Biology subjects

Kaminska, B.

Publications and source records attributed to Kaminska, B..

16 recordsLinked to original sources

bioRxiv2024

Integration of spatial transcriptomics with immunofluorescence staining reveals spatial heterogeneity and plasticity of astrocytes in experimental glioblastomas

Astrocytes comprise [~]50% of all brain cells and present distinct morphological, molecular and functional properties in different brain regions. In glioblastoma (GBM), an aggressive primary brain tumour, tumour-associated astrocytes (TAAs) become activated and exhibit different transcriptomic profiles, morphology and functions supporting disease progression. Heterogeneity and specific roles of TAAs within various regions of tumours are poorly known. Advancements of single-cell and spatial transcriptomics allow to profile tumours at unprecedented resolution revealing cell phenotypes, hidden functionalities and spatial architecture in disease-specific context. We combined spatial transcriptomics and multiple immunofluorescent staining to visualize TAAs heterogeneity and location of various subpopulations in intracranial murine gliomas. Using distinct gene expression profiles, we identified subtypes of TAAs with distinct localization and inferred their specialized functionalities. Gene signatures associated with TAAs reflected their reprograming in the tumour microenvironment (TME), revealed their multiple roles and potential contributing factors shaping the local milieu. Using spatial correlation analysis of the spots, we inferred the interactome of Slc1a2 (encoding a glutamate transporter) with the other markers of TAAs based on segregated areas of the tumour. The designer RGD peptide blocking tumour-microglia communications, alters the spatial distribution of TAAs in experimental gliomas providing insights into potential mechanisms. Spatial transcriptomics combined with multiple staining unveils multiple functional phenotypes of TAAs and interactions within TME. It shows their distinct morphology and unveils different roles in various regions of the tumour. We demonstrate the glioma-induced heterogeneity of TAAs and their adaption to the pharmacologically-induced modification of the immunosuppressive TME.

neuroscience↗

bioRxiv2024

Hypoxic stress dysregulates functions of glioma-associated myeloid cells through epigenomic and transcriptional programs

Hypoxia rapidly alters gene expression to allow cellular adaptation to challenging conditions and support tumour growth. Hypoxia also affects the chromatin structure by modifications of histones and DNA methylation. Glioblastoma (GBM) is an aggressive, deadly primary brain tumour for which there is no effective treatment. The tumour microenvironment of GBM is highly heterogeneous, with infiltration of glioma-associated microglia and macrophages (GAMs) and the presence of necrotic, hypoxic regions which significantly impair effectiveness of therapies. The mechanisms through which hypoxia alters the tumour microenvironment and regulates functions of infiltrating immune cells remain poorly understood. Here, we show that hypoxia modulates the expression of myeloid markers in distinct ways: upregulates the expression of monocytic marker Lgals3 and downregulates the microglial markers P2ry12 and Tmem119 in microglial and monocytic GAMs in vitro and in vivo. Underlying genome-wide hypoxia-dependent transcriptomic changes in microglial cells were identified using microglia-glioma co-cultures and validated in human and mouse GBM single- cell transcriptomics datasets. Numerous GAM subtype markers are dysregulated in response to hypoxic stress due to associated changes in chromatin accessibility, as determined using ATACseq. While hypoxia alone drives a decrease of the overall chromatin accessibility at gene promoters, the exposure to glioma cells under hypoxic conditions leads to both increases and decreases of chromatin accessibility at promoter regions in microglial cells. Hypoxia downregulates the chromatin accessibility at the regions enriched in motifs for transcription factors known as master regulators of microglial cell identity and function, including SPI1 or IRF8. Overall, our results highlight the importance of hypoxic stress as a strong intratumoral regulator of myeloid cell functions, which adds a new dimension to the characterisation of particular GAM subpopulations.

cancer biology↗

bioRxiv2024

Integrin blocking peptide reverses immunosuppression in experimental gliomas and improves anti-PD-1 therapy outcome

