Gene Expression Programs In Cells Research Articles

Genetic deletion of JAM-C in preleukemic cells rewires leukemic stem cell gene expression program in AML

AbstractThe leukemic stem cell (LSC) score LSC-17 based on a stemness–related gene expression signature is an indicator of poor disease outcome in acute myeloid leukemia (AML). However, it is not known whether “niche anchoring” of LSC affects disease evolution. To address this issue, we conditionally inactivated the adhesion molecule JAM-C (Junctional Adhesion Molecule-C) expressed by hematopoietic stem cells (HSCs) and LSCs in an inducible mixed-lineage leukemia (iMLL)-AF9–driven AML mouse model. Deletion of Jam3 (encoding JAM-C) before induction of the leukemia–initiating iMLL-AF9 fusion resulted in a shift from long-term to short-term HSC expansion, without affecting disease initiation and progression. In vitro experiments showed that JAM-C controlled leukemic cell nesting irrespective of the bone marrow stromal cells used. RNA sequencing performed on leukemic HSCs isolated from diseased mice revealed that genes upregulated in Jam3-deficient animals belonged to activation protein-1 (AP-1) and tumor necrosis factor α (TNF-α)/NF-κB pathways. Human orthologs of dysregulated genes allowed to identify a score that was distinct from, and complementary to, the LSC-17 score. Substratification of patients with AML using LSC-17 and AP-1/TNF-α genes signature defined 4 groups with median survival ranging from <1 year to a median of “not reached” after 8 years. Finally, coculture experiments showed that AP-1 activation in leukemic cells was dependent on the nature of stromal cells. Altogether, our results identify the AP-1/TNF-α gene signature as a proxy of LSC anchoring in bone marrow niches, which improves the prognostic value of the LSC-17 score. This trial was registered at www.ClinicalTrials.gov as #NCT02320656.

Abstract SY16-01: What IDH mouse models have taught us about hematological diseases, from leukemia to T cell lymphoma

Abstract SY16-01: What IDH mouse models have taught us about hematological diseases, from leukemia to T cell lymphoma

Abstract 1687: Targeted degradation of MECOM in high-risk acute myeloid leukemia reveals a novel repressive function that is amenable to therapeutic small-molecule rewiring

Abstract Hematopoiesis relies on an intricate balance between stem-cell self-renewal and terminal maturation of blood cells. Disrupting this balance can lead to the development of deadly blood cancers, like acute myeloid leukemia (AML). Increasing evidence suggests that acquiring hematopoietic stem cell (HSC) gene expression programs in AML confers a particularly poor prognosis and increases risk of relapse with conventional therapy. Acquisition of these features in AML is commonly driven by dysregulation of MECOM, a transcription factor and master regulator of HSC self-renewal. To define the role of MECOM in AMLs, we engineered cell-line models of high-risk AML by endogenously tagging MECOM with an FKBP12F36V degron. These models enable targeted degradation of MECOM which we employed in concert with multi-omic readouts to assess changes in nascent transcription and chromatin accessibility to identify direct transcriptional targets of MECOM. We demonstrate that MECOM degradation results in a rapid increase in chromatin accessibility at MECOM-bound sites and increased expression of cognate genes to establish a pro-differentiation gene program. These findings suggest a previously unappreciated mechanism for MECOM in repressing differentiation-promoting cis-regulatory elements (cisREs) and their cognate target genes in AML. We then employed a pooled CRISPR inhibition (CRISPRi) screen to functionally characterize the role that each cisRE plays in promoting high-risk features in AML. Utilizing CRISPRi to repress MECOM-bound cisREs in concert with dTAG-mediated MECOM degradation, we examined if repression of over 500 individual MECOM-bound cisREs is sufficient to maintain AML progenitors in the absence of MECOM. This functional dissection revealed that the individual repression of three cisREs that regulate CEBPA, GFI1B, and RUNX1 is sufficient to maintain AML progenitor cells in the absence of MECOM. This result implicates MECOM as a direct repressor of known regulators of myeloid differentiation. Given these findings, we hypothesized that small molecule-based recruitment of a transcriptional coactivator to MECOM could rewire its repressive function to activate myeloid differentiation and subsequent leukemia cell death. To test this approach, we treated a MECOM-FKBP12F36V AML degron model with the bifunctional small molecule NICE-01 (AP1867-PEG2-JQ1) to recruit an epigenetic activator (BRD4) to MECOM to functionally rewire this transcriptional network. NICE-01 treatment induced significant differentiation of AML progenitor cells relative to JQ1 treatment alone. Our work highlights the utility of targeted protein degradation to mechanistically interrogate the function of a key driver of high-risk AMLs and suggests the potential for small molecule-rewiring of stem cell gene regulatory networks to confer therapeutic benefit. Citation Format: Travis J. Fleming, Michael Gundry, Richard Voit, Mateusz Antoszewski, William J. Gibson, Ananthan Sadagopan, Vijay G. Sankaran. Targeted degradation of MECOM in high-risk acute myeloid leukemia reveals a novel repressive function that is amenable to therapeutic small-molecule rewiring [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1687.

