Biology subjects

Bartlett, A.

Publications and source records attributed to Bartlett, A..

At least 19 recordsLinked to original sources

bioRxiv2026

A Multimodal Single-Cell Epigenomic and 3D Genome Atlas of the Human Basal Ganglia

The basal ganglia (BG) underlie motor control, reward processing, and many neurological and psychiatric disorders, but a comprehensive epigenomic and 3D-genome atlas of the human BG is lacking. Here we present a multimodal single-cell atlas profiling DNA methylation and 3D chromatin conformation in 261,331 nuclei (snm3C-seq) across eight subregions, resolving 12 classes, 31 subclasses, and 59 groups. Harmonized under the HMBA basal-ganglia consensus taxonomy, this atlas integrates with matched RNA, ATAC-seq, and histone-modification data across five regulatory layers. We identify millions of cell-type- and region-specific differentially methylated regions enriched for distinct transcription factor motifs and link them to disease-associated heritability. Neuron-specific loops dominate cell-type-specific 3D contact remodeling, while most non-neuron-specific loops are constitutive. Among spiny projection neurons (SPNs), chromatin loops, rather than TAD boundaries, distinguish D1, D2, and eccentric SPN subclasses, with eccentric SPNs showing the most loop-level reorganization among the three. We characterize STR D2 SMYD2-HTR7 SPN, a newly recognized POU6F2 D2-SPN subtype, and reveal region-specific methylation and contact gradients of disease-associated genes, including CADM1 and PDE8B. Integrative gene-regulatory networks reconstruct cell-type-resolved enhancer-promoter links to interpret Parkinsons disease risk variants at SNCA. Finally, MERFISH spatial profiling combined with cross-species Patch-seq identifies non-SPN neuronal subtypes, including a MOXD1 striosomal STR FS PTHLH-PVALB GABA subtype with distinct electrophysiology, partitioning across the striatal matrix-striosome boundary. HighlightsO_LIA multimodal single-cell atlas maps DNA methylation and 3D genome architecture across human basal ganglia cell types and subregions. C_LIO_LINeuron-specific loops dominate cell-type-specific 3D contact remodeling in the human BG, whereas most non-neuron-specific loops are constitutive. C_LIO_LISpiny Projection Neuron (SPN) subtypes exhibit regionally organized epigenomic and 3D genome signatures that align with dorsal-ventral identities. C_LIO_LIChromatin loops are the primary distinguishing feature among D1, D2, and eccentric SPN subclasses, with eccentric SPNs being the most 3D-reorganized. C_LIO_LIIntegrated regulatory maps link cell-type-specific enhancers to disease-associated genetic risk in the human basal ganglia. C_LIO_LINon-SPN interneurons differ in distribution and in transcriptional and epigenetic identity across the matrix-striosome boundary. C_LI

neuroscience↗

bioRxiv2026

An Integrated Single-Cell and Epigenomic Resource for Comparative Analysis of the Basal Ganglia

The basal ganglia regulate motor, cognitive, and affective behaviors, and their dysfunction underlies diverse neurological and psychiatric disorders. Comprehensive, accessible multi-omics resources are needed to understand the regulatory mechanisms governing basal ganglia cell types. Here we present an open, interactive web-based platform for exploring single-cell multi-omics datasets from basal ganglia, generated using 10X Multiome, snm3C-seq, and Paired-Tag technologies from the BICAN (NIH BRAIN Initiative Cell Atlas Network) consortium. The platform is available at https://basalganglia.epigenomes.net/ and enables integrated visualization of gene expression, chromatin accessibility, DNA methylation, histone modifications, and chromatin conformation across cell types and human, macaque, marmoset, and mouse species, with direct genome browser support and comparative epigenomic functionality. Representative analyses demonstrate cell-type-specific regulatory landscapes, conserved and species-specific regulatory elements, and links between epigenomic regulation and transcription. This resource provides a scalable, community-oriented foundation for advancing basal ganglia biology and interpreting regulatory mechanisms relevant to brain function and disease. HighlightsO_LIIntegrated single-cell epigenomic resource for basal ganglia C_LIO_LIInteractive genome browser enables multi-omics and cross-species exploration C_LIO_LIReveals cell-type-specific and species-specific regulatory landscapes C_LIO_LISupports community access to complex brain epigenomic datasets C_LI

genomics↗

bioRxiv2026

Aging-related Transcriptomic Changes with Spatial Resolution in the Human Prefrontal Cortex