Immune checkpoint inhibitors (ICI) presented clinical benefits in many cancer patients but invariably fail in glioblastoma (GBM), the most common and deadly primary brain tumor. Lack of ICI efficacy in GBM is attributed to the accumulation of immunosuppressive myeloid cells that create the "cold" tumor microenvironment (TME) impeding infiltration and activation of effector T cells. We developed a designer RGD peptide that hindered glioma-instigated, integrin-mediated pro-tumoral reprogramming of myeloid cells and blocked microglia-dependent invasion of human and mouse glioma cells in co-cultures in vitro. Intratumorally-delivered RGD alone did not reduce glioma growth in syngeneic mice but prevented the emergence of immunosuppressive myeloid cells and led to peritumoral blood vessels normalization. Furthermore, combining RGD with immunotherapy using PD-1 blockade reduced tumor growth, led to upsurge of proliferating, interferon-{gamma} producing CD8+T cells and depleted regulatory T cells. Transcriptomic profiles of myeloid cells were altered by the combined treatment, consistently with the restored "hot" inflammatory TME and boosted immunotherapy responses. RGD modified the phenotypes of myeloid cells in human gliomas in nude mice. Thus, combining the integrin blockade with ICI reinvigorates antitumor immunity and paves the way to improve immunotherapy outcomes in GBM.

cancer biology↗

bioRxiv2024

Tracking changes in functionality and morphology of repopulated microglia in young and old mice

Microglia (MG) are myeloid cells of the central nervous system supporting its homeostasis and instigating neuroinflammation in pathologies. Single-cell RNA sequencing (scRNA-seq) revealed the functional heterogeneity of MG in mice brains. Inhibition of colony-stimulating factor 1 receptor (CSF1R) signaling with inhibitors deplete microglia which rapidly repopulate. Functionalities of repopulated microglia are poorly known. We combined scRNA-seq, bulk RNA-seq, immunofluorescence and confocal imaging to study functionalities and morphology of repopulated microglia. CSRF1R inhibitor (BLZ-945) depleted MG in 21 days and their numbers were restored 7 days later as evidenced by TMEM119 staining and flow cytometry. ScRNA-seq and computational analyses demonstrate that repopulated MG originate from preexisting MG progenitors and reconstitute functional clusters but upregulate inflammatory genes. Percentages of proliferating, immature MG displaying inflammatory gene expression increase in aging mice. Morphometric analysis of MG cell body and branching shows distinct morphology of repopulated MG, particularly in old mouse brains. We demonstrate that with aging some repopulated MG fail to reach the homeostatic phenotype. These differences microglia may contribute to the deterioration of microglia protective functions with age.

neuroscience↗

bioRxiv2023

scRNA-seq reveals the diversity of the developing cardiac cell lineage and molecular building blocks of the primary pacemaker

The heart is comprised of a variety of specialized cell types that work in unison to maintain blood flow. Here we utilized scRNA-seq analysis to delineate the diversity of cardiac cell types in the zebrafish. With the growing use of the zebrafish to model human heart biology, a deeper insight into its complex cellular composition is critical for a better understanding of heart function, development, and associated malformations. We present a high resolution atlas of zebrafish heart single cells transcriptomics, consisting of over 50 000 cells representing the building blocks of the zebrafish heart at 48 and 72 hpf. We defined 18 discrete cell populations comprising major cell lineages and sublineages of the developing heart. We pinpointed a population of cells likely to be the primary pacemaker and identified the transcriptome profile defining this critical cell type. Our analyses identified two genes, atp1b3b and colec10, which were enriched in the sinoatrial pacemaker cells. CRISPR/Cas9-mediated knockout of these two genes significantly reduced heart rate which is accompanied by arrhythmia or morphological defects, suggesting their novel function in cardiac development and conduction. Additionally, we describe other subpopulations of cardiac cell lineages, including the endothelial and neural cells, whose expression profiles we provide as a resource for further investigations into the cellular and molecular mechanisms of this organ.

developmental biology↗

bioRxiv2023

Deep Learning Glioma Grading with the Tumor Microenvironment Analysis Protocol for A Comprehensive Learning, Discovering, and Quantifying Microenvironmental Features