Highly sensitive spatial transcriptomics using FISHnCHIPs of multiple co-expressed genes

High-dimensional, spatially resolved analysis of intact tissue samples promises to transform biomedical research and diagnostics, but existing spatial omics technologies are costly and labor-intensive. We present Fluorescence In Situ Hybridization of Cellular HeterogeneIty and gene expression Programs (FISHnCHIPs) for highly sensitive in situ profiling of cell types and gene expression programs. FISHnCHIPs achieves this by simultaneously imaging ~2-35 co-expressed genes (clustered into modules) that are spatially co-localized in tissues, resulting in similar spatial information as single-gene Fluorescence In Situ Hybridization (FISH), but with ~2-20-fold higher sensitivity. Using FISHnCHIPs, we image up to 53 modules from the mouse kidney and mouse brain, and demonstrate high-speed, large field-of-view profiling of a whole tissue section. FISHnCHIPs also reveals spatially restricted localizations of cancer-associated fibroblasts in a human colorectal cancer biopsy. Overall, FISHnCHIPs enables fast, robust, and scalable cell typing of tissues with normal physiology or undergoing pathogenesis.

Plasma Cell Differentiation, Antibody Quality, and Initial Germinal Center B Cell Population Depend on Glucose Influx Rate.

Serum Ab concentrations, selection for higher affinity BCRs, and generation of higher Ab affinities are important elements of immune response optimization and functions of germinal center (GC) reactions. B cell proliferation requires nutrients to support the anabolism inherent in clonal expansion. Glucose usage by mouse GC B cells has been reported to contribute little to their energy needs, with questions raised as to whether glucose uptake or glycolysis increases in GC B cells compared with their naive precursors. Indeed, metabolism can be highly flexible, such that supply shortage along one pathway may be compensated by increased flux on others. We now show that reduction of the glucose transporter GLUT1 in mice after establishment of a preimmune B cell repertoire, even after initiation of the GC B cell gene expression program, decreased initial GC B cell population numbers, affinity maturation, and plasma cell outputs. Glucose oxidation was heightened in GC B cells, but this hexose flowed more into the pentose phosphate pathway, whose activity was important in controlling reactive oxygen species (ROS) and Ab-secreting cell production. In modeling how glucose usage by B cells promotes the Ab response, the control of ROS appeared insufficient. Surprisingly, the combination of galactose, which mitigated ROS, with provision of mannose, an efficient precursor to glycosylation, supported robust production of and normal Ab secretion by Ab-secreting cells under glucose-free conditions. Collectively, the findings indicate that GCs depend on normal glucose influx, especially in plasma cell production, but reveal an unexpected metabolic flexibility in hexose requirements.

Enhancers dysfunction in the 3D genome of cancer cells.

Eukaryotic genomes are spatially organized inside the cell nucleus, forming a threedimensional (3D) architecture that allows for spatial separation of nuclear processes and for controlled expression of genes required for cell identity specification and tissue homeostasis. Hence, it is of no surprise that mis-regulation of genome architecture through rearrangements of the linear genome sequence or epigenetic perturbations are often linked to aberrant gene expression programs in tumor cells. Increasing research efforts have shed light into the causes and consequences of alterations of 3D genome organization. In this review, we summarize the current knowledge on how 3D genome architecture is dysregulated in cancer, with a focus on enhancer highjacking events and their contribution to tumorigenesis. Studying the functional effects of genome architecture perturbations on gene expression in cancer offers a unique opportunity for a deeper understanding of tumor biology and sets the basis for the discovery of novel therapeutic targets.

