TNRC18 engages H3K9me3 to mediate silencing of endogenous retrotransposons.

Trimethylation of histone H3 lysine 9 (H3K9me3) is crucial for the regulation of gene repression and heterochromatin formation, cell-fate determination and organismal development1. H3K9me3 also provides an essential mechanism for silencing transposable elements1-4. However, previous studies have shown that canonical H3K9me3 readers (for example, HP1 (refs. 5-9) and MPP8 (refs. 10-12)) have limited roles in silencing endogenous retroviruses (ERVs), one of the main transposable element classes in the mammalian genome13. Here we report that trinucleotide-repeat-containing 18 (TNRC18), a poorly understood chromatin regulator, recognizes H3K9me3 to mediate the silencing of ERV class I (ERV1) elements such as LTR12 (ref. 14). Biochemical, biophysical and structural studies identified the carboxy-terminal bromo-adjacent homology (BAH) domain of TNRC18 (TNRC18(BAH)) as an H3K9me3-specific reader. Moreover, the amino-terminal segment of TNRC18 is a platform for the direct recruitment of co-repressors such as HDAC-Sin3-NCoR complexes, thus enforcing optimal repression of the H3K9me3-demarcated ERVs. Point mutagenesis that disrupts the TNRC18(BAH)-mediated H3K9me3 engagement caused neonatal death in mice and, in multiple mammalian cell models, led to derepressed expression of ERVs, which affected the landscape of cis-regulatory elements and, therefore, gene-expression programmes. Collectively, we describe a new H3K9me3-sensing and regulatory pathway that operates to epigenetically silence evolutionarily young ERVs and exert substantial effects on host genome integrity, transcriptomic regulation, immunity and development.

Quantitative classification of chromatin dynamics reveals regulators of intestinal stem cell differentiation.

Intestinal stem cell (ISC) plasticity is thought to be regulated by broadly permissive chromatin shared between ISCs and their progeny. Here, we have used a Sox9EGFP reporter to examine chromatin across ISC differentiation. We find that open chromatin regions (OCRs) can be defined as broadly permissive or dynamic in a locus-specific manner, with dynamic OCRs found primarily in loci consistent with distal enhancers. By integrating gene expression with chromatin accessibility at transcription factor (TF) motifs in the context of Sox9EGFP populations, we classify broadly permissive and dynamic chromatin relative to TF usage. These analyses identify known and potential regulators of ISC differentiation via association with dynamic changes in chromatin. Consistent with computational predictions, Id3-null mice exhibit increased numbers of cells expressing the ISC-specific biomarker OLFM4. Finally, we examine the relationship between gene expression and 5-hydroxymethylcytosine (5hmC) in Sox9EGFP populations, which reveals 5hmC enrichment in absorptive lineage-specific genes. Our data demonstrate that intestinal chromatin dynamics can be quantitatively defined in a locus-specific manner, identify novel potential regulators of ISC differentiation and provide a chromatin roadmap for further dissecting cis regulation of cell fate in the intestine.

An Interactive Database for the Assessment of Histone Antibody Specificity.

Access to high-quality antibodies is a necessity for the study of histones and their posttranslational modifications (PTMs). Here we debut the Histone Antibody Specificity Database (http://www.histoneantibodies.com), an online and expanding resource cataloging the behavior of widely used, commercially available histone antibodies by peptide microarray. This interactive web portal provides a critical resource to the biological research community that routinely uses these antibodies as detection reagents for a wide range of applications.

Sox4 Promotes Atoh1-Independent Intestinal Secretory Differentiation Toward Tuft and Enteroendocrine Fates.