Gliomas are primary brain tumors that arise from neural stem cells or glial precursors. Diagnosis of glioma is based on histological evaluation of pathological cell features and molecular markers. Gliomas are infiltrated by myeloid cells that accumulate preferentially in malignant tumors and their abundance inversely correlates with survival, which is of interest for cancer immunotherapies. To avoid time-consuming and laborious manual examination of the images, a deep learning approach for automatic multiclass classification of tumor grades was proposed. Importantly, as an alternative way of investigating characteristics of brain tumor grades, we implemented a protocol for learning, discovering, and quantifying tumor microenvironment elements on our glioma dataset. Using only single-stained biopsies we derived characteristic differentiating tumor microenvironment phenotypic neighborhoods. A challenge of the study was given by a small sample size of human leukocyte antigen stained on glioma tissue microarrays dataset - 203 images from 5 classes - and imbalanced data distribution. This has been addressed by image augmentation of the underrepresented classes. For this glioma multiclass classification task, a residual neural network architecture has been adapted. On the validation set the average accuracy was 0.72 when the model was trained from scratch, and 0.85 with the pre-trained model. Moreover, the tumor microenvironment analysis suggested a relevant role of the myeloid cells and their accumulation to characterize glioma grades. This promising approach can be used as an additional diagnostic tool to improve assessment during intra-operative examination or sub-typing tissues for treatment selection, despite the challenges caused by the difficult dataset. We present here the distributions and visualizations of extracted tumor inter-dependencies. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/544739v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@a85f48org.highwire.dtl.DTLVardef@7a49forg.highwire.dtl.DTLVardef@1c8df75org.highwire.dtl.DTLVardef@c6bad7_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIResearch highlight 1: We demonstrate that the ResNet-18 architecture with simple data augmentation trained in 10-fold cross-validation performs the multiclass classification relatively well even with a small imbalanced dataset with a high degree of similarities between classes. C_LIO_LIResearch highlight 2: After supervised subtyping of the tumor, we investigated the usefulness of the protocol for discovery and learning tumor microenvironment elements for the same task. The protocol designed for deriving new biomarkers based on multiplex stained histological samples proved the ability to detect features characteristic of malignant tumors based only on single target stained tissue microarrays. We propose further studies on this topic can help in formulating specific criteria for improvements in diagnosis of gliomas, allowing to avoid the necessity of conducting advanced histopathological analysis or complementing genetic testing of tumor samples. C_LI

bioinformatics↗

bioRxiv2023

H2A.Z histone variants facilitate HDACi-dependent removal of H3.3K27M mutant protein in paediatric high-grade glioma cells

Diffuse intrinsic pontine gliomas (DIPG) are deadly paediatric brain tumours, non-resectable due to brainstem localisation and diffusive growth. Patients with DIPG have a dismal prognosis of 9-12 months of survival with no effective therapy. Over 80% of DIPGs harbour a mutation in histone 3 (H3.3 or H3.1) resulting in a lysine to methionine substitution (H3K27M). H3K27M causes global epigenetic alterations (a loss of H3K27 trimethylation and an increase in H3K27 acetylation) resulting in aberrant gene expression. To date, no therapeutic strategy exists to suppress the levels of oncogenic H3K27M. We show that pan-HDAC inhibitors (HDACi) lead to the temporary but significant reduction in the H3.3K27M protein (up to 80%) in multiple glioma cell lines expressing the H3.3K27M histone variant, without changes in the H3F3A mRNA expression. The H3.3K27M occupancy at the chromatin is greatly reduced upon HDACi (SB939) treatment, as shown by ChIPseq analysis. H3.3K27M loss is most striking at SB939-upregulated genes suggesting the role in repression of these genes. In addition, genes previously reported as H3K27M-dependent become downregulated in response to SB939 treatment. We discover that the SB939-mediated loss of H3.3K27M is partially blocked by a lysosomal inhibitor, chloroquine. Moreover, the loss of H3.3K27M is facilitated by co-occurrence of H2A.Z, as evidenced by the knock-down of H2A.Z histone isoforms. ChIPseq analysis confirms the occupancy of H3.3K27M and H2A.Z at the same SB939-inducible genes. Altogether, we provide new insight into disease-specific mechanism of HDAC inhibition and demonstrate pharmacological modulation of the oncogenic H3.3K27M protein levels. These findings open a new possibility to directly target the H3.3K27M oncohistone, which may be exploited in future therapies.

