Biology subjects

Bar, D. Z.

Publications and source records attributed to Bar, D. Z..

11 recordsLinked to original sources

bioRxiv2026

Loss of cell-identity DNA methylation tracks division history, not DNA damage

One influential proposal holds that DNA damage drives aging by degrading the epigenome, so that a cell loses the information that specifies what it is. We test that specific proposed driver, not the broader idea that aging involves a loss of epigenetic information. Cell-type-unique CpGs are a direct place to test it. They are the sites where a cell's methylome differs most from every other cell type, so they are where its identity information can be accurately measured. We built such a panel for human skin fibroblasts against twenty primary cell types, and found that most of its 9,150 CpGs behave as a record of proliferation rather than identity. Their methylation state in fibroblasts is shared with continuously proliferating cell lines, consistent with the bulk hypomethylation of late-replicating chromatin that accumulates in culture, and this layer must be separated from identity marks before identity loss can be tested. We then expressed every perturbation as the fraction of the panel's identity information lost beyond baseline-matched background CpGs, using a Bernoulli cross-entropy between the fibroblast code and a generic comparator cell. In 24 matched fibroblast cultures, replicative exhaustion cost 24.3% of the identity information (95% CI 20.2 to 28.7) and RAS activation cost 5.4% (4.6 to 6.1), both at p = 1.2 X 10^-7;. A senescence-inducing dose of X-rays cost nothing: -0.5% (-0.7 to -0.2), with identity CpGs if anything moving slightly further from other cell types than matched background did. The design could detect a loss of 0.4%, about one sixtieth of the exhaustion effect, so the X-ray result is a bounded negative rather than an absence of power. Doxorubicin-induced and irradiation-induced senescence did not move the panel either. Clones that kept dividing after irradiation moved further from other cell types rather than collapsing toward them. In human fibroblasts, loss of identity information tracks prolonged division, not acute DNA damage.

molecular biology↗

bioRxiv2026

A fusion Cell-Permeable C16orf74 Peptide Selectively Disrupts Calcineurin-NFAT Interaction and Inhibits T-cell Activation Without Cytotoxicity

Calcineurin (Cn) is a protein phosphatase that initiates T-cell activation by dephosphorylating the transcription factor NFAT, driving its nuclear translocation and the transcription of immune-related genes. While clinical immunosuppressants like Cyclosporine A (CsA) potently inhibit Cn, they completely block its catalytic site, leading to non-specific inhibition and severe off-target toxicity. Selectively targeting the specific protein-protein interaction (PPI) between Cn and NFAT presents a safer therapeutic strategy. We previously identified the C16orf74 (C16) peptide as a high-affinity Cn-NFAT PPI inhibitor; however, its utility in cellular systems is restricted by poor membrane permeability. In this study, we evaluated cell-penetrating peptide (CPP) conjugates of C16 with an N-terminus transactivator of transcription (TAT) and polyarginine (R11) to enable efficient intracellular delivery. Structural modeling, fluorescence polarization displacement, and pull-down assays confirmed that the CPP-C16 conjugates retain the ability to compete with an NFAT-derived peptide and bind Cn. Fluorescence microscopy demonstrated efficient intracellular entry of TAT-C16 and R11-C16 in mammalian cells, and effective inhibition of NFAT nuclear translocation and attenuation of downstream NFAT-dependent transcriptional activity of the IL-2 gene in human T cells at concentrations of 10 {micro}M or lower. Crucially, unlike CsA, the CPP-C16 peptides exhibited minimal cytotoxicity even at high concentrations of up to 50 {micro}M, establishing a potential safe therapeutic window. These findings establish CPP-C16 conjugates as effective, cell-permeable, and non-toxic inhibitors of the Cn-NFAT signaling axis, providing the basis for the development of PPI-directed immunosuppressants.

bioengineering↗

bioRxiv2025

Colon-Specific Epigenetic Clocks from Minimal Features Reveal Disease-Driven Aging