BACKGROUND & AIMS: The intestinal epithelium is maintained by intestinal stem cells (ISCs), which produce postmitotic absorptive and secretory epithelial cells. Initial fate specification toward enteroendocrine, goblet, and Paneth cell lineages requires the transcription factor Atoh1, which regulates differentiation of the secretory cell lineage. However, less is known about the origin of tuft cells, which participate in type II immune responses to parasite infections and appear to differentiate independently of Atoh1. We investigated the role of Sox4 in ISC differentiation. METHODS: We performed experiments in mice with intestinal epithelial-specific disruption of Sox4 (Sox4fl/fl:vilCre; SOX4 conditional knockout [cKO]) and mice without disruption of Sox4 (control mice). Crypt- and single-cell-derived organoids were used in assays to measure proliferation and ISC potency. Lineage allocation and gene expression changes were studied by immunofluorescence, real-time quantitative polymerase chain reaction, and RNA-seq analyses. Intestinal organoids were incubated with the type 2 cytokine interleukin 13 and gene expression was analyzed. Mice were infected with the helminth Nippostrongylus brasiliensis and intestinal tissues were collected 7 days later for analysis. Intestinal tissues collected from mice that express green fluorescent protein regulated by the Atoh1 promoter (Atoh1GFP mice) and single-cell RNA-seq analysis were used to identify cells that coexpress Sox4 and Atoh1. We generated SOX4-inducible intestinal organoids derived from Atoh1fl/fl:vilCreER (ATOH1 inducible knockout) mice and assessed differentiation. RESULTS: Sox4cKO mice had impaired ISC function and secretory differentiation, resulting in decreased numbers of tuft and enteroendocrine cells. In control mice, numbers of SOX4+ cells increased significantly after helminth infection, coincident with tuft cell hyperplasia. Sox4 was activated by interleukin 13 in control organoids; SOX4cKO mice had impaired tuft cell hyperplasia and parasite clearance after infection with helminths. In single-cell RNA-seq analysis, Sox4+/Atoh1- cells were enriched for ISC, progenitor, and tuft cell genes; 12.5% of Sox4-expressing cells coexpressed Atoh1 and were enriched for enteroendocrine genes. In organoids, overexpression of Sox4 was sufficient to induce differentiation of tuft and enteroendocrine cells-even in the absence of Atoh1. CONCLUSIONS: We found Sox4 promoted tuft and enteroendocrine cell lineage allocation independently of Atoh1. These results challenge the longstanding model in which Atoh1 is the sole regulator of secretory differentiation in the intestine and are relevant for understanding epithelial responses to parasitic infection.

Genome-Wide Transcriptional Regulation Mediated by Biochemically Distinct SWI/SNF Complexes.

Multiple positions within the SWI/SNF chromatin remodeling complex can be filled by mutually exclusive subunits. Inclusion or exclusion of these proteins defines many unique forms of SWI/SNF and has profound functional consequences. Often this complex is studied as a single entity within a particular cell type and we understand little about the functional relationship between these biochemically distinct forms of the remodeling complex. Here we examine the functional relationships among three complex-specific ARID (AT-Rich Interacting Domain) subunits using genome-wide chromatin immunoprecipitation, transcriptome analysis, and transcription factor binding maps. We find widespread overlap in transcriptional regulation and the genomic binding of distinct SWI/SNF complexes. ARID1B and ARID2 participate in wide-spread cooperation to repress hundreds of genes. Additionally, we find numerous examples of competition between ARID1A and another ARID, and validate that gene expression changes following loss of one ARID are dependent on the function of an alternative ARID. These distinct regulatory modalities are correlated with differential occupancy by transcription factors. Together, these data suggest that distinct SWI/SNF complexes dictate gene-specific transcription through functional interactions between the different forms of the SWI/SNF complex and associated co-factors. Most genes regulated by SWI/SNF are controlled by multiple biochemically distinct forms of the complex, and the overall expression of a gene is the product of the interaction between these different SWI/SNF complexes. The three mutually exclusive ARID family members are among the most frequently mutated chromatin regulators in cancer, and understanding the functional interactions and their role in transcriptional regulation provides an important foundation to understand their role in cancer.

Human tRNA genes function as chromatin insulators.

Insulators help separate active chromatin domains from silenced ones. In yeast, gene promoters act as insulators to block the spread of Sir and HP1 mediated silencing while in metazoans most insulators are multipartite autonomous entities. tDNAs are repetitive sequences dispersed throughout the human genome and we now show that some of these tDNAs can function as insulators in human cells. Using computational methods, we identified putative human tDNA insulators. Using silencer blocking, transgene protection and repressor blocking assays we show that some of these tDNA-containing fragments can function as barrier insulators in human cells. We find that these elements also have the ability to block enhancers from activating RNA pol II transcribed promoters. Characterization of a putative tDNA insulator in human cells reveals that the site possesses chromatin signatures similar to those observed at other better-characterized eukaryotic insulators. Enhanced 4C analysis demonstrates that the tDNA insulator makes long-range chromatin contacts with other tDNAs and ETC sites but not with intervening or flanking RNA pol II transcribed genes.

Insulators and promoters: closer than we think.

