Chromatin Gene Research Articles

Integrated ATAC-seq and RNA-seq Analysis of In Vitro Cultured Skeletal Muscle Satellite Cells to Understand Changes in Cell Proliferation.

Skeletal muscle satellite cells, the resident stem cells in pig skeletal muscle, undergo proliferation and differentiation to enable muscle tissue repair. The proliferative and differentiative abilities of these cells gradually decrease during in vitro cultivation as the cell passage number increases. Despite extensive research, the precise molecular mechanisms that regulate this process are not fully understood. To bridge this knowledge gap, we conducted transcriptomic analysis of skeletal muscle satellite cells during in vitro cultivation to quantify passage number-dependent changes in the expression of genes associated with proliferation. Additionally, we explored the relationships between gene transcriptional activity and chromatin accessibility using transposase-accessible chromatin sequencing. This revealed the closure of numerous open chromatin regions, which were primarily located in intergenic regions, as the cell passage number increased. Integrated analysis of the transcriptomic and epigenomic data demonstrated a weak correlation between gene transcriptional activity and chromatin openness in expressed genic regions; although some genes (e.g., GNB4 and FGD5) showed consistent relationships between gene expression and chromatin openness, a substantial number of differentially expressed genes had no clear association with chromatin openness in expressed genic regions. The p53-p21-RB signaling pathway may play a critical regulatory role in cell proliferation processes. The combined transcriptomic and epigenomic approach taken here provided key insights into changes in gene expression and chromatin openness during in vitro cultivation of skeletal muscle satellite cells. These findings enhance our understanding of the intricate mechanisms underlying the decline in cellular proliferation capacity in cultured cells.

Chromatin modifiers in endometriosis pathogenesis

Introduction. It was revealed that various epigenetic abnormalities may play an important role in the endometriosis pathogenesis. The regulation of chromatin structure is carried out mainly by chromatin modifiers (CMs), which stimulate generation of genomic regions with different functional structures and thus change the patterns or levels of gene expression by exerting expected biological functions and causing epigenetic changes.Aim: to consider СМs role in endometriosis pathogenesis and their regulation mechanism assessing current publications.Materials and Methods. The search was conducted in the databases PubMed, Scopus, Web of Science, Google Scholar and eLibrary. Keywords and phrases in Russian and English related to the research topic were used as follows: "endometriosis", "chromatin modifiers", "histone acetylation", "DNA methylation", "microRNA", "endometriosis", "chromatin modifiers", "histone acetylation", "DNA methylation", "microRNA". The evaluation of articles was carried out in accordance with PRISMA recommendations.Results. Chromatin modifiers control differentiation, growth and development, aging and cell death by interacting with various functional chromatin elements. They can cause abnormal gene expression by regulating chromatin structure affecting emergence and development of endometriosis. DNA methylation determines cell types, controls gene expression and genome stability. Abnormal DNA methylation in gene promoter regions necessary for normal endometrial response affects endometriosis development. DNA methyltransferase (DNMT) inhibitors reduce the methylation of human homeobox A10 (HOXA10) and progesterone receptor (PR) genes and potentiate their expression in endometrial cells, improving endometrial susceptibility and inhibiting cell cycle progression. Abnormal histone modifications in endometrial cells may facilitate or hinder the access of transcription mechanisms to chromatin DNA. Histone deacetylase inhibitors effectively eliminate the effects of abnormal histone modifications in endometriosis cells and prevent endometriosis progression. The expression of non-coding RNAs and chromatin remodeling complexes also alters chromatin structure being involved in arising endometriosis and is associated with infertility by promoting proliferation, invasion and migration of endometriod cells.Conclusion. Chromatin modifiers play a key role in developing endometriosis by controlling gene expression and chromatin structure. Understanding underlying mechanisms provides valuable information for diagnostics and development of new approaches to treat endometriosis.