cancer biology↗

bioRxiv2023

SorLA restricts TNFalpha release from microglia to shape glioma-supportive brain microenvironment

SorLA, encoded by the gene SORL1, is an intracellular sorting receptor of the VPS10P domain receptor gene family. Although SorLA is best recognized for its ability to shuttle target proteins between intracellular compartments in neurons, recent data suggest that also its microglial expression can be of high relevance for the pathogenesis of brain diseases, including glioblastoma (GBM). Here we interrogated the impact of SorLA on the functional properties of glioma-associated microglia and macrophages (GAMs). In the GBM microenvironment, GAMs are re-programmed and in turn lose the ability to elicit anti-tumor responses. Instead, they acquire glioma-supporting phenotype, which is a key mechanism promoting glioma progression. Our analysis of scRNA-seq data from GBM patients revealed that the pro-tumorigenic and pro-inflammatory properties of GAMs are linked to high and low SORL1 expression, respectively. Using cell models, we confirm that SorLA levels are differentially regulated by the presence of glioma cells and by inflammatory cues. We further show that SorLA acts as a sorting receptor for the pro-inflammatory cytokine TNF to restrain its secretion from microglia. As a consequence, loss of SorLA enhanced the pro-inflammatory potential of microglia, having a remarkable impact on glioma progression. In a murine model of glioma, SorLA-deficient mice develop smaller tumors and show hallmarks of anti-tumor response including altered microglia morphology, enhanced necroptosis, and massive neutrophil influx into the tumor parenchyma. Our findings indicate that SorLA is a key player in shaping the phenotype of GAMs, and its depletion can unlock an anti-tumor response. Significance statementOur study provides insight into the mechanisms shaping the tumor microenvironment in glioblastoma (GBM), the most prevalent and aggressive brain malignancy in adults. Poor prognosis in GBM largely results from the properties of the glioma milieu that blocks the anti-tumor response. We show that SorLA restricts release of the pro-inflammatory cytokine TNF from microglia, thereby hampering their anti-glioma response. SorLA depletion reinforces the pro-inflammatory properties of tumor microenvironment and inhibits glioma growth. These findings have significant implications for our understanding of glioma biology, indicating SorLA-TNF interaction as a potential target in GBM therapies. They also offer a new perspective on SorLA activities in microglia which emerge as highly relevant not only for the pathogenesis GBM, but also of other brain diseases such as Alzheimers disease.

neuroscience↗

bioRxiv2022

Comprehensive analysis of the transcription factor REST regulatory networks in IDH-mutant and IDH-wild type glioma

REST is a widely expressed, dual role transcription factor that acts either as a transcriptional repressor or transcriptional activator depending on the genomic and cellular context. REST is an important oncogenic factor, a key player in brain cell differentiation and has a role in establishing DNA methylation status in proximity of its binding sites. Mutations in IDH cause significant changes to the epigenome contributing to blocking cell differentiation and are considered an oncogenic driver in glioma. We aimed at defining the REST role in the IDH mutation-related phenotype in gliomas accounting for its role in gene activation and repression. We studied the effects of REST knockdown, REST binding sites, and REST motifs methylation in context of IDH mutation, and found that both REST binding patterns and TF motif composition proximal to REST binding sites differed in IDH wild-type and mutant glioma. Among such REST targets were genes involved in glial cell differentiation and ECM organization. REST knockdown differently impacted glioma cell invasion depending on the IDH phenotype. DNA methylation of REST activated gene promoters showed positive correlation with gene expression. The canonical REST-repressed gene targets correlated with NPC-like cellular state properties in IDH-MUT grade 2/3 gliomas. The identified REST targets, gene regulatory networks and putative REST cooperativity with other TFs point to differential control of REST target gene expression in IDH-WT and IDH-MUT gliomas. We conclude that REST could be considered as a key factor in the design of targeted glioma therapies.

cancer biology↗

bioRxiv2022

Regulatory networks driving expression of genes critical for glioblastoma are controlled by the transcription factor c-Jun and the pre-existing epigenetic modifications.

