in steroidogenesis, its use may increase the risk for endometrial pathologies resulting from imbalanced hormones concentrations. [ABSTRACT FROM AUTHOR]