Abstract
WashU Epigenome Browser (https://epigenomegateway.wustl.edu/browser/) is a web-based genomic data exploration tool that provides visualization, integration, and analysis of epigenomic ...datasets. The newly renovated user interface and functions have enabled researchers to engage with the browser and genomic data more efficiently and effectively since 2018. Here, we introduce a new integrated panel design in the browser that allows users to interact with 1D (genomic features), 2D (such as Hi-C), 3D (genome structure), and 4D (time series) data in a single web page. The browser can display three-dimensional chromatin structures with the 3D viewer module. The 4D tracks, called ‘Dynamic’ tracks, animatedly display time-series data, allowing for a more striking visual impact to identify the gene or genomic region candidates as a function of time. Genomic data, such as annotation features, numerical values, and chromatin interaction data can all be viewed in the dynamic track mode. Imaging data from microscopy experiments can also be displayed in the browser. In addition to software development, we continue to service and expand the data hubs we host for large consortia including 4DN, Roadmap Epigenomics, TaRGET and ENCODE, among others. Our growing user/developer community developed additional track types as plugins, such as qBed and dynseq tracks, which extend the utility of the browser. The browser serves as a foundation for additional genomics platforms including the WashU Virus Genome Browser (for COVID-19 research) and the Comparative Genome Browser. The WashU Epigenome Browser can also be accessed freely through Amazon Web Services at https://epigenomegateway.org/.
Graphical Abstract
Graphical Abstract
New components of WashU Epigenome Browser: 3D chromatin viewer, imaging data viewer and dynamic tracks.
Lineage-specific epigenomic changes during human corticogenesis have been difficult to study owing to challenges with sample availability and tissue heterogeneity. For example, previous studies using ...single-cell RNA sequencing identified at least 9 major cell types and up to 26 distinct subtypes in the dorsal cortex alone
. Here we characterize cell-type-specific cis-regulatory chromatin interactions, open chromatin peaks, and transcriptomes for radial glia, intermediate progenitor cells, excitatory neurons, and interneurons isolated from mid-gestational samples of the human cortex. We show that chromatin interactions underlie several aspects of gene regulation, with transposable elements and disease-associated variants enriched at distal interacting regions in a cell-type-specific manner. In addition, promoters with increased levels of chromatin interactivity-termed super-interactive promoters-are enriched for lineage-specific genes, suggesting that interactions at these loci contribute to the fine-tuning of transcription. Finally, we develop CRISPRview, a technique that integrates immunostaining, CRISPR interference, RNAscope, and image analysis to validate cell-type-specific cis-regulatory elements in heterogeneous populations of primary cells. Our findings provide insights into cell-type-specific gene expression patterns in the developing human cortex and advance our understanding of gene regulation and lineage specification during this crucial developmental window.
Full text
Available for:
FZAB, GEOZS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
BTB domain And CNC Homolog 2 (Bach2) is a transcription repressor that actively participates in T and B lymphocyte development, but it is unknown if Bach2 is also involved in the development of ...innate immune cells, such as natural killer (NK) cells. Here, we followed the expression of Bach2 during murine NK cell development, finding that it peaked in immature CD27
CD11b
cells and decreased upon further maturation. Bach2 showed an organ and tissue-specific expression pattern in NK cells. Bach2 expression positively correlated with the expression of transcription factor TCF1 and negatively correlated with genes encoding NK effector molecules and those involved in the cell cycle. Lack of Bach2 expression caused changes in chromatin accessibility of corresponding genes. In the end, Bach2 deficiency resulted in increased proportions of terminally differentiated NK cells with increased production of granzymes and cytokines. NK cell-mediated control of tumor metastasis was also augmented in the absence of Bach2. Therefore, Bach2 is a key checkpoint protein regulating NK terminal maturation.
Abstract Comparative genomics has revealed the rapid expansion of multiple gene families involved in immunity. Members within each gene family often evolved distinct roles in immunity. However, less ...is known about the evolution of their epigenome and cis -regulation. Here we systematically profile the epigenome of the recently expanded murine Ly49 gene family that mainly encode either inhibitory or activating surface receptors on natural killer cells. We identify a set of cis -regulatory elements (CREs) for activating Ly49 genes. In addition, we show that in mice, inhibitory and activating Ly49 genes are regulated by two separate sets of proximal CREs, likely resulting from lineage-specific losses of CRE activity. Furthermore, we find that some Ly49 genes are cross-regulated by the CREs of other Ly49 genes, suggesting that the Ly49 family has begun to evolve a concerted cis- regulatory mechanism. Collectively, we demonstrate the different modes of cis -regulatory evolution for a rapidly expanding gene family.