BackgroundGlioblastoma (GBM, WHO grade IV) is an aggressive, primary brain tumor. Despite gross surgery and forceful radio- and chemotherapy, survival of GBM patients did not improve over decades. Several studies reported transcription deregulation in GBMs but regulatory mechanisms driving overexpression of GBM-specific genes remain largely unknown. Transcription in open chromatin regions is directed by transcription factors (TFs) that bind to specific motifs, recruit co-activators/repressors and the transcriptional machinery. Identification of GBM-related TFs-gene regulatory networks may reveal new and targetable mechanisms of gliomagenesis. ResultsWe predicted TFs-regulated networks in GBMs in silico and intersected them with putative TF binding sites identified in the accessible chromatin in human glioma cells and GBM patient samples. The Cancer Genome Atlas and Glioma Atlas datasets (DNA methylation, H3K27 acetylation, transcriptomic profiles) were explored to elucidate TFs-gene regulatory networks and effects of the epigenetic background. In contrast to the majority of tumors, c-Jun expression was higher in GBMs than in normal brain and c-Jun binding sites were found in multiple genes overexpressed in GBMs such as VIM, FOSL2 or UPP1. Binding of c-Jun to the VIM gene promoter is stronger in GBM cells than in cells derived from benign glioma as evidenced by gel shift and supershift assays. Regulatory regions of a majority of the c-Jun targets have distinct DNA methylation in GBMs suggesting the contribution of DNA methylation to the c-Jun-dependent regulation. ConclusionsWe identified distinct TFs-gene networks in GBMs compared to benign gliomas, a predominant role of c-Jun in controlling genes driving gliomagenesis and a modulatory role of DNA methylation.

cancer biology↗

bioRxiv2022

Hypoxia-mediated regulation of DDX5 through decreased chromatin accessibility and post-translational targeting restricts R-loop accumulation

Local hypoxia occurs in most solid tumors and is associated with aggressive disease and therapy resistance. Widespread changes in gene expression play a critical role in the biological response to hypoxia. However, most research has focused on hypoxia-inducible genes as opposed to those which are decreased in hypoxia. We demonstrate that chromatin accessibility is decreased in hypoxia, predominantly at gene promoters and specific pathways are impacted including DNA repair, splicing and the R-loop interactome. One of the genes with decreased chromatin accessibility in hypoxia was DDX5, encoding the RNA helicase, DDX5, which showed reduced expression in various cancer cell lines in hypoxic conditions, tumor xenografts and in patient samples with hypoxic tumors. Most interestingly, we found that when DDX5 is rescued in hypoxia, replication stress and R-loop levels accumulate further, demonstrating that hypoxia-mediated repression of DDX5 restricts R-loop accumulation. Together these data support the hypothesis that a critical part of the biological response to hypoxia is the repression of multiple R-loop processing factors, however, as shown for DDX5, their role is specific and distinct.

cancer biology↗

bioRxiv2021

ApoE4 disrupts interaction of sortilin with fatty acid-binding protein 7 essential to promote lipid signaling

Sortilin is a receptor for neuronal uptake of apolipoprotein E. Sortilin-dependent uptake of lipidated apoE promotes conversion of polyunsaturated fatty acids (PUFA) into neuromodulators that induce anti-inflammatory gene expression in the brain. This neuroprotective pathway works with apoE3 but is lost with apoE4, the main risk factor for Alzheimers disease (AD). Here, we elucidated steps in cellular handling of lipids through sortilin, and why they are disrupted by apoE4. Combining unbiased proteome screens with analyses in mouse models, we uncover interaction of sortilin with fatty acid-binding protein (FABP) 7, the intracellular carrier for PUFA in the brain. In the presence of apoE3, sortilin promotes functional expression of FABP7 and its ability to elicit lipid-dependent gene transcription. By contrast, apoE4 binding blocks sortilin sorting, causing catabolism of FABP7 and impairing lipid signaling. Reduced FABP7 levels in the brain of AD patients expressing apoE4 substantiate the relevance of these interactions for neuronal lipid homeostasis. Taken together, we document interaction of sortilin with mediators of extracellular and intracellular lipid transport that provides a mechanistic explanation for loss of a neuroprotective lipid metabolism in AD. SUMMARY STATEMENTLipids are central to brain health and defects in brain lipid homeostasis are causal to neurodegenerative processes in Alzheimers disease. Here, we uncovered how the neuronal lipoprotein receptor sortilin interacts with apoE and FABP7, the carriers for extra- and intracellular transport of lipids in the brain, respectively. We show that this interaction enables lipids to control gene transcription via nuclear receptors; and why this presumed neuroprotective lipid action is disturbed in humans who carry the {varepsilon}4 variant of apoE, the most important risk factor for sporadic Alzheimers disease.

