Biology subjects

Singh, S.

Publications and source records attributed to Singh, S..

At least 73 records · Page 4Linked to original sources

bioRxiv2026

Disruption of PI3K/OxPhos Coupling by Trehalose Drives a BCAA to Lipid Metabolic Switch in Hormone Receptor Positive Breast Cancer

Cancer cells sustain proliferation through dynamic coordination between mitochondrial oxidative phosphorylation (OxPhos) and anabolic carbon metabolism. How this metabolic coupling can be selectively destabilized in subtype-specific contexts remains poorly defined. Here we identify trehalose, a disaccharide previously linked to autophagy modulation, as a regulator of mitochondrial-anabolic integration in breast cancer. Using high-resolution respirometry, untargeted metabolomics, and signalling analyses across estrogen/progesterone receptor-positive (ER) and triple-negative models, we show that trehalose preferentially impairs mitochondrial bioenergetics in OxPhos-dependent ER cells. Trehalose reduced electron transport system capacity, NADH-linked respiration, mitochondrial membrane potential, and coupling efficiency, while suppressing mitochondrial biogenesis markers. These bioenergetic effects coincided with attenuation of PI3K/Akt signalling and induction of p21-associated growth arrest. Metabolomic profiling revealed a coordinated redistribution of carbon flux characterized by depletion of branched-chain amino acids (BCAAs) and glycolytic intermediates alongside accumulation of long-chain fatty acids and cholesterol. Correlation network analysis uncovered a strong inverse relationship between BCAA-linked metabolism and lipid abundance, indicating a regulated metabolic trade-off rather than nonspecific stress. Functionally, trehalose enhanced the efficacy of mitochondrial-interfering agents such as tamoxifen and colchicine, while exerting minimal effects in metabolically flexible triple-negative cells. Together, these findings define trehalose as a metabolic modulator that constrains mitochondrial plasticity and enforces a lipid-buffered, growth-restrictive state in ER breast cancer, revealing a therapeutic vulnerability linked to mitochondrial dependency.

cancer biology↗

bioRxiv2026

CGRP receptor-expressing neurons in the central amygdala contributes to injury-induced pain hypersensitivity

The central nucleus of amygdala (CeA) comprises diverse populations of neurons, forming a complex network responsible for regulating various behavioral responses. Among these, neurons expressing calcitonin gene-related peptide receptors (CGRPR) have emerged as key players in CGRP neuropeptide-mediated pain modulation. While previous studies emphasize CGRPs key role in synaptic plasticity and its connection with pain behavior in the CeA, the precise functional attributes and contributions of CeA-CGRPR-expressing neurons in pain processing remain elusive. This study reveals the co-localization of CGRPR-expressing neurons in the CeA with phosphorylated extracellular signal-regulated kinase (pERK), a marker indicating pain plasticity, in a neuropathic pain model. Electrophysiological assessments of these neurons in slice preparations unveiled heightened intrinsic excitability after sciatic nerve cuff implantation, contingent upon their rostro-caudal positioning within the CeA. Furthermore, our behavioral experiments using chemogenetic inhibition of CeA-CGRPR neurons demonstrated the ability to reverse nerve injury-induced hypersensitivity. Conversely, activating these neurons induced pain-related hypersensitivity even in the absence of injury. Our findings also highlight a sex-specific role of CeA-CGRPR neurons in formalin-induced spontaneous pain response. Collectively, these data reinforce the involvement of CeA-CGRPR neurons in pain processing, contributing to a better understanding of how neural circuits are affected in persistent pain conditions. Significance StatementOur study shows the role of CGRPR-expressing neurons within the CeA during pain processing. Using a cuff-implanted neuropathic mouse model, we discovered that CGRPR-expressing neurons co-localize with phosphorylated extracellular signal-regulated kinase (pERK), a hallmark of pain plasticity, in both male and female mice. Furthermore, our electrophysiological investigations reveal that posterior CeA-CGRPR neurons exhibit increased excitability following sciatic nerve cuff implantation. Importantly, we demonstrate that CeA-CGRPR neurons exert bidirectional effects on pain behavior in mice, irrespective of sex differences in nerve injury-induced pain responses while showing sex-specific spontaneous pain responses in the formalin-induced model. These findings show the role of CeA-CGRPR neurons in pain modulation, underscoring their potential significance in understanding and addressing persistent pain conditions.

neuroscience↗

bioRxiv2026

A composite frailty index enables quantification of functional aging and identification of gerotherapeutic drugs in the house cricket.