The human prefrontal cortex (PFC), whose laminar organization is essential for cognitive function, is among the first regions to show age-related functional decline1,2. Single-cell sequencing studies revealed cell type-dependent aging effects but lacked spatial specificity3-6. Spatial transcriptomics (ST) advanced our molecular understanding of the human PFC7, yet whether aging-driven changes differ across PFC layers remains unclear. Here, we performed whole-transcriptome ST on postmortem PFC from 37 individuals across the adult lifespan. We mapped cortical layers and revealed aging mechanisms across layers. This represents one of the largest and most comprehensive lifespan ST analysis of the human PFC brain, offering crucial insight into how the brain ages and identifying potential molecular targets to mitigate cognitive aging and extend healthspan.

neuroscience↗

bioRxiv2025

Dissecting Epstein-Barr Virus Dependence Across Diverse Infected Cell Models

Epstein-Barr virus (EBV) is maintained clonally in a wide array of tumors and cells of lymphoproliferative disorders, yet the degree to which these infected cells continuously depend on EBV remains unresolved. To directly assess EBV dependence, we used two complementary Cas9-based viral eviction strategies: 1) targeting EBNA1 to promote episome dilution during cell division, and 2) targeting repetitive viral genomic regions to rapidly degrade EBV episomes. Using these approaches, we evaluated EBV-loss sensitivity across newly-derived, phenotypically diverse EBV-positive Burkitt lymphoma lines. We also evaluated cell lines from patients with EBV-associated T/NK and epithelial malignancies. We included the Burkitt Lymphoma cell line, Akata-GFP, as our EBV-loss-tolerant control and a lymphoblastoid cell line as our EBV-loss-sensitive control. We observed EBV-loss sensitivity in all the EBV-positive Burkitt Lymphoma cell lines and the chronic active EBV disease cell line. In contrast, the EBV-positive gastric adenocarcinoma lines were markedly less sensitive to EBV loss. EBV latency type did not predict EBV dependence, indicating that viral gene expression programs alone do not dictate reliance on the virus. These findings demonstrate that EBV dependence is prevalent across multiple EBV-associated disease models and is especially pronounced in early-state or newly derived lymphoid malignancies. AUTHOR SUMMARYEpstein-Barr virus (EBV) infects most people worldwide and is linked to many malignancies. Although EBV is present in cancer cells, it is unclear whether these cells require the virus to survive. To answer this question, we used CRISPR/Cas9-based methods to remove EBV from infected cells. Using these tools, we studied many different EBV-positive cell lines, including new Burkitt lymphoma cell lines. We found that most EBV-positive lymphoid cells required EBV for growth. In contrast, EBV-positive gastric cells were more tolerant of viral loss. These results show that many EBV-associated malignancies, especially early or newly established diseases, rely on EBV to survive.

microbiology↗

bioRxiv2025

Brain Region-Specific Epigenomic Reorganization and Altered Cell States in Alzheimer's Disease

Alzheimers disease (AD) is the most common neurodegenerative disorder, yet the molecular mechanisms underlying its region- and cell-type-specific pathogenesis remain poorly defined. Here, we generated a large-scale, single-cell multi-omic atlas--integrating DNA methylation and 3D genome architecture--from postmortem brain tissue of matched AD patients and cognitively normal controls. Samples were collected from three brain regions with distinct vulnerability to AD pathology: the temporal cortex (TC), primary visual cortex (VC), and prefrontal cortex (PFC). Our dataset comprises over 230,000 individual cells, spanning major neuronal and glial populations, and provides a high-resolution view of multi-layer epigenomic regulation. We identified widespread AD-associated DNA methylation changes and marked reorganization of 3D genome structure, including alterations in A/B compartments, topologically associating domains (TADs), and chromatin loops. These changes are strongly region-specific: TC displays pronounced hypermethylation, transcriptional downregulation, and elevated boundary density, whereas VC shows opposing trends and PFC an intermediate profile. We further uncovered previously unrecognized AD-associated glial and neuronal states defined by coordinated epigenomic dysregulation and recurrent genomic deletions, particularly near telomeric regions. This region-resolved, single-cell multi-omic atlas reveals divergent epigenomic trajectories across brain regions and cell types in AD, offering new mechanistic insights and a framework for targeted therapeutic strategies.

