Biology subjects

Saul, D.

Publications and source records attributed to Saul, D..

13 recordsLinked to original sources

bioRxiv2026

SenoQuant: One-stop AI software for senescence marker analysis and prediction

Senescent cells accumulate with age and contribute to tissue dysfunction, yet their identification in tissues is challenging due to low abundance, heterogeneous phenotypes, and the lack of specific markers. Senescence-associated features span multiple subcellular compartments, including nuclear DNA damage foci, cytosolic protein changes, and perinuclear alterations, each requiring tailored detection strategies. To overcome these challenges, we developed SenoQuant (https://github.com/HaamsRee/senoquant), a versatile software designed for comprehensive, accurate, and unbiased spatial quantification and prediction of senescence markers across diverse tissue contexts. Utilizing AI models, SenoQuant enables precise nuclear and cytoplasmic segmentation and detection of senescence markers across low- and high-plex imaging modalities, applicable to cultured cells and tissue sections from mice and humans. The platform also supports custom AI models; for example, we built SenCeption, a proof-of-concept predictor of single-cell p21 status from DAPI-stained nuclei in human skin. Available as a free napari plugin, SenoQuant is widely accessible to researchers. By providing a unified approach to senescence analysis and prediction, SenoQuant opens new opportunities for exploring the complex biology of senescence and its impacts on aging and disease.

Cell Biology↗

bioRxiv2026

Intrinsically disordered N-terminal and structured DNA-binding domains jointly regulate progesterone receptor transcriptional condensates

Transcriptional condensates (TCs) are dynamic, membrane-less assemblies of proteins and nucleic acids that form at specific genomic loci to control gene expression. The primary drivers of TC formation and function are long stretches of polypeptide chains that lack well-defined tertiary structure, known as the intrinsically disordered regions (IDRs) of Transcription Factors (TFs). However, the role of well-structured domains such as the DNA-binding domain (DBD) in condensate formation and downstream regulation remains poorly understood. Here, we investigated whether the DBD of the Progesterone Receptor (PR), a model TF regulated by hormone binding, contributes to the formation of PR TCs. By combining Single Particle Tracking (SPT) with deep-learning-based single-trajectory analysis, we demonstrated that both the IDR and the DBD are required for the formation of PR condensates. Furthermore, transcriptomic analysis showed that mutations of either of the two domains result in the expression of a distinct set of genes, with functional assays corroborating these findings and linking domain-specific transcription to differential effects on cell proliferation and migration. Together, our results show that domains beyond IDRs also play a role in TC formation and function and uncover domain-specific regulation of transcriptional and oncogenic programs.

biophysics↗

bioRxiv2025

Epigenetic Context Defines the Transcriptional Activity of Canonical and Noncanonical NF-κB Signaling in Pancreatic Cancer

NF-{kappa}B signaling can be subdivided into canonical and noncanonical pathways, culminating in the transcriptional activity of RELA and RELB, respectively. However, the upstream signals that activate these transcription factors and their specific regulatory roles in pancreatic ductal adenocarcinoma (PDAC) remain incompletely understood. Here, we demonstrate that TNF is the primary activator of canonical NF-{kappa}B signaling via RELA, while TWEAK engages noncanonical signaling through RELB in PDAC. Using transcriptome-wide gene expression, genome-wide occupancy profiling, and epigenome mapping, we delineate distinct temporal dynamics and regulatory activities of each pathway. Single-cell RNA-seq analysis and multiplex immunofluorescence staining in primary PDAC samples further reveal extensive and distinct interactions with components of the tumor microenvironment. Motif analysis of RELA- and RELB-bound regions uncovers a particularly strong association of RELB with AP1 elements. Notably, while RELA binds to both open and closed chromatin, RELB exclusively occupies regions with pre-existing chromatin accessibility and AP1 co-binding. Collectively, these findings underscore the distinct and complementary roles of TNF and TWEAK in PDAC, with TNF engaging a broader transcriptional program via RELA and TWEAK selectively targeting genes marked by accessible chromatin and epigenetic activity. This study provides critical insights into the regulatory dynamics of NF-{kappa}B signaling in PDAC, highlighting unique and complementary functions of RELA and RELB in modulating downstream gene expression. SignificanceTNF and TWEAK selectively activate RELA and RELB in pancreatic cancer, revealing that RELB-driven noncanonical signaling depends on chromatin accessibility and AP-1, uncovering distinct, targetable epigenetic dependencies from canonical signaling. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=59 SRC="FIGDIR/small/661561v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@14aa356org.highwire.dtl.DTLVardef@1297741org.highwire.dtl.DTLVardef@1545741org.highwire.dtl.DTLVardef@2c982b_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

IL-17A potentiates malignant T-cell viability via electron transport chain complex I in cutaneous T-cell lymphoma