A snoRNA–tRNA modification network governs codon-biased cellular states

The dynamic balance between tRNA supply and codon usage demand is a fundamental principle in the cellular translation economy. However, the regulation and functional consequences of this balance remain unclear. Here, we use PARIS2 interactome capture, structure modeling, conservation analysis, RNA-protein interaction analysis, and modification mapping to reveal the targets of hundreds of snoRNAs, many of which were previously considered orphans. We identify a snoRNA-tRNA interaction network that is required for global tRNA modifications, including 2'-O-methylation and others. Loss of Fibrillarin, the snoRNA-guided 2'-O-methyltransferase, induces global upregulation of tRNA fragments, a large group of regulatory RNAs. In particular, the snoRNAs D97/D133 guide the 2'-O-methylation of multiple tRNAs, especially for the amino acid methionine (Met), a protein-intrinsic antioxidant. Loss of D97/D133 snoRNAs in human HEK293 cells reduced target tRNA levels and induced codon adaptation of the transcriptome and translatome. Both single and double knockouts of D97 and D133 in HEK293 cells suppress Met-enriched proliferation-related gene expression programs, including, translation, splicing, and mitochondrial energy metabolism, and promote Met-depleted programs related to development, differentiation, and morphogenesis. In a mouse embryonic stem cell model of development, knockdown and knockout of D97/D133 promote differentiation to mesoderm and endoderm fates, such as cardiomyocytes, without compromising pluripotency, consistent with the enhanced development-related gene expression programs in human cells. This work solves a decades-old mystery about orphan snoRNAs and reveals a function of snoRNAs in controlling the codon-biased dichotomous cellular states of proliferation and development.

Novel insights into the role of acetyl-CoA producing enzymes in epigenetic regulation.

Inflammation-dependent changes in gene expression programs in innate immune cells, such as macrophages, involve extensive reprogramming of metabolism. This reprogramming is essential for the production of metabolites required for chromatin modifications, such as acetyl-CoA, and regulate their usage and availability impacting the macrophage epigenome. One of the most transcriptionally induced proinflammatory mediator is nitric oxide (NO), which has been shown to inhibit key metabolic enzymes involved in the production of these metabolites. Recent evidence indicates that NO inhibits mitochondrial enzymes such as pyruvate dehydrogenase (PDH) in macrophages induced by inflammatory stimulus. PDH is involved in the production of acetyl-CoA, which is essential for chromatin modifications in the nucleus, such as histone acetylation. In addition, acetyl-CoA levels in inflamed macrophages are regulated by ATP citrate lyase (ACLY) and citrate transporter SLC25A1. Interestingly, acetyl-CoA producing enzymes, such as PDH and ACLY, have also been reported to be present in the nucleus and to support the local generation of cofactors such as acetyl-CoA. Here, we will discuss the mechanisms involved in the regulation of acetyl-CoA production by metabolic enzymes, their inhibition by prolonged exposure to inflammation stimuli, their involvement in dynamic inflammatory expression changes and how these emerging findings could have significant implications for the design of novel therapeutic approaches.

A lamprey neural cell type atlas illuminates the origins of the vertebrate brain

The vertebrate brain emerged more than ~500 million years ago in common evolutionary ancestors. To systematically trace its cellular and molecular origins, we established a spatially resolved cell type atlas of the entire brain of the sea lamprey—a jawless species whose phylogenetic position affords the reconstruction of ancestral vertebrate traits—based on extensive single-cell RNA-seq and in situ sequencing data. Comparisons of this atlas to neural data from the mouse and other jawed vertebrates unveiled various shared features that enabled the reconstruction of cell types, tissue structures and gene expression programs of the ancestral vertebrate brain. However, our analyses also revealed key tissues and cell types that arose later in evolution. For example, the ancestral brain was probably devoid of cerebellar cell types and oligodendrocytes (myelinating cells); our data suggest that the latter emerged from astrocyte-like evolutionary precursors in the jawed vertebrate lineage. Altogether, our work illuminates the cellular and molecular architecture of the ancestral vertebrate brain and provides a foundation for exploring its diversification during evolution.

EIF4EHP promotes Ldh mRNA translation in and fruit fly adaptation to hypoxia.

Hypoxia induces profound modifications in the gene expression program of eukaryotic cells due to lowered ATP supply resulting from the blockade of oxidative phosphorylation. One significant consequence of oxygen deprivation is the massive repression of protein synthesis, leaving a limited set of mRNAs to be translated. Drosophila melanogaster is strongly resistant to oxygen fluctuations; however, the mechanisms allowing specific mRNA to be translated into hypoxia are still unknown. Here, we show that Ldh mRNA encoding lactate dehydrogenase is highly translated into hypoxia by a mechanism involving a CA-rich motif present in its 3' untranslated region. Furthermore, we identified the cap-binding protein eIF4EHP as a main factor involved in 3'UTR-dependent translation under hypoxia. In accordance with this observation, we show that eIF4EHP is necessary for Drosophila development under low oxygen concentrations and contributes to Drosophila mobility after hypoxic challenge. Altogether, our data bring new insight into mechanisms contributing to LDH production and Drosophila adaptation to oxygen variations.