Abstract PR012: KAT6A/B and Menin-MLL complexes coordinately regulate estrogen receptor-driven gene expression programs in breast cancer

Abstract The estrogen receptor (ER) is expressed in ∼70% of breast cancers where it functions as a critical transcription factor that regulates cell growth and tumor progression. ER inhibition is a mainstay treatment for ER-positive (ER+) breast cancer, but despite the clinical benefit of targeting ER signaling, breast cancer remains the second leading cause of cancer-related death in women, with ∼50% of deaths resulting from ER+ tumors. While recurrent and metastatic ER+ tumors are typically resistant to endocrine therapies they continue to rely on ER signaling suggesting that disruption of other chromatin associated complexes that mediate ER-driven gene expression may present an alternative therapeutic strategy. One chromatin modifier important in regulating ER function is the histone acetyltransferase KAT6A. KAT6A is amplified and/or overexpressed in ∼15% of ER+ tumors and a novel KAT6A/B inhibitor has entered clinical trials for advanced breast cancer. However, as with most single agent therapies, compensatory mechanisms may attenuate the effects of KAT6A/B inhibition on ER-driven gene expression. We hypothesized that the dual targeting of distinct chromatin complexes would enhance the effects of KAT6A/B inhibition, by altering chromatin state and transcriptional output. To that end we performed an epigenetic focused CRISPR-Cas9-based functional genetic screen to identify chromatin-associated proteins that enhance KAT6A/B inhibition. CRISPR guides targeting MEN1 (encoding Menin) were among our top hits as sensitizers to treatment with the KAT6A/B inhibitor PF-9363. We confirmed that co-treatment of PF-9363, and the Menin inhibitor, SNDX-5613, had strong synergistic anti-proliferative effects in a panel of ER+ breast cancer cell lines. Mechanistically, our data revealed that KAT6A/B and Menin cooperatively regulate ER-driven gene expression and chromatin accessibility, both through direct effects on ESR1 expression as well as through effects at ER target genes. Specifically, we found that KAT6A and Menin-KMT2A complexes co-localized at promoters of ER target genes and treatment with KAT6A/B or Menin inhibitors displaced KAT6A and Menin/KMT2A from ER target genes, respectively. Yet only the combination effectively displaced both complexes from chromatin, which produced more dramatic changes in ER-directed gene expression than either small molecule alone. Combined displacement of KAT6A and Menin/KMT2A at ER-regulated genes induced loss of an active chromatin landscape characterized by reduction of activating histone modifications and loss of RNA Polymerase II binding to ER target loci. Importantly, the combination of KAT6A/B and Menin inhibition was effective in a panel of ER+ patient-derived organoid models as well as in various models of endocrine resistance, including models with ESR1 mutations, NF1 loss, and FOXA1 mutations. Both KAT6A/B and Menin inhibitors are in Phase I/II clinical trials and have shown manageable toxicity profiles to date. This combination represents a novel potential therapeutic combination for ER+ breast cancer. Citation Format: Sarah Naomi Olsen, Bryn Anderson, Charles Hatton, Rinath Jeselsohn, Myles Brown, Eneda Toska, Scott Armstrong. KAT6A/B and Menin-MLL complexes coordinately regulate estrogen receptor-driven gene expression programs in breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Expanding and Translating Cancer Synthetic Vulnerabilities; 2024 Jun 10-13; Montreal, Quebec, Canada. Philadelphia (PA): AACR; Mol Cancer Ther 2024;23(6 Suppl):Abstract nr PR012.

Cohesin — bridging the gap among gene transcription, genome stability, and human diseases

The intricate landscape of cellular processes governing gene transcription, chromatin organization, and genome stability is a fascinating field of study. A key player in maintaining this delicate equilibrium is the cohesin complex, a molecular machine with multifaceted roles. This review presents an in‐depth exploration of these intricate connections and their significant impact on various human diseases.

Cell type specific roles of FOXP1 during early neocortical murine development.