Cutaneous T-cell lymphoma (CTCL) is a rare malignancy of skin-resident T cells characterized by an aberrant cytokine profile, notably elevated Interleukin-17A (IL-17A) levels. However, the functional importance of IL-17A dysregulation in the CTCL pathogenesis remains poorly understood. Our analysis revealed increased IL-17A receptor (IL-17RA) expression in CTCL patient samples, supporting the importance of IL-17A signaling in CTCL progression. Proteomic and metabolomic profiling of IL-17A-treated malignant T cells revealed mitochondrial electron transport chain (ETC) complex I as a key downstream target of IL-17A signaling. Furthermore, we confirmed the elevated expression of ETC complex I in CTCL patient samples compared to healthy control T cells, hence highlighting the clinical relevance of the IL-17A signaling-ETC complex 1 axis. Altogether, our data unveil the function of IL-17A signaling in the metabolic axis in CTCL progression, which could be useful for developing advanced therapeutic strategies. Significance statement: We uncover an IL-17A-induced electron transport chain (ETC) complex I pro-tumorigenic circuit and provide proof of concept evidence that ETC complex I can be targeted to combat the challenges posed by Cutaneous T-cell Lymphoma aggressiveness.

cancer biology↗

bioRxiv2024

The Multifaceted Phenotype of Senescent HL-60/S4 Macrophages

Every cell has a multifaceted phenotype. Transcriptional analysis of functionally defined groups of genes can provide insight into this phenotypic complexity. In the present study, the mRNA transcriptome of phorbol ester (TPA) differentiated HL-60/S4 macrophage cells was scrutinized using Gene Set Enrichment Analysis (GSEA), which evaluates the strengths of various cellular phenotypes by examining the enrichment of functionally different gene sets. Employing GSEA, we obtained supporting evidence that HL-60/S4 macrophages are senescent, probably a consequence of enriched TGF{beta} and NOTCH signaling transcripts. There appears to be a reduction of transcripts for heterochromatin, nucleosome formation, and chromatin remodeling phenotypes. In addition, despite upregulated oxidative stress gene transcription, we observed a reduction of DNA damage and repair transcripts. GSEA indicated that transcripts for autophagy, extracellular matrix, and inflammation/inflammasomes are enriched. We also observed that the HL-60/S4 macrophage is enriched for apoptosis gene transcripts, which may promote necrotic death by pyroptosis. The long-term goal of this research direction is to see whether this complex multifaceted phenotypic pattern is shared with other types of macrophages and to determine what mechanisms might exist to coordinate these phenotypic facets within a single cell.

cell biology↗

bioRxiv2024

Osteochondroprogenitor cells and neutrophils expressing p21 and senescence markers modulate fracture repair

Cells expressing features of senescence, including upregulation of p21 and p16, appear transiently following tissue injury, yet the properties of these cells or how they contrast with age-induced senescent cells remains unclear. Here, we used skeletal injury as a model and identified the rapid appearance following fracture of p21+ cells expressing senescence markers, mainly as osteochondroprogenitors (OCHs) and neutrophils. Targeted genetic clearance of p21+ cells suppressed senescence-associated signatures within the fracture callus and accelerated fracture healing. By contrast, p21+ cell clearance did not alter bone loss due to aging; conversely, p16+ cell clearance, known to alleviate skeletal aging, did not affect fracture healing. Following fracture, p21+ neutrophils were enriched in signaling pathways known to induce paracrine stromal senescence, while p21+ OCHs were highly enriched in senescence-associated secretory phenotype factors known to impair bone formation. Further analysis revealed an injury-specific stem cell-like OCH subset that was p21+ and highly inflammatory, with a similar inflammatory mesenchymal population (fibro-adipogenic progenitors) evident following muscle injury. Thus, intercommunicating senescent-like neutrophils and mesenchymal progenitor cells are key regulators of tissue repair in bone and potentially across tissues. Moreover, our findings establish contextual roles of p21+ vs p16+ senescent/senescent-like cells that may be leveraged for therapeutic opportunities.

physiology↗

bioRxiv2023

Distinct secretomes in p16- and p21- positive senescent cells across tissues

Senescent cells drive age-related tissue dysfunction via the induction of a chronic senescence-associated secretory phenotype (SASP). The cyclin-dependent kinase inhibitors p21Cip1 and p16Ink4a have long served as markers of cellular senescence. However, their individual roles remain incompletely elucidated. Thus, we conducted a comprehensive examination of multiple single-cell RNA sequencing (scRNA-seq) datasets spanning both murine and human tissues during aging. Our analysis revealed that p21Cip1 and p16Ink4a transcripts demonstrate significant heterogeneity across distinct cell types and tissues, frequently exhibiting a lack of co-expression. Moreover, we identified tissue-specific variations in SASP profiles linked to p21Cip1 or p16Ink4a expression. Our study underscores the extraordinary diversity of cellular senescence and the SASP, emphasizing that these phenomena are inherently cell- and tissue-dependent. However, a few SASP factors consistently contribute to a shared "core" SASP. These findings highlight the need for a more nuanced investigation of senescence across a wide array of biological contexts.