Epigenetic clocks estimate chronological and biological age from DNA methylation patterns, but conventional models typically train on hundreds of thousands of CpG sites and large training cohorts. We previously demonstrated that tissue-unique methylation sites change in a predictable manner upon aging and disease. Here, we demonstrate that clocks built from tissue-unique methylation sites enable accurate age prediction in the human colon using a compact feature set and limited training data. We trained a machine learning model on healthy colon tissue, identifying CpG sites that capture both chronological age and anatomical location (proximal vs. distal). This clock maintains high predictive performance (r = 0.978; MAE 3.9 years) while using an order of magnitude fewer sites and samples than traditional approaches. Applying the model to tissues from individuals with HIV infection, inflammatory bowel disease (IBD), and colonic polyps reveals consistent patterns of accelerated aging, while aspirin treatment is associated with partial deceleration. Our findings establish tissue-unique CpGs as a powerful basis for efficient, interpretable clocks and offer new insights into how chronic inflammation and neoplasia shape the aging landscape of the colon.

molecular biology↗

bioRxiv2025

Reversible and Causal Epigenetic Information Loss in Liver Aging and Disease

The loss of epigenetic information has been proposed as a driver of aging and diseases, but the reversibility and causality of this process remain underexplored. Here we analyze liver-unique methylation sites - genomic loci that show distinct methylation patterns in the liver compared to other tissues. Upon disease progression, these sites overwhelmingly regress toward the pan-tissue average. In addition, we demonstrate that this regression also occurs in a majority of these sites during normal aging. Using Mendelian randomization analysis, we identify significant enrichment of liver-unique methylation sites in causal aging-associated loci, particularly sites that are highly methylated in healthy liver. Remarkably, repeated fasting, a metabolic intervention known to improve liver function, partially restores the liver-unique methylation patterns at these sites. This restoration also occurs in isolated hepatocytes subjected to fasting-mimicking conditions, suggesting the effect is cell-autonomous rather than due to changes in tissue composition. The liver-unique methylation sites are enriched for binding sites of key metabolic transcription factors and show significant overlap with genetic variants associated with liver disease risk, suggesting a mechanistic link between epigenetic information loss and liver dysfunction. Our findings establish epigenetic information loss as both a marker and mediator of liver aging and disease, while demonstrating its potential reversibility through metabolic interventions. Graphical abstract: Reversible information loss at liver-unique methylation sitesLiver-unique sites, showing higher (UH) or lower (UL) methylation levels, regress to the pan-tissue average upon aging and disease. UH are enriched for methylation sites causal to the aging process, while UL are enriched for liver-specific enhancers and PPAR- binding sites. Upon repeated fasting, both UL and UH diverge away from the pan-tissue average, partially restoring the more youthful and disease-free epigenetic state. O_FIG O_LINKSMALLFIG WIDTH=174 HEIGHT=200 SRC="FIGDIR/small/639802v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@6d9ce9org.highwire.dtl.DTLVardef@58a2f6org.highwire.dtl.DTLVardef@132ff20org.highwire.dtl.DTLVardef@10b9acc_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Anti-fungal recombinant psoriasin effectively inhibits Candida albicans growth on denture base

Oral candidiasis leading to denture stomatitis is a fungal infection resulting from unregulated growth and adhesion mainly of Candida albicans onto acrylic denture base. Once the biofilm is formed, it is immune resistant and mainstay treatments involve toxic chemical antifungal agents or mechanical cleaning techniques, both offer limited efficacy. Consequently, there is an urgent need for more effective and safer therapeutic approaches. While biological modalities are expanding in general medicine, the exploration of protein-based therapeutics in dental medicine remains limited. This research evaluates the inhibitory effect of recombinantly expressed psoriasin on the growth of Candida albicans on polymethyl methacrylate denture bases. Psoriasin, also known as S100-A7, has shown promise in treating microbial skin infections, and its natural presence in saliva makes it a promising candidate for treating oral microbial infections. Our findings indicate that psoriasin exhibits a strong, dose-dependent inhibition of Candida albicans growth. Further, we incubated a polymethyl methacrylate denture base within the psoriasin solution. Notably, immersing the denture base in the solution completely eradicated fungal growth. Our research utilizes natural antifungal proteins within biomedical devices like denture bases, suggesting psoriasin as a safe alternative to chemical antifungals in dental medicine.