BORIS/CTCFL-mediated chromatin accessibility alterations promote a pro-invasive transcriptional signature in melanoma cells.

Melanoma is the deadliest form of skin cancer, due to its tendency to metastasize early. Brother of regulator of imprinted sites (BORIS), also known as CCCTC binding factor-like (CTCFL), is a transcription regulator that becomes ectopically expressed in melanoma. We recently showed that BORIS contributes to melanoma phenotype switching by altering the gene expression program of melanoma cells from an intermediate melanocytic state toward a more mesenchymal-like state. However, the mechanism underlying this transcriptional switch remains unclear. Here, ATAC-seq was used to study BORIS-mediated chromatin accessibility alterations in melanoma cells harboring an intermediate melanocytic state. The gene set that gained promoter accessibility, following ectopic BORIS expression, showed enrichment for biological processes associated with melanoma invasion, while promoters of genes associated with proliferation showed reduced accessibility. Integration of ATAC-seq and RNA-seq data demonstrated that increased chromatin accessibility was associated with transcriptional upregulation of genes involved in tumor progression processes, and the aberrant activation of oncogenic transcription factors, while reduced chromatin accessibility and downregulated genes were associated with repressed activity of tumor suppressors and proliferation factors. Together, these findings indicate that BORIS mediates transcriptional reprogramming in melanoma cells by altering chromatin accessibility and gene expression, shifting the cellular transcription landscape of melanoma cells toward a mesenchymal-like genetic signature.

Super-enhancer-associated transcription factors collaboratively regulate trophoblast-active gene expression programs in human trophoblast stem cells.

The placenta is an essential organ that supports the growth and development of the fetus during pregnancy. However, cell type-specific enhancers and transcription factors (TFs), and the mechanisms underlying the maintenance and differentiation of trophoblast stem cell (TSC) populations in the human placenta remain elusive. Here, using human TSCs as a model system, we identify 31,362 enhancers that are enriched with the motifs of previously reported TSC-pivotal TFs, including TEAD4, GATA2/3 and TFAP2C. Subsequently, we identify 580 super-enhancers (SEs) and 549 SE-associated genes. These genes are robustly expressed in the human placenta and include numerous TFs, implying that SE-associated TFs (SE-TFs) may play crucial roles in placental development. Additionally, we identify the global binding sites of five TSC-pivotal SE-TFs (FOS, GATA2, MAFK, TEAD4 and TFAP2C), revealing that they preferentially co-occupy enhancers, regulate each otherand form a trophoblast-active gene regulatory network. Loss-of-function studies unveil that the five TFs promote self-renewal of TSCs by activating proliferation-associated genes while repressing developmental genes. We further reveal that the five TFs exert conserved and unique functions on placental development between humans and mice. Our study provides important insights into the roles of human TSC-pivotal TFs in regulating placenta-specific gene expression programs.

Stress-induced cell depolarization through the MAP kinase-Cdc42 axis.

General stress responses, which sense environmental or endogenous signals, aim at promoting cell survival and fitness during adverse conditions. In eukaryotes, mitogen-activated protein (MAP) kinase-driven cascades trigger a shift in the cell's gene expression program as a cellular adaptation to stress. Here, we review another aspect of activated MAP kinase cascades reported in fission yeast: the transient inhibition of cell polarity in response to oxidative stress. The phosphorylation by a stress-activated MAP kinase of regulators of the GTPase cell division cycle 42 (Cdc42) causes a transient inhibition of polarized cell growth. The formation of growth sites depends on limiting and essential polarity components. We summarize here some processes in which inhibition of Cdc42 may be a general mechanism to regulate polarized growth also under physiological conditions.