genomics↗

bioRxiv2025

PIN1 Drives Cellular Plasticity and Immune Modulation in Chronic Pancreatitis

Background and AimsChronic pancreatitis (CP) is characterized by inflammation, fibrosis, and acinar-to-ductal metaplasia (ADM). PIN1, known to drive oncogenic signaling and cellular plasticity in cancer, has an unexplored role in CP. This study investigates PIN1s expression and function in CP pathogenesis using human tissues and mouse models. MethodsPIN1 expression was assessed in human CP tissue microarrays (TMAs) via immunohistochemistry (IHC) and cyclic immunofluorescence (CyCIF). Acute and chronic pancreatitis were induced in wild-type (WT) and PIN1 knockout (PIN1KO) mice using caerulein. Disease progression was monitored histologically, and immune profiling was conducted using flow cytometry. Pharmacological inhibition was performed using a small molecule PIN1 inhibitor-Sulfopin, and effects were evaluated by histology, qPCR, and cytokine analysis. Single-cell RNA sequencing (scRNA-seq) was performed on pancreatic tissues to perform pathway analysis and intercellular communication. ResultsPIN1 expression was elevated in human CP tissues, correlating with disease severity and ADM. In mice, both acute and chronic pancreatitis increased PIN1 expression, but only in our chronic PIN1KO mice displayed reduced pancreatic injury, fibrosis, ADM, and modulated immune infiltration. Pharmacological PIN1 inhibition mimicked the protective effects of genetic knockout, dampening inflammatory pathways. scRNA-seq revealed that PIN1 inhibition altered the intercellular communication networks between epithelial, immune, and stromal cells. ConclusionPIN1 drives cellular plasticity, immune modulation, and disease progression in CP. Targeting PIN1 may offer a therapeutic strategy to mitigate CP. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=184 HEIGHT=200 SRC="FIGDIR/small/653850v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@67f9fcorg.highwire.dtl.DTLVardef@4d4c1forg.highwire.dtl.DTLVardef@c099d7org.highwire.dtl.DTLVardef@b41aa5_HPS_FORMAT_FIGEXP M_FIG C_FIG Created in BioRender. Shah, V. (2025) https://BioRender.com/undefined

cell biology↗

bioRxiv2025

DNA Methylation Dynamics of Dose-dependent Acute Exercise, Training Adaptation, and Detraining

Exercise and diet are direct physical contributors to human health, wellness, resilience, and performance1-5. Endurance and resistance training are known to improve healthspan through various biological processes such as mitochondrial function6-8, telomere maintenance9, and inflammaging10. Although several training prescriptions have been defined with specific merits 1,10-20, the long-term effects of these in terms of their molecular alterations have not yet been well explored. In this study, we focus on two combined endurance and resistance training programs: (1) traditional moderate-intensity continuous endurance and resistance exercise (TRAD) and (2) a variation of high-intensity interval training (HIIT) we refer to as high intensity tactical training (HITT), to assess the dynamics of DNA methylation (DNAm) in blood and muscle derived from males (N=23) and females (N=31), over a 12-week period of training followed by a 4-week period of detraining, sampled at pre-exercise and acute time points, totaling 528 samples. Due to its rapid responsiveness to stimuli and its stability, DNAm has been known to facilitate regulatory cascades that significantly affect various physiological processes and pathways. We find that several thousand differentially methylated regions (DMRs) associated with acute exercise in blood, many of which are shared across males and females. This trend is reversed when comparing the baseline (pre-exercise) time points or post-exercise timepoints at the untrained state with those at the post-conditioned state. Here, muscle shows majority of DNAm changes, with most of those being unique. We also find several hundred "memory" DMRs in muscle that maintain the gain or loss of methylation after four weeks of inactivity. Comparing phenotypic measurements, we find specific DMRs that correlate significantly with mitochondrial function and myofiber switching. Using machine learning, we select a subset of DMRs that are most characteristic of training modalities, sex and timepoint. Most of the DMRs are enriched in pathways associated with immune function, cell differentiation, and exercise adaptation. These findings reveal mechanisms by which exercise- and training-induced epigenetic changes alter immune surveillance, mitochondrial function, and inflammatory response, and underscore the relevance of epigenetic plasticity to health monitoring and wellness.