cell biology↗

bioRxiv2023

In vitro and in vivo effects of zoledronate on senescence and senescence-associated secretory phenotype markers

In addition to reducing fracture risk, zoledronate has been found in some studies to decrease mortality in humans and extend lifespan and healthspan in animals. Because senescent cells accumulate with aging and contribute to multiple co-morbidities, the non-skeletal actions of zoledronate could be due to senolytic (killing of senescent cells) or senomorphic (inhibition of the secretion of the senescence-associated secretory phenotype [SASP]) actions. To test this, we first performed in vitro senescence assays using human lung fibroblasts and DNA repair-deficient mouse embryonic fibroblasts, which demonstrated that zoledronate killed senescent cells with minimal effects on non-senescent cells. Next, in aged mice treated with zoledronate or vehicle for 8 weeks, zoledronate significantly reduced circulating SASP factors, including CCL7, IL-1{beta}, TNFRSF1A, and TGF{beta}1 and improved grip strength. Analysis of publicly available RNAseq data from CD115+ (CSF1R/c-fms+) pre-osteoclastic cells isolated from mice treated with zoledronate demonstrated a significant downregulation of senescence/SASP genes (SenMayo). To establish that these cells are potential senolytic/senomorphic targets of zoledronate, we used single cell proteomic analysis (cytometry by time of flight [CyTOF]) and demonstrated that zoledronate significantly reduced the number of pre-osteoclastic (CD115+/CD3e-/Ly6G-/CD45R-) cells and decreased protein levels of p16, p21, and SASP markers in these cells without affecting other immune cell populations. Collectively, our findings demonstrate that zoledronate has senolytic effects in vitro and modulates senescence/SASP biomarkers in vivo. These data point to the need for additional studies testing zoledronate and/or other bisphosphonate derivatives for senotherapeutic efficacy.

physiology↗

bioRxiv2023

Multiparametric senescent cell phenotyping reveals CD24 osteolineage cells as targets of senolytic therapy in the aged murine skeleton

Senescence drives organismal aging, yet the deep characterization of senescent cells in vivo remains incomplete. Here, we applied mass cytometry by time-of-flight (CyTOF) using carefully validated antibodies to analyze senescent cells at single-cell resolution. We used multiple criteria to identify senescent mesenchymal cells that were growth arrested and resistant to apoptosis (p16+/Ki67-/BCL-2+; "p16KB" cells). These cells were highly enriched for senescence-associated secretory phenotype (SASP) and DNA damage markers and were strongly associated with age. p16KB cell percentages were also increased in CD24+ osteolineage cells, which exhibited an inflammatory SASP in aged mice and were robustly cleared by both genetic and pharmacologic senolytic therapies. Following isolation, CD24+ skeletal cells exhibited growth arrest, SA-{beta}gal positivity, and impaired osteogenesis in vitro. These studies thus provide a new approach using multiplexed protein profiling by CyTOF to define senescent mesenchymal cells in vivo and identify a highly inflammatory, senescent CD24+ osteolineage population cleared by senolytics.

cell biology↗

bioRxiv2022

Identification of a suitable endogenous control miRNA in bone aging and senescence

ObjectiveMicroRNAs (miRNAs) are promising tools as biomarkers and therapeutic agents in various chronic diseases such as osteoporosis, cancers, type I and II diabetes, and cardiovascular diseases. Considering the rising interest in the regulatory role of miRNAs in bone metabolism, aging, and cellular senescence, accurate normalization of qPCR-based miRNA expression data using an optimal endogenous control becomes crucial. MethodsWe used a systematic approach to select candidate endogenous control miRNAs that exhibit high stability with aging from our miRNA sequence data and literature search. Validation of miRNA expression was performed using qPCR and their comprehensive stability was assessed using the RefFinder tool which is based on four statistical algorithms: GeNorm, NormFinder, BestKeeper, and comparative delta CT. The selected endogenous control was then validated for its stability in mice and human bone tissues, and in bone marrow stromal cells (BMSCs) following induction of senescence and senolytic treatment. Finally, the utility of selected endogenous control versus U6 was tested by using each as a normalizer to measure the expression of miR-34a, a miRNA known to increase with age and senescence. ResultsOur results show that Let-7f did not change across the groups with aging, senescence or senolytic treatment, and was the most stable miRNA, whereas U6 was the least stable. Moreover, using Let-7f as a normalizer resulted in significantly increased expression of miR-34a with aging and senescence and decreased expression following senolytic treatment. However, the expression pattern for miR-34a reversed for each of these conditions when U6 was used as a normalizer. ConclusionsWe show that optimal endogenous control miRNAs, such as Let-7f, are essential for accurate normalization of miRNA expression data to increase the reliability of results and prevent misinterpretation. Moreover, we present a systematic strategy that is transferrable and can easily be used to identify endogenous control miRNAs in other biological systems and conditions.