Frailty, defined by progressive loss of physiological resilience, neuromuscular function, and cognitive capacity, is a central manifestation of biological aging yet remains difficult to quantify in scalable experimental systems. Here, we introduce a Composite Frailty Index (CFI) in the house cricket (Acheta domesticus) that integrates automated measures of locomotion, exploratory behavior, and freezing into a unified, quantitative framework of functional decline. Ten behavioral parameters derived from automated open-field tracking, including locomotor performance, exploratory behavior, and freezing were integrated into the CFI. Locomotor states were classified using k-means clustering (k = 2) of velocity distributions, and all features were normalized to age- or treatment-matched reference populations, discretized into quintiles, and summed to generate a 0-40 frailty score. Aging cohorts (young adult: 4-6 weeks; geriatric: 10-12 weeks, N = 103) and pharmacological cohorts treated at mid-life (8-10 weeks) with rapamycin (14 ppm), acarbose (1000 ppm), or phenylbutyrate (1000 ppm) were evaluated (N = 122). Across chronological aging cohorts, CFI increased from young adults to geriatrics in both females (d = 1.14 [95% CI: 0.53, 1.76], P = 0.0003) and males (d = -1.17 [95% CI: -1.75 to -0.59], P < 0.0001). Using pharmacological intervention cohorts, mid-life rapamycin treatment reduced late-life frailty relative to controls in both females (d = -1.31 [95% CI: -2.09, -0.53], P = 0.0017) and males (d = -1.33 [95% CI: -2.09, -0.58], P = 0.0004), whereas acarbose and phenylbutyrate produced inconclusive effects (ds = -0.54 to -0.03; Ps > 0.05). Together, these findings establish the cricket CFI as a scalable, high-throughput platform for quantifying multidimensional functional aging and prioritizing candidate geroprotective interventions based on clinically relevant endpoints beyond lifespan.

animal behavior and cognition↗

bioRxiv2026

Myeloid Gi signaling acts as a weight-independent immunometabolic switch controlling systemic insulin sensitivity

Metabolic dysfunction does not necessarily correlate with adiposity. Metabolically healthy obese individuals and insulin-resistant lean individuals represent a fundamental paradox that implicates immune cell intrinsic mechanisms in the pathogenesis of type 2 diabetes. Here, we identify myeloid Gi signaling as a previously unrecognized determinant of whole-body glucose homeostasis. Single-cell transcriptomic analysis of adipose tissue macrophages from obese mice and humans reveals marked alteration in Gnai isoform, suggesting that myeloid Gi signaling is functionally engaged during metabolic disease. Using complementary myeloid-specific rodent models of Gi inhibition (pertussis toxin) and chemogenetic Gi activation (DREADD), we demonstrate that inhibition of Gi signaling improves glucose tolerance and enhances insulin sensitivity under both regular chow and high-fat diet conditions, independent of body weight and energy expenditure. Whereas acute Gi activation in lean mice modestly enhances glucose disposal, the same intervention during diet-induced obesity markedly impairs systemic glucose homeostasis, revealing context-dependent pathway function. Mechanistically, Gi inhibition amplifies macrophage cAMP-CREB signaling to drive IL-6 production, engaging STAT3- and AMPK-dependent pathways in adipose tissue and skeletal muscle to support insulin action. Conversely, Gi activation engages a previously uncharacterized G{beta}{gamma}-mTOR/AKT-JNK cascade, driving IL-1{beta} secretion that directly impairs insulin signaling in adipocytes and myotubes. Pharmacological IL-6 receptor blockade abolishes the metabolic benefits of Gi inhibition, whereas IL-1 receptor antagonism fully rescues Gi activation-induced metabolic dysfunction, establishing these cytokines as obligate downstream effectors. This signaling architecture is conserved in human macrophages, and ATAC-seq profiling reveals chromatin remodeling at cAMP-CREB and IL-6 regulatory pathway loci, consistent with the observed transcriptional reprogramming. Together, these findings establish myeloid Gi signaling as a weight-independent immunometabolic switch that couples opposing cytokine programs to systemic insulin sensitivity and identify this pathway as a therapeutic target in obesity-associated metabolic disease. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=141 SRC="FIGDIR/small/713834v1_ufig1.gif" ALT="Figure 1"> View larger version (56K): org.highwire.dtl.DTLVardef@15e4576org.highwire.dtl.DTLVardef@224cd5org.highwire.dtl.DTLVardef@1e24888org.highwire.dtl.DTLVardef@1080bf3_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG

physiology↗

bioRxiv2026

Sphingosine-1-Phosphate Receptor 1 regulates competition dependent astrocyte morphogenesis and tiling in murine cortex.

Astrocytes are highly abundant in the mammalian brain and coordinate with neurons and other glial cells to regulate neural circuit structure, function, and blood brain barrier integrity among many to maintain proper brain homeostasis. Astrocytes perform most of these functions owing to their highly complex morphologies and hundreds of thousands of fine processes that are important in contacting neuronal synapses and other glial cells. In fact, the morphological complexity of astrocytes is regulated by the presence and activity of neurons and helps establish astrocyte territory/tiling in a non-overlapping pattern; however, the mechanisms of astrocyte tiling are not well characterized. Using a human astrocyte-mouse neuron coculture system, we previously showed that sphingosine-1-phosphate receptor 1 (S1PR1) regulates astrocyte morphogenesis in a neuronal contact dependent manner. In this study, we find that S1PR1, in vivo, regulates astrocyte morphogenesis in a cortical layer specific manner. Using astrocyte-specific S1PR1 knock out mouse models and adenoassociated viral labeling methods, we show that S1PR1 is crucial in establishing competition driven astrocyte tiling and morphogenesis in the developing brain. Furthermore, we show that JAK-STAT3 signaling regulates neuronal contact induced expression of S1PR1 in cocultured astrocytes. These studies therefore uncover a lipid signaling receptor as a major regulator of astrocyte morphogenesis and tiling in murine cortical layers.

neuroscience↗

bioRxiv2026

Novel regimens for treatment of Mycobacterium avium lung disease based on advanced in vitro systems and the mathematics of basis functions

Azithromycin plus ethambutol plus rifabutin (azithromycin-ethambutol-rifabutin) is the standard-of-care (SOC) for Mycobacterium avium-complex lung disease. The SOC achieves sustained sputum culture conversion in only 43-53% of patients, after an average of 18 months of therapy. Recent quantitative analyses ranked omadacycline, ceftriaxone, and minocycline highest for microbial kill. Azithromycin-minocycline-ethambutol, azithromycin-omadacycline-ethambutol, epetraborole-omadacycline-ethambutol, ceftriaxone-omadacycline-rifabutin, and the SOC were compared in the intracellular hollow fiber system model of M. avium lung disease (HFS-MAC). HFS-MAC units were treated once daily for 28 days to mimic the intrapulmonary pharmacokinetics of each drug. The ceftriaxone concentrations measured in the HFS-MAC were only 1% of those achieved in the lung by standard clinical doses. Changes in the bacterial burden were described using basis functions (BF). For liquid cultures, BF 1 (BF1) was described by a linear regression-based slope, with steepest kill slope (95% Confidence interval) of 7.87 (1.52 to14.23) by ceftriaxone-omadacycline-rifabutin versus 1.04 (-0.84 to 2.92) for SOC. For the CFU/mL readout, the BF1 steepest non-linear kill slope was for ceftriaxone-omadacycline-rifabutin of 0.55 (0.35 to 0.98) log10 CFU/mL/day versus 0.16 (0.07 to 0.25) log10 CFU/mL/day for the SOC. Thus, ceftriaxone-omadacycline-rifabutin is potentially better than the SOC, even though further ceftriaxone dose optimization is required. BF2 described rebound growth and drug-resistant subpopulation growth, and demonstrated that contrary to popular belief, SOC rebound was best explained by ethambutol-resistance (r2>0.99, p=0.01) and not by azithromycin-resistance (r2=0.27, p=0.32), questioning ethambutols role in the SOC. The BF framework is potentially easy to adapt for modeling other anti-infective agents across many infectious diseases.