Chromatin priming promotes cell-type-specific gene expression, lineage differentiation, and development. The mechanism of chromatin priming has not been fully understood. Here, we report that mouse ...hematopoietic stem and progenitor cells (HSPCs) lacking the Baf155 subunit of the BAF (BRG1/BRM-associated factor) chromatin remodeling complex produce a significantly reduced number of mature blood cells, leading to a failure of hematopoietic regeneration upon transplantation and 5-fluorouracil (5-FU) injury. Baf155-deficient HSPCs generate particularly fewer neutrophils, B cells, and CD8+ T cells at homeostasis, supporting a more immune-suppressive tumor microenvironment and enhanced tumor growth. Single-nucleus multiomics analysis reveals that Baf155-deficient HSPCs fail to establish accessible chromatin in selected regions that are enriched for putative enhancers and binding motifs of hematopoietic lineage transcription factors. Our study provides a fundamental mechanistic understanding of the role of Baf155 in hematopoietic lineage chromatin priming and the functional consequences of Baf155 deficiency in regeneration and tumor immunity.
Display omitted
•Baf155 is needed for lineage-specific chromatin remodeling•Baf155 is critical for hematopoietic lineage differentiation•Baf155 is indispensable for hematopoietic regeneration•Baf155 deficiency compromises anti-tumor immunity
Wu et al. report that Baf155-deficient hematopoietic stem and progenitor cells show defects in lineage differentiation, likely due to the failure of transcription factors to open chromatin at target gene loci, leading to defects in hematopoietic regeneration and tumor control.
Full text
Available for:
GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Cryptic promoters within transposable elements (TEs) can be transcriptionally reactivated in tumors to create new TE-chimeric transcripts, which can produce immunogenic antigens. We performed a ...comprehensive screen for these TE exaptation events in 33 TCGA tumor types, 30 GTEx adult tissues and 675 cancer cell lines, and identified 1,068 TE-exapted candidates with the potential to generate shared tumor-specific TE-chimeric antigens (TS-TEAs). Whole-lysate and HLA-pulldown mass spectrometry data confirmed that TS-TEAs are presented on the surface of cancer cells. In addition, we highlight tumor-specific membrane proteins transcribed from TE promoters that constitute aberrant epitopes on the extracellular surface of cancer cells. Altogether, we showcase the high pan-cancer prevalence of TS-TEAs and atypical membrane proteins that could potentially be therapeutically exploited and targeted.
Full text
Available for:
GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
Abstract
NK cells exhibit memory properties after activation with haptens, viral infection, or cytokines with each stimulus having distinct biology. Memory-like (ML) NK cells differentiate over one ...week after brief activation with combined IL-12/15/18, and have augmented responses upon restimulation by cytokines, activating receptor, or tumor targets. While much is known about their functional characteristics, there are large gaps in our understanding of the molecular mechanisms involved in programming conventional (c)NK cells into ML NK cells. As ML NK cells have enhanced ability to produce IFN-γ after multiple cell divisions, this suggests that the IFNG gene, and others, may be epigenetically poised. Therefore, we hypothesize that activation with IL-12/15/18 triggers epigenetic remodeling of cNK cells to ML NK cells that drives enhanced functionality. To test this, we performed ATAC-seq on baseline cNK cells, IL-12/15/18 activated NK cells, and in vitro differentiated cNK and ML NK cells after one week. Activation with IL-12/15/18 induces dramatic early epigenetic remodeling with >10,000 differentially accessible regions (DARs) compared to cNK cells. Moreover, ML differentiation generates epigenetically distinct ML NK cells (>1,000 DARs) compared to in vitro differentiated cNK cells, including increased IFNG locus accessibility, suggesting epigenetic mechanisms regulate their enhanced functionality. Also, ML NK cells are markedly different from IL-12/15/18 activated NK cells (>15,000 DARs) suggesting not all epigenetic changes are immediately induced, and time is required for full implementation of the memory-like state. Together, this supports that ML NK cells rely on epigenetic programming for enhanced function.
Supported by grants from NIH (P50CA171963, R01CA205239, P30CA91842, 1F31GM146361-01).