Insulators prevent promiscuous gene regulation by restricting the action of enhancers and silencers. Recent studies have revealed a number of similarities between insulators and promoters, including binding of specific transcription factors, chromatin-modification signatures and localization to specific subnuclear positions. We propose that enhancer-blockers and silencing barrier-insulators might have evolved as specialized derivatives of promoters and that the two types of element use related mechanisms to mediate their distinct functions. These insights can help to reconcile different models of insulator action.

TFIIIC bound DNA elements in nuclear organization and insulation.

tRNA genes (tDNAs) have been known to have barrier insulator function in budding yeast, Saccharomyces cerevisiae, for over a decade. tDNAs also play a role in genome organization by clustering at sites in the nucleus and both of these functions are dependent on the transcription factor TFIIIC. More recently TFIIIC bound sites devoid of pol III, termed Extra-TFIIIC sites (ETC) have been identified in budding yeast and these sites also function as insulators and affect genome organization. Subsequent studies in Schizosaccharomyces pombe showed that TFIIIC bound sites were insulators and also functioned as Chromosome Organization Clamps (COC); tethering the sites to the nuclear periphery. Very recently studies have moved to mammalian systems where pol III genes and their associated factors have been investigated in both mouse and human cells. Short interspersed nuclear elements (SINEs) that bind TFIIIC, function as insulator elements and tDNAs can also function as both enhancer - blocking and barrier insulators in these organisms. It was also recently shown that tDNAs cluster with other tDNAs and with ETCs but not with pol II transcribed genes. Intriguingly, TFIIIC is often found near pol II transcription start sites and it remains unclear what the consequences of TFIIIC based genomic organization are and what influence pol III factors have on pol II transcribed genes and vice versa. In this review we provide a comprehensive overview of the known data on pol III factors in insulation and genome organization and identify the many open questions that require further investigation. This article is part of a Special Issue entitled: Transcription by Odd Pols.

Expanded detection of BAP1 alterations in cancer and tumor type-specific expression score comparison.

BAP1 is a tumor suppressor gene that was originally studied in uveal melanoma (UVM), kidney renal cell clear cell carcinoma (KIRC), and malignant mesothelioma (MESO). Early analyses focused on single-nucleotide variants, but other alteration types such as larger indels and gene-level copy number (CN) loss can also lead to loss of BAP1 expression. We performed integrated multi-omic analyses using data from The Cancer Genome Atlas (TCGA) for 33 cancer types and more than 10,000 individuals. We combined and manually reviewed existing variant calls and new calls derived from a de novo local realignment pipeline across multiple independent variant callers including indel callers, increasing detection of high-quality somatic variant calls by 30% from 91 to 130, including 7 indels ≥40bp. Including CN loss alterations, 1561 samples from 32 cancer types were BAP1-altered, with alterations being predominantly CN-driven. Differential expression and survival analyses revealed both shared and tissue-specific consequences associated with BAP1 alteration. Our findings broadly emphasize the improvements that are gained by using new computational approaches in large cancer-genome studies such as TCGA.

Sox9EGFP Defines Biliary Epithelial Heterogeneity Downstream of Yap Activity.

BACKGROUND & AIMS: Defining the genetic heterogeneity of intrahepatic biliary epithelial cells (BECs) is challenging, and tools for identifying BEC subpopulations are limited. Here, we characterize the expression of a Sox9EGFP transgene in the liver and demonstrate that green fluorescent protein (GFP) expression levels are associated with distinct cell types. METHODS: Sox9EGFP BAC transgenic mice were assayed by immunofluorescence, flow cytometry, and gene expression profiling to characterize in vivo characteristics of GFP populations. Single BECs from distinct GFP populations were isolated by fluorescence-activated cell sorting, and functional analysis was conducted in organoid forming assays. Intrahepatic ductal epithelium was grown as organoids and treated with a Yes-associated protein (Yap) inhibitor or bile acids to determine upstream regulation of Sox9 in BECs. Sox9EGFP mice were subjected to bile duct ligation, and GFP expression was assessed by immunofluorescence. RESULTS: BECs express low or high levels of GFP, whereas periportal hepatocytes express sublow GFP. Sox9EGFP+ BECs are differentially distributed by duct size and demonstrate distinct gene expression signatures, with enrichment of Cyr61 and Hes1 in GFPhigh BECs. Single Sox9EGFP+ cells form organoids that exhibit heterogeneous survival, growth, and HNF4A activation dependent on culture conditions, suggesting that exogenous signaling impacts BEC heterogeneity. Yap is required to maintain Sox9 expression in biliary organoids, but bile acids are insufficient to induce BEC Yap activity or Sox9 in vivo and in vitro. Sox9EGFP remains restricted to BECs and periportal hepatocytes after bile duct ligation. CONCLUSIONS: Our data demonstrate that Sox9EGFP levels provide readout of Yap activity and delineate BEC heterogeneity, providing a tool for assaying subpopulation-specific cellular function in the liver.