microbiology↗

bioRxiv2026

Ribosome Processing Factor-2 Interacts with RPL10A to Regulate Selective Translation during Plant Immunity and Drought Stress

Processing of ribosomal RNA (rRNA) is essential for ribosome biogenesis, translation, plant development, and stress adaptation. Ribosome processing factor-2 (RPF2), which plays a role in the later stages of rRNA maturation, interacts with ribosomal protein L10A (RPL10A). RPF2 overexpression in Arabidopsis and Nicotiana benthamiana showed enhanced plant growth and trichome development due to increased gibberellic acid (GA) levels. Conversely, RPF2-silenced and mutant plants had a dwarf phenotype, reduced stomatal apertures, and decreased glucosinolate accumulation. RPF2 silenced and mutant plants also showed compromised nonhost disease resistance, whereas RPF2 overexpression lines exhibited enhanced disease resistance to both host and nonhost pathogens. RPL10A and RPF2 overexpression lines were sensitive to abscisic acid (ABA) and tolerant to drought, which is attributed to their unique roles in translation regulation. Despite having larger stomatal apertures, RPF2 overexpression plants displayed low pathogen multiplication rates and reduced water loss, indicating independent resistance mechanisms associated with ribosomal functions in translation regulation. Although both RPL10A and RPF2 proteins interact with each other and are involved in translation regulation, proteomic analysis suggests that they regulate the translation of distinct sets of genes during pathogen or drought stress. These findings indicate that RPF2 and RPL10A play independent roles in the regulation of unique protein translation.

plant biology↗

bioRxiv2026

Biobank-scale genotyping of Robertsonian translocations reveals hidden structural variation on the human acrocentric chromosomes

Balanced Robertsonian translocation (ROB) is the most common chromosomal rearrangement in humans, with an estimated occurrence of 1 in 800 in newborn studies. Carriers are at increased risk of cancer and often diagnosed at fertility clinics after facing recurrent miscarriages, infertility, or aneuploid offspring. Genotyping carriers with DNA sequencing has been challenging because of gaps and misrepresentation of the translocation fusion site in the human reference genome. Only recently, telomere-to-telomere (T2T) human genomes successfully revealed sequences of the acrocentric short arms, including the most common ROB fusion site. A ROB results in loss of two ribosomal DNA (rDNA) arrays and its adjacent distal sequences, including the highly conserved distal junction (DJ). Here, we present a novel method to type ROB carriers directly from short sequencing reads by estimating DJ copy number. We demonstrate that our method successfully genotypes ROBs using a reference-free approach or alignments to either T2T-CHM13v2 or GRCh38. Applying the method to a cohort of healthy newborns and family members (n=4,172) as well as the UK Biobank (n=490,416), we find candidate ROBs at a frequency consistent with the previously reported 1 in 800 incidence (0.11-0.12%). In addition to ROB carriers, we report the frequency of one DJ loss (9, 2.8-3.4%) or gain (11+, 8.4-9.3%) from the two cohorts and the 1000 Genomes Project (n=3,202), and characterize the underlying structural variation in near-T2T genome assemblies from the Human Pangenome Reference Consortium. Importantly, our method provides the first sequencing-based diagnostic for Robertsonian chromosomes and can be applied to low-coverage sequencing data, enhancing its clinical applicability and enabling new studies of structural variation on the acrocentric chromosomes.

genomics↗

bioRxiv2026

A Scalable Design for Proximity-Inducing Molecules

Chimeric molecules, which bring together an effector enzyme and a protein-of-interest (POI) to add/remove post-translational modifications (PTMs), are furnishing transformative modalities (e.g., PROTACs). However, these chimeras scalability is limited as they employ rare, non-inhibitory binders of effectors. We report GRoup-transfer chimeras for Inducing Proximity (GRIPs) that employ abundantly available effectors inhibitors to append POI binder on the effector using group-transfer handles. To demonstrate scalability, we develop 6 GRIPs classes for 3 PTMs utilizing diverse inhibitor, spanning 16 effector-POI pairs. Furthermore, we report a toolbox of 42 tunable group-transfer handles for Cys/Lys residues and [~]5000 inhibitor-residue pairs for diverse effectors. Using global proteomics, we confirm the specificity for group transfer and PTM editing. GRIPs endowed new functionalities to POI drugs, including preventing rebound signaling upon drug withdrawal, a more potent/persistent inhibition, and inhibitor-induced pathway activation in 4 fully-endogenous systems. In diverse hemi-endogenous systems (tagged POI), GRIPs induced condensate formation with reduced off-targets, cleared pathogenic PTMs, and initiated PTM crosstalk. Overall, GRIPs provide a scalable and versatile platform for PTM editing. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=184 SRC="FIGDIR/small/706349v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@132f70dorg.highwire.dtl.DTLVardef@f9aa62org.highwire.dtl.DTLVardef@12143edorg.highwire.dtl.DTLVardef@f296d4_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2026