NK effector functions can be triggered by inflammatory cytokines and engagement of activating receptors. NK cell production of IFN-γ, an important immunoregulatory cytokine, exhibits ...activation-specific IFN-γ regulation. Resting murine NK cells exhibit activation-specific metabolic requirements for IFN-γ production, which are reversed for activating receptor-mediated stimulation following IL-15 priming. Although both cytokine and activating receptor stimulation leads to similar IFN-γ protein production, only cytokine stimulation upregulates Ifng transcript, suggesting that protein production is translationally regulated after receptor stimulation. Based on these differences in IFN-γ regulation, we hypothesized that ex vivo IL-15 priming of murine NK cells allows a switch to IFN-γ transcription upon activating receptor engagement. Transcriptional analysis of primed NK cells compared with naive cells or cells cultured with low-dose IL-15 demonstrated that primed cells strongly upregulated Ifng transcript following activating receptor stimulation. This was not due to chromatin accessibility changes in the Ifng locus or changes in ITAM signaling, but was associated with a distinct transcriptional signature induced by ITAM stimulation of primed compared with naive NK cells. Transcriptional analyses identified a common signature of c-Myc (Myc) targets associated with Ifng transcription. Although Myc marked NK cells capable of Ifng transcription, Myc itself was not required for Ifng transcription using a genetic model of Myc deletion. This work highlights altered regulatory networks in IL-15-primed cells, resulting in distinct gene expression patterns and IFN-γ regulation in response to activating receptor stimulation.
NK cells are innate lymphoid cells critical for mediating anti-tumor responses and are being advanced as a cancer immunotherapy. While conventional (c) NK cell-based therapies have been explored in ...clinical trials, challenges related to cancer target recognition, in vivo persistence, and functionality result in limited activity. To overcome these challenges, NK cell memory programs have been investigated. Cytokine-induced memory-like (CIML or ML) NK cells are generated after brief activation with IL-12, IL-15, and IL-18, resulting in enhanced functionality, cytotoxicity, and in vivo persistence. ML NK cells have been advanced in multiple clinical trials for leukemia with promising results. Despite this translational progress, the underlying cellular dynamics and molecular mechanisms of ML reprogramming remain unclear. Elucidating these molecular programs may optimize NK therapeutics and reveal new translational strategies. We hypothesized that transcriptional and epigenetic mechanisms contribute to ML NK cell heterogeneity arising after IL-12/15/18 activation. To test this, NK cells from healthy donors were activated with IL-12/15/18 (to become ML) or IL-15 (to maintain cNK) and differentiated in vitro for 7 days (Fig 1, n=3-40 donors), followed by the assessment of their transcriptome (CITE-seq) and epigenome (ATAC-seq). CITE-seq clustering revealed unexpected heterogeneity with ~50% clustering with cNK cells (effector, effcNK) and ~50% forming a unique enriched ML cluster (eML). eML clusters were significantly increased in frequency in the IL-12/15/18 condition, compared to cNK (Mean±SEM=53±10.5% for IL-12/15/18, 6.2±1.1% for IL-15, p<0.0001) and discovered to expressed ENTPD1 (CD39). Based on CITE-seq, a flow cytometry strategy was developed to distinguish eML NK cells from effector cNK and cNK cells. Next, we identified eML NK cells 3-4 weeks after adoptive transfer of IL-12/15/18 activated donor NK cells across three different clinical trials (mean=13.3±1.9%, p<0.001). eML NK cells produced significantly more IFNg compared to cNK cells and effector cNK cells when stimulated with leukemia targets (Fig. 2, p<0.0001), and had enhanced cytotoxicity compared to cNK cells (EC50 of eML=0.862, EC50 of cNK=2.223, p<0.0001). Bulk ATAC-seq on flow-sorted populations revealed that eML NK cells have a distinct global epigenetic profile with >4,500 significant differentially accessible regions (DARs) compared to effcNK, while effcNK and cNK cells were epigenetically similar. Further, we interrogated which transcription factors are enriched in DARs of eML NK cells revealing TBOX and IRF family motifs. Within CITE-seq, the eML cluster was comprised of two subsets (eML-1 and eML-2). eML1 had increased expression of the transcription factors TCF7 and TOX2. In contrast, eML-2 had increased expression of IRF4 and PRDM1, as well as terminal exhaustion genes, including LAG3 and TIGIT (p<0.05). To evaluate eML-1 and eML-2 function, we performed CITE-seq after stimulation of eML NK cells with leukemia targets. eML-1 NK cells were the most functional, with significantly increased IFNG compared to eML-2, while eML-2 was hypofunctional and marked by high TOX expression (p<0.05). These findings suggest that optimizing strategies that lead to eML-1 differentiation may enhance the therapeutic activity of IL-12/15/18-induced ML NK cells. In summary, we identify heterogeneity following IL-12/15/18 activation of NK cells, resulting in multiple cellular fates. Further, these data identify an eML NK cell population with a distinct single-cell transcriptional and epigenetic program that directly manifests as enhanced NK cell functional response to leukemia.
Full text
Available for:
IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZRSKP
PDF