General transcription factor TAF4 antagonizes epigenetic silencing by Polycomb to maintain intestine stem cell functions

Taf4 (TATA-box binding protein-associated factor 4) is a subunit of the general transcription factor TFIID, a component of the RNA polymerase II pre-initiation complex that interacts with tissue-specific transcription factors to regulate gene expression. Properly regulated gene expression is particularly important in the intestinal epithelium that is constantly renewed from stem cells. Tissue-specific inactivation of Taf4 in murine intestinal epithelium during embryogenesis compromised gut morphogenesis and the emergence of adult-type stem cells. In adults, Taf4 loss impacted the stem cell compartment and associated Paneth cells in the stem cell niche, epithelial turnover and differentiation of mature cells, thus exacerbating the response to inflammatory challenge. Taf4 inactivation ex vivo in enteroids prevented budding formation and maintenance and caused broad chromatin remodeling and a strong reduction in the numbers of stem and progenitor cells with a concomitant increase in an undifferentiated cell population that displayed high activity of the Ezh2 and Suz12 components of Polycomb Repressive Complex 2 (PRC2). Treatment of Taf4-mutant enteroids with a specific Ezh2 inhibitor restored buddings, cell proliferation and the stem/progenitor compartment. Taf4 loss also led to increased PRC2 activity in cells of adult crypts associated with modification of the immune/inflammatory microenvironment that potentiated Apc-driven tumorigenesis. Our results reveal a novel function of Taf4 in antagonizing PRC2-mediated repression of the stem cell gene expression program to assure normal development, homeostasis, and immune-microenvironment of the intestinal epithelium.

The Xbp1-regulated transcription factor Mist1 restricts antibody secretion by restraining Blimp1 expression in plasma cells.

Antibody secretion by plasma cells provides acute and long-term protection against pathogens. The high secretion potential of plasma cells depends on the unfolded protein response, which is controlled by the transcription factor Xbp1. Here, we analyzed the Xbp1-dependent gene expression program of plasma cells and identified Bhlha15 (Mist1) as the most strongly activated Xbp1 target gene. As Mist1 plays an important role in other secretory cell types, we analyzed in detail the phenotype of Mist1-deficient plasma cells in Cd23-Cre Bhlha15 fl/fl mice under steady-state condition or upon NP-KLH immunization. Under both conditions, Mist1-deficient plasma cells were 1.4-fold reduced in number and exhibited increased IgM production and antibody secretion compared to control plasma cells. At the molecular level, Mist1 regulated a largely different set of target genes compared with Xbp1. Notably, expression of the Blimp1 protein, which is known to activate immunoglobulin gene expression and to contribute to antibody secretion, was 1.3-fold upregulated in Mist1-deficient plasma cells, which led to a moderate downregulation of most Blimp1-repressed target genes in the absence of Mist1. Importantly, a 2-fold reduction of Blimp1 (Prdm1) expression was sufficient to restore the cell number and antibody expression of plasma cells in Prdm1 Gfp/+ Cd23-Cre Bhlha15 fl/fl mice to the same level seen in control mice. Together, these data indicate that Mist1 restricts antibody secretion by restraining Blimp1 expression, which likely contributes to the viability of plasma cells.

Obesity alters the mouse endometrial transcriptome in a cell context-dependent manner

Obesity impacts fertility and is positively correlated with endometrial hyperplasia and endometrial cancer occurrence. Endometrial epithelia often harbor disease driver-mutations, while endometrial stroma are highly regulative of neighboring epithelia. Here, we sought to determine distinct transcriptome changes occurring in individual cell types in the obese mouse uterus. Outbred CD-1 mice were fed high-fat or control diets for 18 weeks, estrous cycle staged, and endometrial epithelia, macrophages, and stroma isolated for transcriptomic analysis. High-fat diet mice displayed increased body mass and developed glucose intolerance, hyperinsulinemia, and fatty liver. Obese mouse epithelia displayed differential gene expression for genes related to innate immunity and leukocyte chemotaxis. The obese mouse stroma differentially expressed factors related to circadian rhythm, and expression of these genes correlated with glucose tolerance or body mass. We observed correlations between F4/80 + macrophage numbers, Cleaved Caspase 3 (CC3) apoptosis marker staining and glucose intolerance among obese mice, including a subgroup of obese mice with high CC3 + luminal epithelia. This subgroup displayed differential gene expression among all cell types, with pathways related to immune escape in epithelia and macrophages, while the stroma dysregulated pathways related to regulation of epithelia. These results suggest an important role for differential response of both the epithelia and stroma in their response to obesity, while macrophages are dysregulated in the context of apoptotic epithelia. The obesity-related gene expression programs in cells within the uterine microenvironment may influence the ability of the endometrium to function during pregnancy and influence disease pathogenesis.

SMNDC1 links chromatin remodeling and splicing to regulate pancreatic hormone expression.