biochemistry↗

bioRxiv2023

Detecting Protein-DNA Binding in Single Molecules using Antibody Guided Methylation

Characterization of DNA binding sites for specific proteins is of fundamental importance in molecular biology. It is commonly addressed experimentally by chromatin immunoprecipitation and sequencing (ChIP-seq) of bulk samples (103-107 cells). We have developed an alternative method that uses a Chromatin Antibody-mediated Methylating Protein (ChAMP) composed of a GpC methyltransferase fused to protein G. By tethering ChAMP to a primary antibody directed against the DNA-binding protein of interest, and selectively switching on its enzymatic activity in situ, we generated distinct and identifiable methylation patterns adjacent to the protein binding sites. This method is compatible with methods of single-cell methylation-detection and single molecule methylation identification. Indeed, as every binding event generates multiple nearby methylations, we were able to confidently detect protein binding in long single molecules. O_FIG O_LINKSMALLFIG WIDTH=199 HEIGHT=200 SRC="FIGDIR/small/567792v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@dac365org.highwire.dtl.DTLVardef@522ecaorg.highwire.dtl.DTLVardef@17ddb38org.highwire.dtl.DTLVardef@1c371b0_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO (i) ChAMP is added to fix and permeabilized cells, where it binds (ii) any antibody, and upon the addition of SAM, methylates nearby GpC sites, to be detected by sequencing (iii-iv). C_FIG

molecular biology↗

bioRxiv2023

Immunorecognition of Streptococcus mutans secreted proteins protects against caries by limiting tooth adhesion

Childhood caries, a chronic disease affecting 60-90% of children in industrialized countries, results in lesions in primary and permanent dentition, leading to hospitalizations, emergency room visits, high treatment costs, and loss of school days. It diminishes the childs ability to learn and increases the risk of caries in adulthood. Despite multiple risk factors for caries, significant interpersonal variability unaccounted for by these factors exists. The immune system generates a personal antibody repertoire that helps maintain a balanced and healthy oral microbiome. Streptococcus mutans is a key bacterium in caries development. Utilizing mass-spectrometry, we examined which S. mutans proteins are identified by antibodies of children with and without caries and identified a core set of proteins recognizable by the immune system of most individuals. This set was enriched with proteins enabling bacterial adhesion. To study the physiological relevance of these findings, we tested the ability of saliva to prevent S. mutans adherence to tooth surfaces. Saliva from caries-free individuals, but not children with caries, was found to hinder the binding of S. mutans to teeth. These findings delineate the S. mutans proteome targeted by the immune system and suggest that the inhibition of bacterial adherence to teeth is a primary mechanism employed by the immune system to maintain oral balance and prevent caries formation. These discoveries offer fresh insights into the immune systems role in preserving oral health and preventing caries development. Targeting S. mutans proteins implicated in bacterial adhesion could be a promising strategy for preventing childhood caries.

microbiology↗

bioRxiv2023

Epigenetic information loss is a common feature of multiple diseases and aging

Aging is a major risk factor for a plethora of diseases. The information theory of aging posits that epigenetic information loss is a principal driver of the aging process. Despite this, the connection between epigenetic information loss and disease has not been thoroughly investigated. Here, we analyzed tissue-unique methylation patterns in healthy and diseased organs, revealing that for several diseases these patterns degrade, regressing to a mean form. We interpret this as epigenetic information loss, where tissue-unique patterns erode. Information loss is not limited to diseases. Age-related erosion of unique methylation patterns was observed in some tissues and cells, while other tissues and cells diverged away from the mean. Our findings demonstrate that analyzing methylation patterns in tissue-unique sites can effectively distinguish between patients and healthy controls across a range of diseases, and underscore the role of epigenetic information loss as a common feature in various pathological conditions. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=191 SRC="FIGDIR/small/539727v3_ufig1.gif" ALT="Figure 1"> View larger version (27K): org.highwire.dtl.DTLVardef@12049e0org.highwire.dtl.DTLVardef@d214c7org.highwire.dtl.DTLVardef@52425forg.highwire.dtl.DTLVardef@839011_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical abstractC_FLOATNO Tissue unique methylation pattern regress toward the mean upon disease. A single methylation site, showing low methylation in the liver and high in every other tissue, becomes more methylated in diseased livers. C_FIG