Cortical development is a tightly controlled process and any deviation during development may increase the susceptibility to neurodevelopmental disorders, such as autism spectrum disorders (ASD). Numerous studies identified mutations in FOXP1, a transcription factor enriched in the neocortex, as causal for ASD and FOXP1 syndrome. Our group has shown that Foxp1 deletion in the mouse cortex leads to overall reduced cortex thickness, alterations in cortical lamination, and changes in the relative thickness of cortical layers. However, the developmental and cell type-specific mechanisms underlying these changes remained unclear. This work characterizes the developmental requirement of neocortical Foxp1 at key embryonic and perinatal ages using a conditional knock-out of Foxp1. We find that Foxp1 deletion results in accelerated pseudo-age during early neurogenesis, increased cell cycle exit during late neurogenesis, altered gene expression and chromatin accessibility, and selective migration deficits in a subset of upper-layer neurons. These data explain the postnatal differences observed in cortical layers and relative cortical thickness. We also highlight genes regulated by FOXP1 and their enrichment with high-confidence ASD or synaptic genes. Together, these results underscore a network of neurodevelopmental disorder-related genes that may serve as potential modulatory targets for postnatal modification relevant to ASD and FOXP1 syndrome.

Identification and characterization of a human MORC2 DNA binding region that is required for gene silencing.

The eukaryotic microrchidia (MORC) protein family are DNA gyrase, Hsp90, histidine kinase, MutL (GHKL)-type ATPases involved in gene expression regulation and chromatin compaction. The molecular mechanisms underlying these activities are incompletely understood. Here we studied the full-length human MORC2 protein biochemically. We identified a DNA binding site in the C-terminus of the protein, and we observe that this region is heavily phosphorylated in cells. Phosphorylation of MORC2 reduces its affinity for DNA and appears to exclude the protein from the nucleus. We observe that DNA binding by MORC2 reduces its ATPase activity and that MORC2 can topologically entrap multiple DNA substrates between its N-terminal GHKL and C-terminal coiled coil 3 dimerization domains. Finally, we observe that the MORC2 C-terminal DNA binding region is required for gene silencing in cells. Together, our data provide a model to understand how MORC2 engages with DNA substrates to mediate gene silencing.

Muscle-specific pyruvate kinase isoforms, PKM1 and PKM2, regulate mammalian SWI/SNF proteins and histone 3 phosphorylation during myoblast differentiation.

Pyruvate kinase is a glycolytic enzyme that converts phosphoenolpyruvate and ADP into pyruvate and ATP. There are two genes that encode pyruvate kinase in vertebrates; Pkm and Pkl encode muscle- and liver/erythrocyte-specific forms, respectively. Each gene encodes two isoenzymes due to alternative splicing. Both muscle-specific enzymes, PKM1 and PKM2, function in glycolysis, but PKM2 also has been implicated in gene regulation due to its ability to phosphorylate histone 3 threonine 11 (H3T11) in cancer cells. Here, we examined the roles of PKM1 and PKM2 during myoblast differentiation. RNA-seq analysis revealed that PKM2 promotes the expression of Dpf2/Baf45d and Baf250a/Arid1A. DPF2 and BAF250a are subunits that identify a specific sub-family of the mammalian SWI/SNF (mSWI/SNF) of chromatin remodeling enzymes that is required for the activation of myogenic gene expression during differentiation. PKM2 also mediated the incorporation of DPF2 and BAF250a into the regulatory sequences controlling myogenic gene expression. PKM1 did not affect expression but was required for nuclear localization of DPF2. Additionally, PKM2 was required not only for the incorporation of phosphorylated H3T11 in myogenic promoters but also for the incorporation of phosphorylated H3T6 and H3T45 at myogenic promoters via regulation of AKT and protein kinase C isoforms that phosphorylate those amino acids. Our results identify multiple unique roles for PKM2 and a novel function for PKM1 in gene expression and chromatin regulation during myoblast differentiation.

De novo prediction of functional effects of genetic variants from DNA sequences based on context-specific molecular information

Deciphering the functional effects of noncoding genetic variants stands as a fundamental challenge in human genetics. Traditional approaches, such as Genome-Wide Association Studies (GWAS), Transcriptome-Wide Association Studies (TWAS), and Quantitative Trait Loci (QTL) studies, are constrained by obscured the underlying molecular-level mechanisms, making it challenging to unravel the genetic basis of complex traits. The advent of Next-Generation Sequencing (NGS) technologies has enabled context-specific genome-wide measurements, encompassing gene expression, chromatin accessibility, epigenetic marks, and transcription factor binding sites, to be obtained across diverse cell types and tissues, paving the way for decoding genetic variation effects directly from DNA sequences only. The de novo predictions of functional effects are pivotal for enhancing our comprehension of transcriptional regulation and its disruptions caused by the plethora of noncoding genetic variants linked to human diseases and traits. This review provides a systematic overview of the state-of-the-art models and algorithms for genetic variant effect predictions, including traditional sequence-based models, Deep Learning models, and the cutting-edge Foundation Models. It delves into the ongoing challenges and prospective directions, presenting an in-depth perspective on contemporary developments in this domain.