Insulin expression is primarily restricted to the pancreatic β cells, which are physically or functionally depleted in diabetes. Identifying targetable pathways repressing insulin in non-β cells, particularly in the developmentally related glucagon-secreting α cells, is an important aim of regenerative medicine. Here, we perform an RNA interference screen in a murine α cell line to identify silencers of insulin expression. We discover that knockdown of the splicing factor Smndc1 triggers a global repression of α cell gene-expression programs in favor of increased β cell markers. Mechanistically, Smndc1 knockdown upregulates the β cell transcription factor Pdx1 by modulating the activities of the BAF and Atrx chromatin remodeling complexes. SMNDC1's repressive role is conserved in human pancreatic islets, its loss triggering enhanced insulin secretion and PDX1 expression. Our study identifies Smndc1 as a key factor connecting splicing and chromatin remodeling to the control of insulin expression in human and mouse islet cells.

The SWI/SNF ATPase BRG1 facilitates multiple pro-tumorigenic gene expression programs in SMARCB1-deficient cancer cells

Malignant rhabdoid tumor (MRT) is driven by the loss of the SNF5 subunit of the SWI/SNF chromatin remodeling complex and then thought to be maintained by residual SWI/SNF (rSWI/SNF) complexes that remain present in the absence of SNF5. rSWI/SNF subunits colocalize extensively on chromatin with the transcription factor MYC, an oncogene identified as a novel driver of MRT. Currently, the role of rSWI/SNF in modulating MYC activity has neither been delineated nor has a direct link between rSWI/SNF and other oncogenes been uncovered. Here, we expose the connection between rSWI/SNF and oncogenic processes using a well-characterized chemical degrader to deplete the SWI/SNF ATPase, BRG1. Using a combination of gene expression and chromatin accessibility assays we show that rSWI/SNF complexes facilitate MYC target gene expression. We also find that rSWI/SNF maintains open chromatin at sites associated with hallmark cancer genes linked to the AP-1 transcription factor, suggesting that AP-1 may drive oncogenesis in MRT. Interestingly, changes in MYC target gene expression are not overtly connected to the chromatin remodeling function of rSWI/SNF, revealing multiple mechanisms used by rSWI/SNF to control transcription. This work provides an understanding of how residual SWI/SNF complexes may converge on multiple oncogenic processes when normal SWI/SNF function is impaired.

MeCP2 and transcriptional control of eukaryotic gene expression

Eukaryotic gene expression is controlled at multiple steps that work in harmony to ensure proper maintenance of cellular morphology and function. Such regulatory mechanisms would include transcriptional gene regulation, which is in turn controlled by chromatin remodeling, distinct topologically associating domains of the chromatin structure, cis-regulatory elements such as enhancers and promoters, action of trans-acting factors, DNA methylation, RNA modifications, and post-translational modification of histones. These guiding mechanisms of gene expression play critical roles in the epigenetic setting of individual cells within the eukaryotic systems. Some epigenetic factors may play multiple functional roles in guarding the accurate gene expression program of the eukaryotic cells, especially within the central nervous system. A well-studied example of such multi-functional factors is the methyl-CpG-binding protein 2 (MeCP2), a nuclear protein that is encoded by the X-linked MECP2 gene. Here, we aim to provide an overview of eukaryotic gene regulation, the three-dimensional chromatin organization, standard techniques to study newly synthesized RNA transcripts, and the role of MeCP2 as an important transcriptional regulator in eukaryotes.

EWSR1-ATF1 dependent 3D connectivity regulates oncogenic and differentiation programs in Clear Cell Sarcoma

Oncogenic fusion proteins generated by chromosomal translocations play major roles in cancer. Among them, fusions between EWSR1 and transcription factors generate oncogenes with powerful chromatin regulatory activities, capable of establishing complex gene expression programs in permissive precursor cells. Here we define the epigenetic and 3D connectivity landscape of Clear Cell Sarcoma, an aggressive cancer driven by the EWSR1-ATF1 fusion gene. We find that EWSR1-ATF1 displays a distinct DNA binding pattern that requires the EWSR1 domain and promotes ATF1 retargeting to new distal sites, leading to chromatin activation and the establishment of a 3D network that controls oncogenic and differentiation signatures observed in primary CCS tumors. Conversely, EWSR1-ATF1 depletion results in a marked reconfiguration of 3D connectivity, including the emergence of regulatory circuits that promote neural crest-related developmental programs. Taken together, our study elucidates the epigenetic mechanisms utilized by EWSR1-ATF1 to establish regulatory networks in CCS, and points to precursor cells in the neural crest lineage as candidate cells of origin for these tumors.