cell biology↗

bioRxiv2020

Epigenetic regulation of differentially expressed genes between various glioma types

Gliomas are the most frequent primary tumors of the central nervous system (CNS) and encompass two major subgroups: diffuse, malignant gliomas and benign, well differentiated gliomas showing a more circumscribed growth. Genome-wide next generation sequencing studies have uncovered specific genetic alterations, transcriptomic patterns and epigenetic profiles associated with different types of gliomas improving tumor diagnosis and having important implications for future clinical trials and patient management. We have recently created a unique resource encompassing genome-wide profiles of open chromatin, histone H3K27ac and H3Kme3 modifications, DNA methylation and transcriptomes of 33 glioma samples of different grades. Here, we took advantage of a wealth of data from those high-throughput experiments, intersected those data with topologically associating domains (TADs) and demonstrated that the chromatin organization and epigenetic landscape of enhancers have a strong impact on genes differentially expressed in low grade versus high grade gliomas. We identified TADs enriched in glioma grade-specific genes and/or epigenetic marks. We found a set of transcription factors, including REST, E2F1 and NFKB1, that are most likely to regulate gene expression in multiple TADs, containing glioma-related genes. Moreover, many genes associated with the cell-matrix adhesion Gene Ontology group, in particular 14 PROTOCADHERINs, were found to be regulated by the long range contacts with enhancers. Overall, the results presented here demonstrate the existence of epigenetic differences associated with chromatin organization driving differential gene expression in gliomas of different malignancy. We demonstrated that integration of whole genome epigenetic data with Hi-C data and transcriptomic profiles described in this work, can segregate low and high grade gliomas and reveal new regulatory networks that could explain some of the functional differences between gliomas of different malignancies. HighlightsO_LIIntegration of ATAC-seq, ChIP-seq and RNA-seq reveals glioma malignancy-related gene regulatory networks. C_LIO_LITADs segmentation contributes to gene-epigenetically modified enhancer relationships. C_LIO_LIContacts of active enhancers in gliomas of different malignancies might affect expression of genes involved in cancerogenesis, such as PROTOCADHERINs or EGFR. C_LI

cancer biology↗

bioRxiv2020

TOP2A substitution enhances topoisomerase activity and causes transcriptional dysfunction in glioblastoma patients

BackgroundHigh grade gliomas (HGGs) are aggressive, primary brain tumors with poor clinical outcomes. We aim to better understand glioma pathobiology and find potential therapeutic susceptibilities. MethodsWe designed a custom panel of 664 cancer- and epigenetics-related genes, and employed targeted next generation sequencing to study the genomic landscape of somatic and germline variants in 182 gliomas of different malignancy grades. mRNA sequencing was performed to detect transcriptomic abnormalities. ResultsIn addition to known alterations in TP53, IDH1, ATRX, EGFR genes found in this cohort, we identified a novel, recurrent mutation in the TOP2A gene coding for Topoisomerase 2A occurring only in glioblastomas (GBM, WHO grade IV gliomas). Biochemical assays with recombinant proteins demonstrated stronger DNA binding and DNA supercoil relaxation activities of the variant proteins. GBM patients carrying the mutated TOP2A had shorter overall survival than those with the wild type TOP2A. Computational analyses of transcriptomic data showed that GBMs with the mutated TOP2A have different transcriptomic patterns suggesting higher transcriptomic activity. ConclusionWe identified a novel TOP2A E948Q variant that strongly binds to DNA and is more active than the wild type protein. Our findings suggest that the discovered TOP2A variant is gain-of-function mutation. Key pointsO_LIThe most frequent genetic alterations in high grade gliomas are reported. C_LIO_LIA new mutation in the TOP2A gene was found in 4 patients from Polish population. C_LIO_LIA E948Q substitution changes TOP2A activities towards DNA. C_LIO_LIThe recurrent TOP2A variant is a gain-of-function mutation. C_LI Importance of the studyGlioblastoma is a deadly disease. Despite recent advancements in genomics and innovative targeted therapies, glioblastoma therapy has not shown improvements. Insights into glioblastoma biology may improve diagnosis, prognosis, and treatment prediction, directing to a better outcome. We performed targeted sequencing of 664 cancer genes, and identified a new variant of the TOP2A gene encoding topoisomerase 2A in glioblastomas. The TOP2A protein variant shows a higher affinity towards DNA and causes transcriptional alterations, suggesting a higher de novo transcription rate.