Evolution of diapause in the African turquoise killifish by remodeling the ancient gene regulatory landscape

Suspended animation states allow organisms to survive extreme environments. The African turquoise killifish has evolved diapause as a form of suspended development to survive a complete drought. However, the mechanisms underlying the evolution of extreme survival states are unknown. To understand diapause evolution, we performed integrative multi-omics (gene expression, chromatin accessibility, and lipidomics) in the embryos of multiple killifish species. We find that diapause evolved by a recent remodeling of regulatory elements at very ancient gene duplicates (paralogs) present in all vertebrates. CRISPR-Cas9-based perturbations identify the transcription factors REST/NRSF and FOXOs as critical for the diapause gene expression program, including genes involved in lipid metabolism. Indeed, diapause shows a distinct lipid profile, with an increase in triglycerides with very-long-chain fatty acids. Our work suggests a mechanism for the evolution of complex adaptations and offers strategies to promote long-term survival by activating suspended animation programs in other species.

Cell-type-specific effects of autism-associated chromosome 15q11.2-13.1 duplications in human brain.

Recurrent copy number variation represents one of the most well-established genetic drivers in neurodevelopmental disorders, including autism spectrum disorder (ASD). Duplication of 15q11.2-13.1 (dup15q) is a well-described neurodevelopmental syndrome that increases the risk of ASD by over 40-fold. However, the effects of this duplication on gene expression and chromatin accessibility in specific cell types in the human brain remain unknown. To identify the cell-type-specific transcriptional and epigenetic effects of dup15q in the human frontal cortex we conducted single-nucleus RNA-sequencing and multi-omic sequencing on dup15q cases (n=6) as well as non-dup15q ASD (n=7) and neurotypical controls (n=7). Cell-type-specific differential expression analysis identified significantly regulated genes, critical biological pathways, and differentially accessible genomic regions. Although there was overall increased gene expression across the duplicated genomic region, cellular identity represented an important factor mediating gene expression changes. Neuronal subtypes, showed greater upregulation of gene expression across a critical region within the duplication as compared to other cell types. Genes within the duplicated region that had high baseline expression in control individuals showed only modest changes in dup15q, regardless of cell type. Of note, dup15q and ASD had largely distinct signatures of chromatin accessibility, but shared the majority of transcriptional regulatory motifs, suggesting convergent biological pathways. However, the transcriptional binding factor motifs implicated in each condition implicated distinct biological mechanisms; neuronal JUN/FOS networks in ASD vs. an inflammatory transcriptional network in dup15q microglia. This work provides a cell-type-specific analysis of how dup15q changes gene expression and chromatin accessibility in the human brain and finds evidence of marked cell-type-specific effects of this genetic driver. These findings have implications for guiding therapeutic development in dup15q syndrome, as well as understanding the functional effects CNVs more broadly in neurodevelopmental disorders.

Multiscale 3D genome organization underlies duck fatty liver with no adipose inflammation or serious injury

Non-alcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease. Little is known about how gene expression and chromatin structure are regulated in NAFLD due to lack of suitable model. Ducks naturally develop fatty liver similar to serious human non-alcoholic fatty liver (NAFL) without adipose inflammation and liver fibrosis, thus serves as a good model for investigating molecular mechanisms of adipose metabolism and anti-inflammation. Here, we constructed a NAFLD model without adipose inflammation and liver fibrosis in ducks. By performing dynamic pathological and transcriptomic analyses, we identified critical genes involving in regulation of the NF-κB and MHCII signaling, which usually lead to adipose inflammation and liver fibrosis. We further generated dynamic three-dimensional chromatin maps during liver fatty formation and recovery. This showed that ducks enlarged hepatocyte cell nuclei to reduce inter-chromosomal interaction, decompress chromatin structure, and alter strength of intra-TAD and loop interactions during fatty liver formation. These changes partially contributed to the tight control the NF-κB and the MHCII signaling. Our analysis uncovers duck chromatin reorganization might be advantageous to maintain liver regenerative capacity and reduce adipose inflammation. These findings shed light on new strategies for NAFLD control.