Re-expression of SMARCA4/BRG1 in small cell carcinoma of ovary, hypercalcemic type (SCCOHT) promotes an epithelial-like gene signature through an AP-1-dependent mechanism.

Small cell carcinoma of the ovary, hypercalcemic type (SCCOHT) is a rare and aggressive form of ovarian cancer. SCCOHT tumors have inactivating mutations in SMARCA4 (BRG1), one of the two mutually exclusive ATPases of the SWI/SNF chromatin remodeling complex. To address the role that BRG1 loss plays in SCCOHT tumorigenesis, we performed integrative multi-omic analyses in SCCOHT cell lines +/- BRG1 reexpression. BRG1 reexpression induced a gene and protein signature similar to an epithelial cell and gained chromatin accessibility sites correlated with other epithelial originating TCGA tumors. Gained chromatin accessibility and BRG1 recruited sites were strongly enriched for transcription-factor-binding motifs of AP-1 family members. Furthermore, AP-1 motifs were enriched at the promoters of highly upregulated epithelial genes. Using a dominant-negative AP-1 cell line, we found that both AP-1 DNA-binding activity and BRG1 reexpression are necessary for the gene and protein expression of epithelial genes. Our study demonstrates that BRG1 reexpression drives an epithelial-like gene and protein signature in SCCOHT cells that depends upon by AP-1 activity.

Remodeling the cancer epigenome: mutations in the SWI/SNF complex offer new therapeutic opportunities.

INTRODUCTION: Cancer genome sequencing studies have discovered mutations in members of the SWItch/Sucrose Non-Fermentable (SWI/SNF) chromatin-remodeling complex in nearly 25% of human cancers. The SWI/SNF complex, first discovered in S. cerevisiae, shows strong conservation from yeast to Drosophila to mammals, contains approximately 10-12 subunits and regulates nucleosome positioning through the energy generated by its ATPase subunits. The unexpected finding of frequent mutations in the complex has fueled studies to identify the mechanisms that drive tumor development and the accompanying therapeutic vulnerabilities. Areas covered: In the review, we focus upon the potential roles different SWI/SNF subunit mutations play in human oncogenesis, their common and unique mechanisms of transformation and the potential for translating these mechanisms into targeted therapies for SWI/SNF-mutant tumors. Expert opinion: We currently have limited insights into how mutations in different SWI/SNF subunits drive the development of human tumors. Because the SWI/SNF complex participates in a broad range of normal cellular functions, defining specific oncogenic pathways has proved difficult. In addition, therapeutic options for SWI/SNF-mutant cancers have mainly evolved from high-throughput screens of cell lines with mutations in different subunits. Future studies should follow a more coherent plan to pinpoint common vulnerabilities among these tumors.

SWI/SNF remains localized to chromatin in the presence of SCHLAP1.

SCHLAP1 is a long noncoding RNA that is reported to function by depleting the SWI/SNF complex from the genome. We investigated the hypothesis that SCHLAP1 affects only specific compositions of SWI/SNF. Using several assays, we found that SWI/SNF is not depleted from the genome by SCHLAP1 and that SWI/SNF is associated with many coding and noncoding RNAs, suggesting that SCHLAP1 may function in a SWI/SNF-independent manner.

Combined stimulation with interleukin-18 and CpG induces murine natural killer dendritic cells to produce IFN-gamma and inhibit tumor growth.

Natural killer dendritic cells (NKDC) are a novel subtype of dendritic cells with natural killer (NK) cell properties. IFN-gamma is a pleiotropic cytokine that plays an important role in the innate immune response to tumors. Based on our previous finding that the combination of Toll-like receptor 9 ligand CpG and interleukin (IL)-4 stimulates NKDC to produce IFN-gamma, we hypothesized that NKDC are the major IFN-gamma-producing dendritic cell subtype and may play a significant role in the host antitumor response. We found that under several conditions in vitro and in vivo NKDC accounted for the majority of IFN-gamma production by murine spleen CD11c(+) cells. IL-18 alone induced NKDC to secrete IFN-gamma, and the combination of IL-18 and CpG resulted in a synergistic increase in IFN-gamma production, both in vitro and in vivo. NK cells made 26-fold less IFN-gamma under the same conditions in vitro, whereas dendritic cells produced a negligible amount. The mechanism of IFN-gamma secretion by NKDC depended on IL-12. NKDC selectively proliferated in vitro and in vivo in response to the combination of IL-18 and CpG. Systemic treatment with IL-18 and CpG reduced the number of B16F10 melanoma lung metastases. The mechanism depended on NK1.1(+) cells, as their depletion abrogated the effect. IL-18 and CpG activated NKDC provided greater tumor protection than NK cells in IFN-gamma(-/-) mice. Thus, NKDC are the major dendritic cell subtype to produce IFN-gamma. The combined use of IL-18 and CpG is a viable strategy to potentiate the antitumor function of NKDC.