molecular biology↗

bioRxiv2022

Kidney-specific methylation patterns correlate with kidney function and are lost upon kidney disease progression

Chronological and biological age correlate with DNA methylation levels at specific sites in the genome. Linear combinations of multiple methylation sites, termed epigenetic clocks, can inform us of the chronological age and predict multiple health-related outcomes. However, why some sites correlate with lifespan, healthspan, or specific medical conditions remains poorly understood. Kidney fibrosis is the common pathway for Chronic Kidney Disease, which affects 10% of Europe and USA population. Here we identify epigenetic clocks and methylation sites that correlate with kidney function. Moreover, we identify methylation sites that have a unique methylation signature in the kidney. Methylation levels in the majority of these sites correlates with kidney state and function. When kidney function deteriorates, all of these sites regress towards the common methylation pattern observed in other tissues. Interestingly, while the majority of sites are less methylated in the kidney and become more methylated with loss of function, a fraction of the sites are highly methylated in the kidney and become less methylated when kidney function declines. These methylation sites are enriched for specific transcription-factor binding sites. In a large subset of sites, changes in methylation pattern are accompanied by changes in gene expression in kidneys of chronic kidney disease patients. These results support the information theory of aging, and the hypothesis that the unique tissue identity, as captured by methylation patterns, is lost as tissue function declines. However, this information loss is not random, but guided towards a baseline that is dependent on the genomic loci.

bioinformatics↗

bioRxiv2022

The hidden secrets of the dental calculus: Calibration of a mass spectrometry protocol for dental calculus protein analysis

Dental calculus is a solid deposit that forms and accumulates on the tooth surface, entrapping oral microorganisms, biomolecules, and other micro-debris found in the oral cavity. Mass spectrometry analysis of its protein content opens a vista into the subjects diet, oral flora, and even some aspects of health, thus providing new insight and expanding our knowledge of archaic cultures. Multiple experimental protocols have been proposed for the optimal extraction of proteins from dental calculus. Herein, we compared various experimental conditions in order to calibrate and validate a protocol for protein extraction. Our results show that a high concentration of acetic acid followed by mechanical crushing and sonication provided the highest protein yield, while acetone precipitation enabled the identification of more distinct proteins. We validated this protocol using archeological samples, identifying human and microbial proteins in specimens from the 8th and 17th centuries (approximately 250-1300 years ago). These findings demonstrate that the developed protocol is useful for studying excavated archaeological samples and that it might be utilized to explore the biohistory, dietary habits, and microbiome of archaic populations.

molecular biology↗

bioRxiv2020

Lamin regulates the dietary restriction response via the mTOR pathway in Caenorhabditis elegans

Animals subjected to dietary restriction (DR) have reduced body size, low fecundity, slower development, lower fat content and longer life span. We identified lamin as a regulator of multiple dietary restriction phenotypes. Downregulation of lmn-1, the single Caenorhabditis elegans lamin gene, increased animal size and fat content, specifically in DR animals. The LMN-1 protein acts in the mTOR pathway, upstream to RAPTOR and S6K, key component and target of mTOR complex 1 (mTORC1), respectively. DR excludes the mTORC1 activator RAGC-1 from the nucleus. Downregulation of lmn-1 restores RAGC-1 to the nucleus, a necessary step for the activation of the mTOR pathway. These findings further link lamin to metabolic regulation.

cell biology↗