P22 A single-cell map of the epigenomic landscape in keratinocytes exposed to the cytokine milieu of atopic dermatitis

Abstract Introduction and aims In atopic dermatitis (AD), induction of inflammatory responses regulated by cytokines such as of thymic stromal lymphopoietin exacerbate the disrupted integrity of the skin barrier. Cytokines, such as interleukin (IL)-4, IL-13 and IL-22, present in the skin milieu, bind to their corresponding receptors on the cell surface of keratinocytes, activating multiple signal transduction pathways that lead to changes in the gene expression profile of keratinocytes, in part, by modulating chromatin structure. Understanding the intricate interplay between cytokine-mediated signalling pathways in keratinocytes and the resulting epigenetic changes is essential for elucidating the pathogenesis of AD. In this study, we utilized high-throughput technology to examine the effects of cytokines on modulation of the epigenetic landscape at the keratinocyte single-cell level, and subsequent effects on the gene expression. Methods We utilized the reconstructed human epidermis model, an in vitro three-dimensional human epidermis model consisting of a polycarbonate membrane and keratinocytes mimicking the structure of native human epidermis. Following exposure to AD cytokine(s), the model gene expression and chromatin accessibility of target cells were analysed using single-cell multiomics approaches. Results Our preliminary data show that IL-4, IL-13 and IL-22 reduce the expression levels of the barrier-related gene filaggrin in keratinocytes. Conclusions Unlike genetic changes, epigenetic alterations are dynamic and can be restored to their normal state, which has major implications for the development of therapeutics for various diseases. This study will provide valuable insights into the epigenetic mechanisms underlying AD and present potential targets for epigenetic-related therapeutic interventions. These interventions can be used in combination with available compounds to reduce the recurrence of AD and improve the treatment efficiency.

An ApiAp2 Transcription Factor with a Dispensable Role in Plasmodium berghei Life Cycle.

Malaria parasites have a complex life cycle and undergo replication and population expansion within vertebrate hosts and mosquito vectors. These developmental transitions rely on changes in gene expression and chromatin reorganization that result in the activation and silencing of stage-specific genes. The ApiAp2 family of DNA-binding proteins plays an important role in regulating gene expression in malaria parasites. Here, we characterized the ApiAp2 protein in Plasmodium berghei, which we termed Ap2-D. In silico analysis revealed that Ap2-D has three beta-sheets followed by a helix at the C-terminus for DNA binding. Using gene tagging with 3XHA-mCherry, we found that Ap2-D is expressed in Plasmodium blood stages and is present in the parasite cytoplasm and nucleus. Surprisingly, our gene deletion study revealed a completely dispensable role for Ap2-D in the entirety of the P. berghei life cycle. Ap2-D KO parasites were found to grow in the blood successfully and progress through the mosquito midgut and salivary glands. Sporozoites isolated from mosquito salivary glands were infective for hepatocytes and achieved similar patency as WT in mice. We emphasize the importance of genetic validation of antimalarial drug targets before progressing them to drug discovery.

A Novel Macrophage Subpopulation Conveys Increased Genetic Risk of Coronary Artery Disease.