cell biology↗

bioRxiv2021

A New Gene Set Identifies Senescent Cells and Predicts Senescence-Associated Pathways Across Tissues

Although cellular senescence is increasingly recognized as driving multiple age-related co-morbidities through the senescence-associated secretory phenotype (SASP), in vivo senescent cell identification, particularly in bulk or single cell RNA-sequencing (scRNA-seq) data remains challenging. Here, we generated a novel gene set (SenMayo) and first validated its enrichment in bone biopsies from two aged human cohorts. SenMayo also identified senescent cells in aged murine brain tissue, demonstrating applicability across tissues and species. For direct validation, we demonstrated significant reductions in SenMayo in bone following genetic clearance of senescent cells in mice, with similar findings in adipose tissue from humans in a pilot study of pharmacological senescent cell clearance. In direct comparisons, SenMayo outperformed all six existing senescence/SASP gene sets in identifying senescent cells across tissues and in demonstrating responses to senescent cell clearance. We next used SenMayo to identify senescent hematopoietic or mesenchymal cells at the single cell level from publicly available human and murine bone marrow/bone scRNA-seq data and identified monocytic and osteolineage cells, respectively, as showing the highest levels of senescence/SASP genes. Using pseudotime and cellular communication patterns, we found senescent hematopoietic and mesenchymal cells communicated with other cells through common pathways, including the Macrophage Migration Inhibitory Factor (MIF) pathway, which has been implicated not only in inflammation but also in immune evasion, an important property of senescent cells. Thus, SenMayo identifies senescent cells across tissues and species with high fidelity. Moreover, using this senescence panel, we were able to characterize senescent cells at the single cell level and identify key intercellular signaling pathways associated with these cells, which may be particularly useful for evolving efforts to map senescent cells (e.g., SenNet). In addition, SenMayo represents a potentially clinically applicable panel for monitoring senescent cell burden with aging and other conditions as well as in studies of senolytic drugs.

cell biology↗

bioRxiv2021

Adipocyte-Specific Ablation Of PU.1 Promotes Energy Expenditure and Ameliorates Metabolic Syndrome In Aging Mice

ObjectiveAlthough PU.1/Spi1 is known as a master regulator for macrophage development and function, we have reported previously that it is also expressed in adipocytes and is transcriptionally induced in obesity. Here, we investigated the role of adipocyte PU.1 in the development of age-associated metabolic syndrome. MethodsWe generated mice with adipocyte specific PU.1 knockout, assessed metabolic changes in young and aged PU.1fl/fl (control) and AdipoqCre PU.1fl/fl(aPU.1KO) mice, including body weight, body composition, energy expenditure and glucose homeostasis. We also performed transcriptional analyses using RNA-Sequencing of adipocytes from these mice. ResultsaPU.1KO mice have elevated energy expenditure at a young age and decreased adiposity and increased insulin sensitivity in later life. Corroborating these observations, transcriptional network analysis indicated the existence of validated, aPU.1-modulated regulatory hubs that direct inflammatory and thermogenic gene expression programs. ConclusionsOur data provide evidence for a previously uncharacterized role of PU.1 in the development of age-associated obesity and insulin resistance.

cell biology↗

bioRxiv2021

Modulation of fracture healing by the transient accumulation of senescent cells

Senescent cells have detrimental effects across tissues with aging but may have beneficial effects on tissue repair, specifically on skin wound healing. However, the potential role of senescent cells in fracture healing has not been defined. Here, we performed an in silico analysis of public mRNAseq data and found that senescence and senescence-associated secretory phenotype (SASP) markers increased during fracture healing. We next directly established that the expression of senescence biomarkers increased markedly during murine fracture healing. We also identified a subset of cells in the fracture callus that displayed hallmarks of senescence, including distension of satellite heterochromatin and telomeric DNA damage. Then, using a genetic mouse model (p16LUC) containing a pl6Ink4a-dnven luciferase reporter, we demonstrated transient in vivo senescent cell accumulation during callus formation. Finally, we intermittently treated young adult mice following fracture with drugs that selectively eliminate senescent cells ("senolytics", Dasatinib plus Quercetin), and showed that this regimen both decreased senescence and SASP markers in the fracture callus and significantly accelerated the time course of fracture healing. Our findings thus demonstrate that senescent cells accumulate transiently in the murine fracture callus and, in contrast to the skin, their clearance does not impair but rather may improve fracture healing.

physiology↗