Direct effector recognition by a tandem kinase triggers non-canonical immunity in wheat

Tandem kinase proteins (TKPs) are an important class of disease resistance genes in cereal crops alongside nucleotide-binding (NB-ARC) leucine-rich repeat (LRR) (NLR) receptors. Here, we cloned the wheat (Triticum aestivum) resistance gene Lr41, which encodes a TKP with a C-terminal fusion, and its paired effector gene AvrLr41 from the rust pathogen Puccinia triticina. AvrLr41 is unusually large and highly divergent from most fungal effectors and is directly recognized by Lr41 through its pseudokinase domain. Immune activation mediated by Lr41 require auxiliary helper proteins, including either a minimal coiled-coil (CC) helper protein or its full-length NLR ortholog. These findings reveal a non-canonical immune system in which a kinase-type receptor directly recognizes an effector and recruits either CC or NLR helpers, demonstrating that full-length NLRs are not strictly required for NLR-linked immune execution and expanding the functional repertoire of disease resistance proteins in crops.

plant biology↗

bioRxiv2026

Molecular Dynamics Analysis of Self and Microbial Peptides Bound to HLA-B27: A Multi-Parameter Framework

Molecular mimicry between pathogen-derived and self-peptides shown by MHC molecules is one of the critical mechanisms in the pathophysiology of autoimmune diseases. Numerous studied has been conducted in this field to identify sequence similarity, but evaluating structural and dynamic similarity, systematic computational frameworks remain limited. Therefore, we created an automated multi-parameter molecular dynamics analysis workflow and used it to compare three peptides (KP1, KP2, and KP3) generated from Klebsiella pneumoniae bound to HLA-B class protein with one human self-peptide (Annexin-derived, ANX). We assessed six complementing parameters using one microsecond-scale MD simulation: radius of gyration (Rg), solvent-accessible surface area (SASA), hydrogen bonding dynamics, MM-GBSA binding free energy, root mean square fluctuation (RMSF), and root mean square deviation (RMSD) to understand time-dependent structural and dynamic behaviour of all the peptide-HLA-B complex. Additionally, hydrogen bond occupancy and molecular mechanics generalised Born surface area (MM-GBSA) binding free energy calculations were performed to provide a more comprehensive assessment of complex stability. Our analysis suggests that KP1 exhibits structural features consistent with molecular mimicry, maintaining conformational stability, surface exposure, and interaction patterns comparable to ANX. In contrast, KP2 showed reduced stability, characterised by higher RMSD values and substantial hydrogen bond loss, whereas KP3 displayed intermediate behaviour, with relatively favourable energetics but noticeable conformational variability. Overall, the multi-parameter framework enabled differentiation among the candidate peptides based on combined structural, dynamic, and energetic properties. The workflow can be adapted for the analysis of larger peptide datasets and may provide a systematic approach for investigating potential autoimmune-relevant molecular mimics in microbial proteomes, with required adjustments according to the system.

immunology↗

bioRxiv2026

Evaluation of Significance of SERPINA3 and SPP1 in Driving Glioma Progression Through Dysregulation of Brain Inflammatory Pathway