Coronary artery disease (CAD), the leading cause of death worldwide, is influenced by both environmental and genetic factors. Although over 250 genetic risk loci have been identified through genome-wide association studies, the specific causal variants and their regulatory mechanisms are still largely unknown, particularly in disease-relevant cell types such as macrophages. We utilized single-cell RNA-seq and single-cell multiomics approaches in primary human monocyte-derived macrophages to explore the transcriptional regulatory network involved in a critical pathogenic event of coronary atherosclerosis-the formation of lipid-laden foam cells. The relative genetic contribution to CAD was assessed by partitioning disease heritability across different macrophage subpopulations. Meta-analysis of single-cell RNA-seq data sets from 38 human atherosclerotic samples was conducted to provide high-resolution cross-referencing to macrophage subpopulations in vivo. We identified 18 782 cis-regulatory elements by jointly profiling the gene expression and chromatin accessibility of >5000 macrophages. Integration with CAD genome-wide association study data prioritized 121 CAD-related genetic variants and 56 candidate causal genes. We showed that CAD heritability was not uniformly distributed and was particularly enriched in the gene programs of a novel CD52-hi lipid-handling macrophage subpopulation. These CD52-hi macrophages displayed significantly less lipoprotein accumulation and were also found in human atherosclerotic plaques. We investigated the cis-regulatory effect of a risk variant rs10488763 on FDX1, implicating the recruitment of AP-1 and C/EBP-β in the causal mechanisms at this locus. Our results provide genetic evidence of the divergent roles of macrophage subsets in atherogenesis and highlight lipid-handling macrophages as a key subpopulation through which genetic variants operate to influence disease. These findings provide an unbiased framework for functional fine-mapping of genome-wide association study results using single-cell multiomics and offer new insights into the genotype-environment interactions underlying atherosclerotic disease.

Cancer–stromal cell interactions in breast cancer brain metastases induce glycocalyx-mediated resistance to HER2-targeting therapies

Brain metastatic breast cancer is particularly lethal largely due to therapeutic resistance. Almost half of the patients with metastatic HER2-positive breast cancer develop brain metastases, representing a major clinical challenge. We previously described that cancer-associated fibroblasts are an important source of resistance in primary tumors. Here, we report that breast cancer brain metastasis stromal cell interactions in 3D cocultures induce therapeutic resistance to HER2-targeting agents, particularly to the small molecule inhibitor of HER2/EGFR neratinib. We investigated the underlying mechanisms using a synthetic Notch reporter system enabling the sorting of cancer cells that directly interact with stromal cells. We identified mucins and bulky glycoprotein synthesis as top-up-regulated genes and pathways by comparing the gene expression and chromatin profiles of stroma-contact and no-contact cancer cells before and after neratinib treatment. Glycoprotein gene signatures were also enriched in human brain metastases compared to primary tumors. We confirmed increased glycocalyx surrounding cocultures by immunofluorescence and showed that mucinase treatment increased sensitivity to neratinib by enabling a more efficient inhibition of EGFR/HER2 signaling in cancer cells. Overexpression of truncated MUC1 lacking the intracellular domain as a model of increased glycocalyx-induced resistance to neratinib both in cell culture and in experimental brain metastases in immunodeficient mice. Our results highlight the importance of glycoproteins as a resistance mechanism to HER2-targeting therapies in breast cancer brain metastases.

LSD2 Is an Epigenetic Player in Multiple Types of Cancer and Beyond.

Histone demethylases, enzymes responsible for removing methyl groups from histone proteins, have emerged as critical players in regulating gene expression and chromatin dynamics, thereby influencing various cellular processes. LSD2 and LSD1 have attracted considerable interest among these demethylases because of their associations with cancer. However, while LSD1 has received significant attention, LSD2 has not been recognized to the same extent. In this study, we conduct a comprehensive comparison between LSD2 and LSD1, with a focus on exploring LSD2's implications. While both share structural similarities, LSD2 possesses unique features as well. Functionally, LSD2 shows diverse roles, particularly in cancer, with tissue-dependent roles. Additionally, LSD2 extends beyond histone demethylation, impacting DNA methylation, cancer cell reprogramming, E3 ubiquitin ligase activity and DNA damage repair pathways. This study underscores the distinct roles of LSD2, providing insights into their contributions to cancer and other cellular processes.

Epigenetic reprogramming of CD8+ T cell populations drives exhaustion in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS)