Identification of Two Distinct Classes of the Human INO80 Complex Genome-Wide.

Chromatin remodeling and histone modifying enzymes play a critical role in shaping the regulatory output of a cell. Although much is known about these classes of proteins, identifying the mechanisms by which they coordinate gene expression programs remains an exciting topic of investigation. One factor that may contribute to the targeting and activity of chromatin regulators is local chromatin landscape. We leveraged genomic approaches and publically-available datasets to characterize the chromatin landscape at targets of the human INO80 chromatin remodeling complex (INO80-C). Our data revealed two classes of INO80-C targets with distinct chromatin signatures. The predominant INO80-C class was enriched for open chromatin, H3K27ac, and representative subunits from each of the three INO80-C modules (RUVBL1, RUVBL2, MCRS1, YY1). We named this class Canonical INO80. Notably, we identified an unexpected class of INO80-C targets that contained only the INO80 ATPase and harbored a repressive chromatin signature characterized by inaccessible chromatin, H3K27me3, and the methyltransferase EZH2. We named this class Non-Canonical INO80 (NC-INO80). Biochemical approaches indicated that INO80-C and the H3K27 acetyltransferase P300 physically interact, suggesting INO80-C and P300 may jointly coordinate chromatin accessibility at Canonical INO80 sites. No interaction was detected between INO80-C and EZH2, indicating INO80-C and EZH2 may engage in a separate form of regulatory crosstalk at NC-INO80 targets. Our data indicate that INO80-C is more compositionally heterogenous at its genomic targets than anticipated. Moreover, our data suggest there is an important link between INO80-C and histone modifying enzymes that may have consequences in developmental and pathological contexts.

In vivo overexpression of Flt3 ligand expands and activates murine spleen natural killer dendritic cells.

Natural killer dendritic cells (NKDC) are a unique class of murine immune cells that possess the characteristics of both natural killer (NK) cells and dendritic cells (DC). Because NKDC are able to secrete IFN-gamma, directly lyse tumor cells, and present antigen to naïve T cells, they have immunotherapeutic potential. The relative paucity of NKDC, however, impedes their detailed study. We have found that in vivo, overexpression of the hematopoietic cytokine Flt3 ligand (Flt3L) expands NKDC in various organs from 2-18 fold. Flt3L expanded splenic NKDC retain the ability to lyse tumor cells and become considerably more potent at activating naïve allogeneic and antigen-specific T cells. Compared to normal splenic NKDC, Flt3L-expanded splenic NKDC have a more mature phenotype, a slightly increased ability to capture and process antigen, and a similar cytokine profile. In vivo, we found that Flt3L-expanded splenic NKDC are more effective than normal splenic NKDC in stimulating antigen-specific CD8 T cells. Additionally, we show that NKDC are able to cross-present antigen in vivo. The ability to expand NKDC in vivo using Flt3L will facilitate further analysis of their unique biology. Moreover, Flt3L-expanded NKDC may have enhanced immunotherapeutic potential, given their increased ability to stimulate T cells.

Co-regulation of transcription by BRG1 and BRM, two mutually exclusive SWI/SNF ATPase subunits.