Glioma is a highly aggressive malignancy with a poor prognosis, particularly in grade IV glioblastoma. The monitoring of disease progression remains challenging due to high heterogeneity in the tumour and a lack of progressive markers to keep track of the tumour progression. This scarcity of good progressive biomarkers has led us to search for better options. SERPINA3 and SPP1 were investigated as potential dual biomarkers reflecting tumour microenvironment and enabling assessment of progression. In silico analysis was conducted, where correlation analysis was performed to evaluate the cell-type specificity of SERPINA3 in astrocytes and SPP1 in microglial cells, with comparisons across other neural populations in both low-grade glioma and glioblastoma. Pan-cancer expression analysis was conducted to determine whether these biomarkers remain significantly elevated in glioma relative to other malignancies, including hepatocellular carcinoma, given their hepatic origin. Alzheimers disease datasets were analysed to verify the relevance in neurodegenerative diseases. The in-silico analysis revealed that SERPINA3 and SPP1 show astrocytic and microglial specificity, respectively, and exhibit their highest expression levels in glioma across cancers. Co-expression analysis further identified enrichment of immunoregulatory pathways alongside upregulated oxidative stress-associated markers, highlighting the functional relevance of these biomarkers within the glioma microenvironment.

neuroscience↗

bioRxiv2026

Targeting Galectin-3 C-epitope oligomers associated maladaptive mechanotransductive signaling in pressure-overload induced left ventricular cardiac hypertrophy

BackgroundAging and various pathological conditions lead to pressure-overload in the left ventricle, promoting maladaptive hypertrophic remodeling and subsequent cardiac dysfunction, ultimately increasing the risk of heart failure. Galectin-3 (Gal-3) plays a central role in this process; however, its critical intracellular functions complicate direct therapeutic targeting. Notably, pathological microenvironments trigger the proteolytic cleavage of Gal-3 into distinct N- and C-terminal fragments. The specific contributions of these cleaved epitope forms to adverse cardiomyocyte mechanotransduction, and their potential as precision therapeutic targets in contrast to the full-length protein, remain unresolved. MethodsTo address this gap, we combined rodent models of aging and pressure-overload (PO) -induced cardiac hypertrophy with PO mechanobiology-driven in vitro assays and validation in human cardiac tissue and serum. Gal-3 epitope abundance, localization, phosphorylation, oligomerization, and downstream signaling were quantified using biochemical, imaging, and functional approaches. ResultsWe found that extracellular oligomers of the Gal-3 C-terminal epitope accumulated in serum and on cardiomyocyte surfaces in hypertrophic rodents and human subjects, where they correlated with adverse remodeling and cardiomyocyte loss. Treatment with Amalaki Rasayana (AR), a standardized nutraceutical-based cardioprotective Ayurvedic phytomedicine, and its bioactive component gallic acid (GA) significantly reduced circulating and surface-associated Gal-3 C-epitope oligomers and attenuated hypertrophy-associated cytotoxic signaling. Mechanistically, AR/GA enhanced Ser6 phosphorylation of Gal-3, promoting intracellular retention, while limiting pathological secretion and deleterious extracellular oligomerization. Following AR/GA treatment, the binding of preformed Gal-3 C-epitope oligomers to cardiomyocyte surfaces were further inhibited, thereby suppressing maladaptive mechanotransductive signaling. Importantly, circulating Gal-3 C-epitope oligomers, together with atrial natriuretic peptide (ANP), constituted a drug-responsive biomarker panel that accurately tracked hypertrophy regression, serving as an indicator of drug efficacy. ConclusionsIn summary, Gal-3 C-epitope oligomers represent pathogenic signaling, drug-responsive therapeutic targets and circulating biomarkers of cardiac hypertrophy, with broader relevance to other Gal-3-driven neoplastic, fibrotic, and inflammatory diseases. HighlightsO_LIGalectin-3 C-epitope forms pathogenic extracellular oligomers in pressure-overload induced cardiac hypertrophy. C_LIO_LISurface binding of excess C-epitope oligomers triggers an adverse mechanotransductive remodelling in cardiomyocytes. C_LIO_LIPhytomedicine -Amalaki Rasayana (AR) and its key bioactive compound, gallic acid (GA), effectively inhibit the surface binding and mitigate harmful effects of Gal-3 C-epitope oligomers. C_LIO_LIAR/GA induces the phosphorylation of Gal-3, limiting pathological secretion, extracellular oligomer assembly, and glycan-mediated surface binding. C_LIO_LIIntracellular retention of Gal-3 preserves its crucial cellular functions. C_LIO_LICirculating levels of Gal-3 C-epitope oligomers can be used to monitor the therapeutic regression of cardiac hypertrophy, serving as an indicator of drug efficacy C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=164 HEIGHT=200 SRC="FIGDIR/small/704762v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@4702e9org.highwire.dtl.DTLVardef@10361eorg.highwire.dtl.DTLVardef@51118eorg.highwire.dtl.DTLVardef@57ba37_HPS_FORMAT_FIGEXP M_FIG C_FIG Pressure-overload induces excessive secretion of Galectin-3 and triggers its proteolytic cleavage into the N- and C-terminal epitopes. The extracellular oligomerization of the C-terminal epitope facilitates high-affinity glycan binding and promotes pathological mechanotransductive signaling in cardiomyocytes. Phytochemical modulation by Amalaki rasayana and its bioactive component, gallic acid, induces the phosphorylation of Gal-3, limiting pathological secretion, extracellular oligomer assembly, and glycan-mediated surface binding, while preserving intracellular function. Circulating levels of Gal-3 C-epitope oligomers can be used to monitor the therapeutic regression of cardiac hypertrophy, serving as an indicator of drug efficacy.