genomics↗

bioRxiv2025

Cell-type-specific transposable element demethylation and TAD remodeling in the aging mouse brain

Aging is a major risk factor for neurodegenerative diseases, yet underlying epigenetic mechanisms remain unclear. Here, we generated a comprehensive single-nucleus cell atlas of brain aging across multiple brain regions, comprising 132,551 single-cell methylomes and 72,666 joint chromatin conformation-methylome nuclei. Integration with companion transcriptomic and chromatin accessibility data yielded a cross-modality taxonomy of 36 major cell types. We observed that age-related methylation changes were more pronounced in non-neuronal cells. Transposable element methylation alone distinguished age groups, showing cell-type-specific genome-wide demethylation. Chromatin conformation analysis demonstrated age-related increases in TAD boundary strength with enhanced accessibility at CTCF binding sites. Spatial transcriptomics across 895,296 cells revealed regional heterogeneity during aging within identical cell types. Finally, we developed novel deep-learning models that accurately predict age-related gene expression changes using multi-modal epigenetic features, providing mechanistic insights into gene regulation. This dataset advances our understanding of brain aging and offers potential translational applications. HighlightsO_LISingle-cell multi-omic profiling maps the epigenetic and spatial transcriptomic landscape of brain aging across multiple regions. C_LIO_LICell-type-specific genome-wide demethylation of retrotransposable elements correlates with increased chromatin accessibility and expression. C_LIO_LIElevated TAD boundary strength emerges as a unique marker of brain aging associated with CTCF gaining accessibility. C_LIO_LIA novel deep-learning model reveals the significance of epigenetic features on age-related transcriptomic changes across genes. C_LI

genomics↗

bioRxiv2025

Human Body Single-Cell Atlas of 3D Genome Organization and DNA Methylation

Higher-order chromatin structure and DNA methylation are critical for gene regulation, but how these vary across the human body remains unclear. We performed multi-omic profiling of 3D genome structure and DNA methylation for 86,689 single nuclei across 16 human tissues, identifying 35 major and 206 cell subtypes. We revealed extensive changes in CG and non-CG methylation across almost all cell types and characterized 3D chromatin structure at an unprecedented cellular resolution. Intriguingly, extensive discrepancies exist between cell types delineated by DNA methylation and genome structure, indicating that the role of distinct epigenomic features in maintaining cell identity may vary by lineage. This study expands our understanding of the diversity of DNA methylation and chromatin structure and offers an extensive reference for exploring gene regulation in human health and disease.

genomics↗

bioRxiv2025

Source of dietary protein alters the abundance of proteases, intestinal epithelial and immune proteins both directly and via interactions with the gut microbiota

Dietary protein source differentially impacts long-term health outcomes, yet the specific host responses underlying these health effects remain unclear, including which are direct effects of protein source and which are microbiota-mediated. Using metagenome-informed metaproteomics, we determined that protein source altered the abundance of host proteins related to proteolysis, immunity, and the gut barrier as well as microbial function in mice. These stool host responses further reflected site-specific responses throughout the intestinal tract. Additionally, while some host responses were microbiota-dependent, others were microbiota-independent. These effects of dietary protein source on the host and microbiota translated to humans. Notably, microbial enzymes related to mucin degradation increased consistently in response to egg white protein in both mice and humans. Our results show that protein source impacts multiple aspects of host physiology through microbiota-dependent and independent routes, providing essential insights into mechanisms underpinning protein source-driven differences in health outcomes.

microbiology↗

bioRxiv2024

Fatty acid desaturases link cell metabolism pathways to promote proliferation of Epstein-Barr virus-infected B cells