cancer biology↗

bioRxiv2019

Mapping chromatin accessibility and active regulatory elements reveals new pathological mechanisms in human gliomas

Chromatin structure and accessibility, and combinatorial binding of transcription factors to regulatory elements in genomic DNA control transcription. Genetic variations in genes encoding histones, epigenetics-related enzymes or modifiers affect chromatin structure/dynamics and result in alterations in gene expression contributing to cancer development or progression. Gliomas are brain tumors frequently associated with epigenetics-related gene deregulation. We performed whole-genome mapping of chromatin accessibility, histone modifications, DNA methylation patterns and transcriptome analysis simultaneously in multiple tumor samples to unravel novel epigenetic dysfunctions driving gliomagenesis. Based on the results of the integrative analysis of the acquired profiles, we created an atlas of active enhancers and promoters in benign and malignant gliomas. We explored these elements and intersected with Hi-C data to uncover molecular mechanisms instructing gene expression in gliomas. SignificanceEpigenetics-driven deregulation of gene expression accompanies cancer development, but its comprehensive characterization in cancer patients is fragmentary. We performed whole-genome profiling of gene expression, open chromatin, histone modifications and DNA-methylation profiles in the same samples from benign and malignant gliomas. Our study provides a first comprehensive atlas of active regulatory elements in gliomas, which allowed identification of the functional enhancers and promoters in patient samples. This comprehensive approach revealed epigenetic patterns influencing gene expression in benign gliomas and a new pathogenic mechanism involving FOXM1-driven network in glioblastomas. This atlas provides a common set of elements for cross-comparisons of existing and new datasets, prompting novel discoveries and better understanding of gliomagenesis. HighlightsO_LIWe provide an atlas of cis-regulatory elements active in human gliomas C_LIO_LIEnhancer-promoter contacts operating in gliomas are revealed C_LIO_LIDiverse enhancer activation is pronounced in malignant gliomas C_LIO_LIChromatin loop activates FOXM1-ANXA2R pathological network in glioblastomas. C_LI

cancer biology↗

bioRxiv2019

Single-cell RNA sequencing reveals functional heterogeneity and sex differences of glioma-associated brain macrophages

Microglia are resident myeloid cells in the central nervous system (CNS) that control homeostasis and protect CNS from damage and infections. Microglia and peripheral myeloid cells accumulate and adapt tumor supporting roles in human glioblastomas that show prevalence in men. Cell heterogeneity and functional phenotypes of myeloid subpopulations in gliomas remain elusive. Single-cell RNA sequencing (scRNA-seq) of CD11b+ myeloid cells in naive and GL261 glioma-bearing mice revealed distinct profiles of microglia, infiltrating monocytes/macrophages and CNS border-associated macrophages. We demonstrated an unforeseen molecular heterogeneity among myeloid cells in naive and glioma-bearing brains, validated selected marker proteins and showed distinct spatial distribution of identified subsets in experimental gliomas. We found higher expression of MHCII encoding genes in glioma-activated male microglia, which was corroborated in bulk and scRNA-seq data from human diffuse gliomas. Sex-specific gene expression in glioma-activated microglia may be relevant to sex differences in incidence and outcomes of glioma patients.

cancer biology↗