cell biology↗

bioRxiv2026

Higher-order intrinsic routes support flexible task-evoked communication

Flexible task-evoked communication requires the brain to transiently establish functional interactions that are not evident during rest, yet the network mechanisms supporting these newly formed connections remain unresolved. Using whole-brain functional MRI from 92 participants performing six cognitive tasks, we show that task-evoked communication is implemented within a constrained intrinsic network architecture rather than through large-scale rewiring. Across tasks, we found that approximately 70% of functional connections were preserved from rest, forming a dominant stable core, while task engagement selectively reconfigured a smaller subset of connections, primarily at the between-network level. While existing activity-flow models explain task-evoked information transfer primarily through direct stable core connections, we demonstrate that newly formed connections between regions lacking direct resting-state coupling were supported by selectively strengthened indirect higher-order resting-state routes embedded within the intrinsic scaffold. Network-level analyses revealed a multiscale organization of information transfer: we observed that direct stable routes dominated within-network communication, most prominently in sensory-motor and default mode systems, whereas indirect higher-order routes preferentially supported between-network integration through control and attentional systems. By extending activity-flow modelling to incorporate these higher-order intrinsic routes, we show a significant improvement in the prediction of task-evoked activity at the global, between-network scale, confirming their functional recruitment during task performance. Together, our findings identify higher-order intrinsic routes as a key mechanism enabling flexible task-evoked communication within a stable large-scale network architecture.

neuroscience↗

bioRxiv2026

Cytokinin Senescence Delay Is Shaped by Receptor Specificity and Metabolic Stability

Each of the four different cytokinin (CK) base forms, trans Zeatin (tZ), isopentenyladenine (iP), dihydrozeatin (DHZ), and cis Zeatin (cZ) have distinct chemical metabolism and affinity to the CK Histidine Kinase (CHK) receptors. However, it remains unclear how specific biochemical features of each form such receptor specificity or metabolic differences drives distinct tissue-specific physiological hormone output in response to application of these CK bases. Here, we show that CK receptor preference and metabolic persistence together shape isoform-specific CK signaling strength, including tissue-dependent hormone responses in Arabidopsis leaf versus root assays. Physiological, genetic, and multi-omics integration was used to show that tZ and iP anti senescence activity is matched by DHZ through a distinct receptor metabolic mechanism. DHZ requires Arabidopsis Histidine Kinase 3 (AHK3) signaling to be fully effective in a leaf Dark Induced Senescence (DIS) assay and where it overcomes its lower receptor affinity through higher metabolic persistence, accumulating at levels [~]2.5-fold above tZ and iP early in a senescence time course. Together, these findings provide a framework for integration of receptor preference and metabolic stability to determine CK isoform activity.

plant biology↗

bioRxiv2025

Correlated dynamics, reactive conformations and non-Arrhenius behaviour in the temperature-dependence of enzyme activity: triosephosphate isomerase