Epstein-Barr virus (EBV) is a gamma herpesvirus that infects up to 95% of the human population by adulthood, typically remaining latent in the host memory B cell pool. In immunocompromised individuals, EBV can drive the transformation and rapid proliferation of infected B cells, ultimately resulting in neoplasia. The same transformation process can be induced in vitro, with EBV-infected peripheral blood mononuclear cells (PBMCs) forming immortalized lymphoblastoid cell lines (LCLs) within weeks. In this study, we found that the fatty acid desaturases stearoyl-CoA desaturase 1 (SCD1) and fatty acid desaturase 2 (FADS2) are upregulated by EBV, and crucial for EBV-induced B cell proliferation. Additionally, we found that pharmacological and genetic inhibition of both SCD1 and FADS2 results in a significantly greater reduction in proliferation and cell cycle arrest, compared to perturbing either enzyme individually. Additionally, we found that inhibiting either SCD1 or FADS2 alone hypersensitizes LCLs to palmitate-induced apoptosis, recapitulating studies in other tissue types. Further lipidomic and metabolic analysis of dual SCD1/FADS2-inhibited LCLs revealed an increase in free unsaturated fatty acids, a reduction of oxidative phosphorylation, and a reduction of glycolysis, thereby linking the activity of SCD1 and FADS2 to overall growth-promoting metabolism. Lastly, we show that SCD1 and FADS2 are important in the growth of clinically derived EBV+ immunoblastic lymphoma cells. Collectively, these data demonstrate a previously uncharacterized role of lipid desaturation in EBV+ transformed B cell proliferation, revealing a metabolic pathway that can be targeted in future anti-lymphoma therapies.

microbiology↗

bioRxiv2024

Dietary protein from different sources escapes host digestion and is differentially modified by the microbiota

Protein is an essential macronutrient and variations in its source and quantity have been shown to impact long-term health outcomes. Differential health impacts of dietary proteins from various sources are likely driven by differences in their digestibility by the host and subsequent availability to the intestinal microbiota. However, our current understanding regarding the fate of dietary proteins from different sources in the gut, specifically how component proteins within these sources interact with the host and the gut microbiota, is limited. To determine which dietary proteins are efficiently digested by the host, and which proteins escape host digestion and are used by the gut microbiota, we used high-resolution mass spectrometry to quantify proteins that constitute different dietary protein sources before and after digestion in germ-free and conventionally raised mice. We detected proteins from all sources in fecal samples of both germ-free and conventional mice suggesting that even protein sources with high digestive efficiency make it to the colon where they can serve as metabolic substrate for gut microbiota. Additionally, we found that specific component proteins of dietary protein sources were degraded to a greater extent in the presence of the microbiota. We found that specific proteins with functions that could potentially impact host health and physiology were differentially enriched in germ-free or conventionally raised mice. These findings reveal large differences in the fate of dietary protein from various sources in the gut which could explain some of their differential health impacts.

microbiology↗

bioRxiv2024

Metaproteomics and DNA metabarcoding as tools to assess dietary intake in humans

Objective biomarkers of food intake are a sought-after goal in nutrition research. Most biomarker development to date has focused on metabolites detected in blood, urine, skin or hair, but detection of consumed foods in stool has also been shown to be possible via DNA sequencing. An additional food macromolecule in stool that harbors sequence information is protein. However, the use of protein as an intake biomarker has only been explored to a very limited extent. Here, we evaluate and compare measurement of residual food-derived DNA and protein in stool as potential biomarkers of intake. We performed a pilot study of DNA sequencing-based metabarcoding (FoodSeq) and mass spectrometry-based metaproteomics in five individuals stool sampled in short, longitudinal bursts accompanied by detailed diet records (n=27 total samples). Dietary data provided by stool DNA, stool protein, and written diet record independently identified a strong within-person dietary signature, identified similar food taxa, and had significantly similar global structure in two of the three pairwise comparisons between measurement techniques (DNA-to-protein and DNA-to-diet record). Metaproteomics identified proteins including myosin, ovalbumin, and beta-lactoglobulin that differentiated food tissue types like beef from dairy and chicken from egg, distinctions that were not possible by DNA alone. Overall, our results lay the groundwork for development of targeted metaproteomic assays for dietary assessment and demonstrate that diverse molecular components of food can be leveraged to study food intake using stool samples.