BACKGROUND: SWI/SNF is a large heterogeneous multi-subunit chromatin remodeling complex. It consists of multiple sets of mutually exclusive components. Understanding how loss of one sibling of a mutually exclusive pair affects the occupancy and function of the remaining complex is needed to understand how mutations in a particular subunit might affect tumor formation. Recently, we showed that the members of the ARID family of SWI/SNF subunits (ARID1A, ARID1B and ARID2) had complex transcriptional relationships including both antagonism and cooperativity. However, it remains unknown how loss of the catalytic subunit(s) affects the binding and genome-wide occupancy of the remainder complex and how changes in occupancy affect transcriptional output. RESULTS: We addressed this gap by depleting BRG1 and BRM, the two ATPase subunits in SWI/SNF, and characterizing the changes to chromatin occupancy of the remaining subunit and related this to transcription changes induced by loss of the ATPase subunits. We show that depletion of one subunit frequently leads to loss of the remaining subunit. This could cause either positive or negative changes in gene expression. At a subset of sites, the sibling subunit is either retained or gained. Additionally, we show genome-wide that BRG1 and BRM have both cooperative and antagonistic interactions with respect to transcription. Importantly, at genes where BRG1 and BRM antagonize one another we observe a nearly complete rescue of gene expression changes in the combined BRG/BRM double knockdown. CONCLUSION: This series of experiments demonstrate that mutually exclusive SWI/SNF complexes have heterogeneous functional relationships and highlight the importance of considering the role of the remaining SWI/SNF complexes following loss or depletion of a single subunit.

Epigenetic Regulation by ATP-Dependent Chromatin-Remodeling Enzymes: SNF-ing Out Crosstalk.

Cells utilize precise mechanisms to access genomic DNA with spatiotemporal accuracy. ATP-dependent chromatin-remodeling enzymes (also known simply as "remodelers") comprise a specialized class of enzymes that is intimately involved in genomic organization and accessibility. Remodelers selectively position nucleosomes to either alleviate chromatin compaction or achieve genomic condensation locally, based on a multitude of cellular signals. By dictating nucleosome position, remodelers control local euchromatic and heterochromatic states. These activities govern the accessibility of regulatory regions like promoters and enhancers to transcription factors, RNA polymerases, and coactivators or -repressors. As studies unravel the complexities of epigenetic topography, evidence points to a chromatin-based interactome where regulators interact competitively, cooperatively, and/or codependently through physical and functional means. These types of interactions, or crosstalk, between remodelers raise important questions for tissue development. Here, we briefly review the evidence for remodeler interactions and argue for additional studies examining crosstalk.

Global gene expression profiling of a mouse model of ovarian clear cell carcinoma caused by ARID1A and PIK3CA mutations implicates a role for inflammatory cytokine signaling.

Ovarian clear-cell carcinoma (OCCC) is an aggressive form of epithelial ovarian cancer (EOC). OCCC represents 5-25% of all EOC incidences and is the second leading cause of death from ovarian cancer (Glasspool and McNeish, 2013) [1]. A recent publication by Chandler et al. reported the first mouse model of OCCC that resembles human OCCC both genetically and histologically by inducing a localized deletion of ARID1A and the expression of the PIK3CA(H1047R) substitution mutation (Chandler et al., 2015) [2]. We utilized Affymetrix Mouse Gene 2.1 ST arrays for the global gene expression profiling of mouse primary OCCC tumor samples and animal-matched normal ovaries to identify cancer-dependent gene expression. We describe the approach used to generate the differentially expressed genes from the publicly available data deposited at the Gene Expression Omnibus (GEO) database under the accession number GSE57380. These data were used in cross-species comparisons to publically available human OCCC gene expression data and allowed the identification of coordinately regulated genes in both mouse and human OCCC and supportive of a role for inflammatory cytokine signaling in OCCC pathogenesis (Chandler et al., 2015) [2].

Coexistent ARID1A-PIK3CA mutations promote ovarian clear-cell tumorigenesis through pro-tumorigenic inflammatory cytokine signalling.

Ovarian clear-cell carcinoma (OCCC) is an aggressive form of ovarian cancer with high ARID1A mutation rates. Here we present a mutant mouse model of OCCC. We find that ARID1A inactivation is not sufficient for tumour formation, but requires concurrent activation of the phosphoinositide 3-kinase catalytic subunit, PIK3CA. Remarkably, the mice develop highly penetrant tumours with OCCC-like histopathology, culminating in haemorrhagic ascites and a median survival period of 7.5 weeks. Therapeutic treatment with the pan-PI3K inhibitor, BKM120, prolongs mouse survival by inhibiting the tumour cell growth. Cross-species gene expression comparisons support a role for IL-6 inflammatory cytokine signalling in OCCC pathogenesis. We further show that ARID1A and PIK3CA mutations cooperate to promote tumour growth through sustained IL-6 overproduction. Our findings establish an epistatic relationship between SWI/SNF chromatin remodelling and PI3K pathway mutations in OCCC and demonstrate that these pathways converge on pro-tumorigenic cytokine signalling. We propose that ARID1A protects against inflammation-driven tumorigenesis.