Heat capacity changes are increasingly recognised as crucial for understanding the temperature dependence of enzyme-catalysed reaction rates. Here, we combine experiments and molecular dynamics simulations to investigate triosephosphate isomerase (TIM, TPI) from psychrophilic, mesophilic and thermophilic organisms. Kinetic data show clear curvature in rate-temperature plots, particularly for the cold-adapted enzyme, independent of unfolding. This non-Arrhenius behaviour is accounted for by Macromolecular Rate Theory (MMRT), implying an activation heat capacity, largest for the psychrophile. Simulations reveal the molecular origins of these differences, showing significant conformational and dynamical changes between reactant and transition states in the cold-adapted enzyme, with a smaller activation heat capacity in the mesophilic enzyme, and near-zero for the thermophile. Transition-state-like conformations and reorganized correlated dynamics underlie these adaptations. These results demonstrate that dynamical differences between crucial states along the reaction pathway play a key role in determining the optimum temperature of activity for this archetypal enzyme.

biochemistry↗

bioRxiv2025

Senescent CD8+ T Effector Memory Cells are Functionally Impaired, Enriched in Aging and Disease, and a Barrier to Immunotherapy

Senescent cells play important roles in various biological processes that promote fitness and health, however, their timely elimination by immune cells is critical to maintain tissue homeostasis and prevent disease. Despite this, senescent cells progressively accumulate systemically with age, suggesting that certain immune cells also become senescent and dysfunctional during aging. Supporting this, we previously demonstrated that CD8 T cells, immune cells capable of targeting senescent cells, increasingly develop characteristics of senescence with advancing age in humans. Here, we further characterized the senescence state of human SA-{beta}Gal-expressing CD8 T effector cells, their functional capabilities, and their involvement in aging and disease. Single-cell RNA sequencing revealed that SA-{beta}Gal-expressing CD8 T cells with unique transcriptional signatures develop in all stages of T cell differentiation, including in effector memory (EM) T cells. SA-{beta}Gal-expressing CD8 TEM cells expressed various classical markers of senescence and were significantly impaired in their ability to proliferate, produce cytokines, and eliminate senescent human stromal cells, compared to CD8 TEM cells with low SA-{beta}Gal activity. Gene signatures of senescent SA-{beta}Gal-expressing CD8 TEM cells were enriched in CD8 T cells from older human donors, patients with age-related disorders, cancer, and smokers. Furthermore, our results demonstrate that T cell senescence is distinct from and dominant over T cell exhaustion, limiting the response of CD8 TEM cells to immunotherapy. Collectively, our study demonstrates that the senescence state impairs the functions of CD8 TEM cells and reveals the involvement of senescent and dysfunctional CD8 TEM cells in aging, disease, exposure to toxins, and responses to immunotherapy.

cell biology↗

bioRxiv2025

Activation of cannabinoid receptor CB1 leads to aberrant myelination in development

The endocannabinoid system (ECS) has a widespread role in the development and function of the central nervous system (CNS). Cannabinoid receptors like CB1 and CB2 can be activated with exogenous cannabinoids most popularly known as tetrahydrocannabinol (THC) or cannabis and cannabidiol (CBD). The components of the ECS are expressed early in fetal development, and prenatal exposure to cannabis can lead to structural changes in white matter. White matter is composed of neuronal axons ensheathed in myelin, a lipid-rich insulation that facilitates saltatory conduction and maintains axon integrity. In the CNS, myelin is made by specialized glial cells called oligodendrocytes (OLs), which in addition to neurons also express components of the ECS. However, while several studies have focused on how the ECS regulates neuronal development, there is a limited understanding of its impact on OL development or myelin formation. Therefore, our current study set out to understand how pharmacological activation of the ECS alters OL differentiation and myelin formation in vivo. We administered WIN 55,212-2 (WIN 55), a CB1 and CB2 agonist, to larval zebrafish and longitudinally analyzed OL development and myelination in vivo. Interestingly, we observed an increase in non-axonal ensheathments in the spinal cord, which appeared to be surrounding neuronal cell bodies. These non-axonal ensheathments were dependent on CB1, as the addition of WIN 55 in a global CB1 mutant prevented this phenotype. Furthermore, this ectopic cell body ensheathment occurred independently from normal myelination processes, as individual OLs did not exhibit changes in the number of myelin sheaths, sheath length, or total myelin output. This study shows that activation of CB receptors in vivo leads to increased non-axonal ensheathment without significantly changing OL differentiation or normal myelin formation. Future studies can further investigate the pathways that drive this phenotype to better understand how exogenous cannabinoid activation can regulate the precision of oligodendrocyte ensheathment.

neuroscience↗