molecular biology↗

bioRxiv2024

Dietary protein source strongly alters gut microbiota composition and function

The source of protein in a persons diet affects their total life expectancy. However, the mechanisms by which dietary protein sources differentially impact human health and life expectancy are poorly understood. Dietary choices have major impacts on the composition and function of the intestinal microbiota that ultimately modulate host health. This raises the possibility that health outcomes based on dietary protein sources might be driven by interactions between dietary protein and the gut microbiota. In this study, we determined the effects of seven different sources of dietary protein on the gut microbiota of mice using an integrated metagenomics-metaproteomics approach. The protein abundances measured by metaproteomics can provide microbial species abundances, and evidence for the molecular phenotype of microbiota members because measured proteins indicate the metabolic and physiological processes used by a microbial community. We showed that dietary protein source significantly altered the species composition and overall function of the gut microbiota. Different dietary protein sources led to changes in the abundance of microbial proteins involved in the degradation of amino acids and the degradation of glycosylations conjugated to dietary protein. In particular, brown rice and egg white protein increased the abundance of amino acid degrading enzymes. Egg white protein increased the abundance of bacteria and proteins usually associated with the degradation of the intestinal mucus barrier. These results show that dietary protein sources can change the gut microbiotas metabolism, which could have major implications in the context of gut microbiota mediated diseases.

microbiology↗

bioRxiv2023

Large Quantities of Bacterial DNA and Protein in Common Dietary Protein Source Used in Microbiome Studies

Diet has been shown to greatly impact the intestinal microbiota. To understand the role of individual dietary components, defined diets with purified components are frequently used in diet-microbiota studies. Many of the frequently used defined diets use purified casein as the protein source. Previous work indicated that this casein contains microbial DNA potentially impacting results of microbiome studies. Other diet-based microbially derived molecules that may impact microbiome measurements, such as proteins detected by metaproteomics, have not been determined for casein. Additionally, other protein sources used in microbiome studies have not been characterized for their microbial content. We used metagenomics and metaproteomics to identify and quantify microbial DNA and protein in a casein-based defined diet to better understand potential impacts on metagenomic and metaproteomic microbiome studies. We further tested six additional defined diets with purified protein sources with an integrated metagenomic-metaproteomic approach and show that contaminating microbial protein is unique to casein within the tested set as microbial protein was not identified in diets with other protein sources. We also illustrate the contribution of diet-derived microbial protein in diet-microbiota studies by metaproteomic analysis of stool samples from germ-free mice (GF) and mice with a conventional microbiota (CV) following consumption of diets with casein and non-casein protein. This study highlights a potentially confounding factor in diet-microbiota studies that must be considered through evaluation of the diet itself within a given study. ImportanceMany diets used in diet-microbiota studies use casein as the source of dietary protein. We found large quantities of microbial DNA and protein in casein-based diets. This microbial DNA and protein are resilient to digestion as it is present in fecal samples of mice consuming casein-based diets. This contribution of diet-derived microbial DNA and protein to microbiota measurements may influence results and conclusions and must therefore be considered in diet-microbiota studies. We tested additional dietary protein sources and did not detect microbial DNA or protein. Our findings highlight the necessity of evaluating diet samples in diet-microbiota studies to ensure that potential microbial content of the diet can be accounted for in microbiome measurements.

microbiology↗

bioRxiv2023

Cell type-specific effects of age and sex on human cortical neurons

Excitatory and inhibitory neurons establish specialized identities early in life through cell type-specific patterns of epigenetic regulation and gene expression. Although cell types are largely stable throughout the lifespan, altered transcriptional and epigenetic regulation may contribute to cognitive changes with advanced age. Using single-nucleus multiomic DNA methylation and transcriptome sequencing (snmCT-seq) in frontal cortex samples from young adult and aged donors, we found widespread age- and sex-related variability in specific neuronal cell types. The proportion of GABAergic inhibitory cells, including SST and VIP expressing cells, was reduced in aged donors. On the other hand, excitatory neurons had more profound age-related changes in their gene expression and DNA methylation compared with inhibitory cells. Hundreds of genes involved in synaptic activity were downregulated, while genes located in subtelomeric regions were upregulated with age and anti-correlated with telomere length. We further mapped sex differences in autosomal gene expression and escape from X-inactivation in specific neuron types. Multiomic single-nucleus epigenomes and transcriptomes provide new insight into the effects of age and sex on human neurons.