Abstract Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a severe, debilitating disease with striking similarities to long-COVID/PASC, but its etiology remains unclear. Mounting evidence implicates immune dysfunction as a potential cause for ME/CFS. We previously reported metabolic changes in ME/CFS CD8+ T cells, including increased fatty acid oxidation, decreased glycolysis, and reduced mitochondrial membrane potential - features commonly found in exhausted T cells. To comprehensively characterize the gene regulatory state underlying potential transcriptional dysregulation, we first re-analyzed a single cell transcriptomic atlas of ME/CFS by reclassifying CD8+ T cells, where we found that various clusters upregulated exhaustion-associated genes. We then generated high resolution profiles of gene expression and chromatin accessibility in effector memory (TEM) and naïve CD8+ T cells by RNA-seq and ATAC-seq at baseline and further analyzed different CD8+ subsets following symptom provocation. We observed in ME/CFS TEM upregulation of key transcription factors associated with exhaustion and downregulation of genes controlling key metabolic activities, as well as an altered chromatin landscape consistent with exhaustion programs, while ME/CFS TN displayed a potential loss of naïve identity. Our dataset serves as an important resource for understanding ME/CFS pathology and supports a novel state of immune dysregulation in ME/CFS which may offer new treatment options.

Consequences of Transient YAP Signaling in the Liver

The transcriptional co-activator YAP plays a complex role in liver homeostasis and disease development. In response to liver injuries and hepatotoxins, YAP is transiently activated to promote liver regeneration by coordinating cell proliferation and metabolism. By contrast, chronic activation of YAP in experimental models has been associated with liver diseases and cancer. These primary responses to YAP are well established; however, it is presently unknown how acute YAP signaling contributes to the eventual development of liver diseases. Here, we developed an in vivo model of dynamic YAP signaling using a doxycycline-inducible system to control the transient expression of a constitutively active YAP mutant in hepatocytes. We found that while YAP-dependent effects on liver growth and cell proliferation are reversible, there are metabolic changes in the liver that remain stable even after YAP inactivation. By coupling RNAseq and ATACseq analyses, we unraveled persistent changes in gene expression and chromatin accessibility, particularly those involved in lipid metabolism. Furthermore, we demonstrated that transient YAP signaling sensitizes the liver to develop more severe liver injuries than normally induced in an unprimed liver. These data suggest that transient YAP signaling imparts a form of lipid metabolic memory that promotes susceptibility to liver diseases. Overall, our current work reveals long-term consequences of acute YAP signaling that may influence the development of liver diseases. 5R01HL128850-07 (FC) and 5T32DK007477-40 (KA). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Dissecting gene activation and chromatin remodeling dynamics in single human cells undergoing reprogramming

During cell fate transitions, cells remodel their transcriptome, chromatin, and epigenome; however, it has been difficult to determine the temporal dynamics and cause-effect relationship between these changes at the single-cell level. Here, we employ the heterokaryon-mediated reprogramming system as a single-cell model to dissect key temporal events during early stages of pluripotency conversion using super-resolution imaging. We reveal that, following heterokaryon formation, the somatic nucleus undergoes global chromatin decompaction and removal of repressive histone modifications H3K9me3 and H3K27me3 without acquisition of active modifications H3K4me3 and H3K9ac. The pluripotency gene OCT4 (POU5F1) shows nascent and mature RNA transcription within the first 24h after cell fusion without requiring an initial open chromatin configuration at its locus. NANOG, conversely, has significant nascent RNA transcription only at 48h after cell fusion but, strikingly, exhibits genomic reopening early on. These findings suggest that the temporal relationship between chromatin compaction and gene activation during cellular reprogramming is gene context dependent.

Variant-to-function analysis of the childhood obesity chr12q13 locus implicates rs7132908 as a causal variant within the 3′ UTR of FAIM2

The ch12q13 locus is among the most significant childhood obesity loci identified in genome-wide association studies. This locus resides in a non-coding region within FAIM2; thus, the underlying causal variant(s) presumably influence disease susceptibility via cis-regulation. We implicated rs7132908 as a putative causal variant by leveraging our in-house 3D genomic data and public domain datasets. Using a luciferase reporter assay, we observed allele-specific cis-regulatory activity of the immediate region harboring rs7132908. We generated isogenic human embryonic stem cell lines homozygous for either rs7132908 allele to assess changes in gene expression and chromatin accessibility throughout a differentiation to hypothalamic neurons, a key cell type known to regulate feeding behavior. The rs7132908 obesity risk allele influenced expression of FAIM2 and other genes and decreased the proportion of neurons produced by differentiation. We have functionally validated rs7132908 as a causal obesity variant that temporally regulates nearby effector genes and influences neurodevelopment and survival.