neuroscience↗

bioRxiv2023

Epigenome erosion in Alzheimer's disease brain cells and induced neurons

Late-onset Alzheimers disease (LOAD) is typically sporadic, correlated only to advanced age, and has no clear genetic risk factors. The sporadic nature of LOAD presents a challenge to understanding its pathogenesis and mechanisms. Here, we comprehensively investigated the epigenome of LOAD primary entorhinal cortex brain tissues via single-cell multi-omics technologies, simultaneously capturing DNA methylation and 3D chromatin conformation. We identified AD-specific DNA methylation signatures and found they interact with bivalent promoters of AD differentially expressed genes. In addition, we discovered global chromosomal epigenome erosion of 3D genome structure within and across brain cell types. Furthermore, to evaluate whether these age- and disease-dependent molecular signatures could be detected in the in vitro cellular models, we derived induced neurons (iNs) converted directly from AD patients fibroblasts and found a set of conserved methylation signatures and shared molecular processes. We developed a machine-learning algorithm to identify robust and consistent methylation signatures of LOAD in vivo primary brain tissues and in vitro fibroblast-derived iNs. The results recapitulate the age- and disease-related epigenetic features in iNs and highlight the power of epigenome and chromatin conformation for identifying molecular mechanisms of neuronal aging and generating biomarkers for LOAD. HIGHLIGHTO_LIAD-specific DNA methylation signatures are identified in entorhinal cortex brain cell types C_LIO_LIThe AD differentially expressed genes linked with differentially methylated regions via loop interactions are enriched in a bivalent chromatin state C_LIO_LIChromosomal epigenome erosion of 3D genome structures occurs in LOAD brain cell types. C_LIO_LIShared and reliable methylation signatures are observed in both in vitro cellular iN models and primary brain tissues. C_LIO_LIMachine learning models identify robust and reliable methylation loci as AD biomarkers across cell types. C_LI

genomics↗

bioRxiv2023

Intracellular K+ limits T cell exhaustion and preserves antitumor function

The cancer-killing activity of T cells is often compromised within tumors, allowing disease progression. We previously found that intratumoral elevations in extracellular K+ related to ongoing cell death constrained CD8+ T cell Akt-mTOR signaling and effector function (1,2). To alleviate K+ mediated T cell suppression, we pursued genetic means to lower intracellular K+. Transcriptomic analysis of CD8+ T cells demonstrated the Na+/K+ ATPase to be robustly and dynamically expressed. CRISPR-Cas9 mediated deletion of the catalytic alpha subunit of the Na+/K+ ATPase lowered intracellular K+ but produced tonic hyperactivity in multiple signal transduction cascades along with the acquisition of co-inhibitory receptors and terminal differentiation in mouse and human CD8+ T cells. Mechanistically, Na+/K+ ATPase disruption led to ROS accumulation due to depletion of intracellular K+ in T cells. Antioxidant treatment or high K+ media prevented Atp1a1 deficient T cells from exhausted T (TEx) cell formation. Consistent with transcriptional and proteomic data suggesting a TEx cell phenotype, T cells lacking Atp1a1 had compromised persistence and antitumor activity in a syngeneic model of orthotopic murine melanoma. Translational application of these findings will include efforts to lower intracellular K+ while limiting ROS accumulation within tumor specific T cells. SynopsisHigh extracellular K+ ({uparrow}[K+]e) is found within tumors and suppresses T cell effector function. Collier et al. find that deletion of the Na+/K+ ATPase in T cells lowers intracellular K+ and promotes ROS accumulation, tonic signal transduction and T cell exhaustion owing to ROS accumulation. Engineering T cell ion transport is an important consideration for cancer immunotherapy.

immunology↗