Biology subjects

Roy, S.

Publications and source records attributed to Roy, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2026

Neutrophil Extracellular Trap Formation and Complement Activation Pathways Dominate Microbiota-dependent disease bias in lupus-prone female NZM2328 mice

Women are predisposed to systemic lupus erythematosus (SLE) with a prevalence ratio of up to 9:1 over men. Multiple mouse strains including NZM2328 exhibit strong female dominance in developing spontaneous lupus as in humans with SLE. While lupus-prone mice can develop disease under germ free (GF) condition, the role of gut microbiota in female bias for lupus nephritis is not investigated systematically. Here, using specific pathogen free (SPF) and GF NZM2328 mice, and employing microbiota-depletion and microbial-association strategies, we show that microbiota influences lupus-like disease outcomes differently in males and females. Female NZM2328 mice with intact microbiota presents higher inflammation factor expression, including X-chromosome linked TLRs, in the distal gut and systemic compartments, and higher activation of genes and biological pathways such as neutrophil extracellular trap (NET) formation and complement and coagulation cascade (CCC) pathways, associating with their higher disease susceptibility. Gut microbiota-depletion as well as GF derivation eliminated not only the modest differences in the serum and fecal antibody levels and nAg reactivity, but also the gender bias in the timing of clinical stage disease onset as well as systemic NET and CCC pathway activation. Reciprocally, conventionalization of GF NZM2328 mice at juvenile age restored the female bias in intestinal and systemic autoantibody levels, pro-inflammatory immune pathway activation, and the timing of clinical stage disease onset. Overall, our observations show that, while genetic susceptibility appears to be the cause of lupus-like disease in NZM2328 mice, differential activation of NET and CCC pathways in males and females upon exposure to gut microbes, in combination with host-factors, causes gender bias in disease outcomes. We conclude that microbiota exposure-dependent protection of males and overactivation of NET and CCC pathways in females could be contributing to the female bias in lupus-like disease in NZM2328 mice.

immunology↗

bioRxiv2026

ESCRT-0 regulates AMPA receptor currents and Ca2+- dependent signaling

Membrane protein trafficking is essential for synaptic growth, maintenance, function, and plasticity, and involves the regulated exocytosis and endocytosis of proteins to and from the pre-and post-synaptic membranes. Defects in the clearance of membrane proteins can lead to the accumulation of ubiquitinated membrane proteins and contribute to neurodegenerative disease. The ESCRT (endosomal sorting complexes required for transport) machinery binds and sorts ubiquitinated membrane proteins into lysosomes for degradation, yet the presence and function of ESCRTs in sorting ubiquitinated AMPA and other receptors at the post-synapse remain unclear. Here we show that the ubiquitin-binding ESCRT-0 protein, Hrs, localizes to both pre- and post-synapses, and levels are modulated by neuronal activity, increasing and decreasing with higher and lower neuronal activity, respectively. Phosphoproteomic profiling of Hrs-depleted post-synaptic membranes revealed a role for Hrs in glutamatergic synaptic transmission, including long-term potentiation. In addition, Hrs-depleted neurons showed faster AMPAR current kinetics and reduced amplitude in whole-cell patch-clamp recordings. Genetic deletion of neuronal Hgs in mice led to reductions in phosphorylated CaMKII- and -{beta} (T286/T287) and structural proteins, PSD-95 and gephyrin, suggestive of LTD (long-term depression)-like synaptic depression. In contrast, Hrs overexpression led to increases in Ca2+-dependent signaling, including protein kinase C (PKC) and PKC substrate, AMPAR subunit GluA1-S831, a site which increases conductance. Together, these findings identify a dynamic, bidirectional role for Hrs at the post-synapse as it both senses and is modulated by neuronal activity, ultimately impacting excitatory synaptic strength. Significance StatementSynaptic plasticity relies on dynamic trafficking and turnover of membrane proteins, including AMPA-type glutamate receptors (AMPARs), yet how receptor trafficking intersects with ubiquitin-mediated sorting pathways at synapses remains unclear. We show that the ubiquitin-binding ESCRT-0 protein, Hrs, localizes to both pre- and post-synapses, and its abundance is bidirectionally regulated by neuronal activity. Genetic depletion of Hrs in mice reduces CaMKII phosphorylation and impacts AMPAR channel surface localization. In contrast, neuronal-specific Hrs overexpression led to enhanced GluA1 and protein kinase C substrate phosphorylation, suggesting altered AMPAR trafficking, subunit composition, and/or function. Thus, Hrs emerges as a modulator of glutamatergic signaling, coupling ubiquitin-mediated receptor sorting to the fine-tuning of synaptic transmission, with direct implications for learning and memory in health and disease.

neuroscience↗

bioRxiv2026

Single-cell, single-shot stimulation reveals heterogeneous network recruitment not explained by standard functional properties.

Understanding how minimal perturbations influence cortical network dynamics remains a central challenge in neural engineering. While single-cell stimulation has been shown to affect population activity, the resulting variability is often treated as noise rather than an informative feature of network behavior. Here, we investigated how single-cell stimulation reflects heterogeneous modulation of neuronal recruitment and the extent to which these effects can be explained by the functional state of the stimulated cell. For this, we combined single-cell optogenetic stimulation with wide-field calcium imaging in cortical cultures. In each network, a single stimulation event was induced, and subsequent alterations in stimulus-coupled recruitment, synchrony, and pairwise correlations were quantified. Additionally, we evaluated whether the baseline functional state of the stimulated neurons, including their event activity levels and Pearson correlation structures, were linked to the observed network responses. Single cell stimulation induced effects were transient, and the network dynamics recovered over a few seconds within the responder population. Importantly, Our findings demonstrate that the observed direction and magnitude of recruitment changes were not significantly explained by the functional state of the stimulated neurons, indicating that these parameters do not capture the determinants of perturbation-induced network responses. This highlights a possibilities in the future approaches for characterizing network responsiveness and suggests that additional unobserved features govern the response of microcircuits to localized inputs.

neuroscience↗

bioRxiv2026

Prediction of fMRI activity using vector autoregressive models: a comparison of sparse and low-rank approaches

Vector autoregressive (VAR) models have a history of being used to examine functional connectivity in the brain, as captured by functional MRI studies. Such models allow for an estimation of Granger-causal relationships between regions of interest across the brain. Unfortunately, since the number of parameters in the VAR model scales as the square of the number of regions, and this is typically large compared to the number of temporal observations, these parameter estimates will exhibit high variance. To address this challenge, we introduce a low-rank pre-smoothing method that applies a low-rank approximation to the observations before fitting a VAR model. We estimate these models using individual subject data from both task-based and resting-state conditions, tuning hyperparameters at the population level. Our low-rank approach is directly compared against sparse and unconstrained estimation methods. Evaluations of predictive performance and model structure reveal that our pre-smoothing technique enables robust individual-level parameter estimation and significantly reduces prediction error, a finding further validated by synthetic experiments where the ground-truth parameters are known.

neuroscience↗

bioRxiv2026

Matched pancreatic cancer liver metastatic model system reveals cancer cell-dependent organotropism and site-specific tumor microenvironment reflective of human disease

Background: Pancreatic ductal adenocarcinoma (PDAC) mortality is driven largely by liver metastatic disease; however, the malignant cell states and tumor microenvironmental features of PDAC liver metastases remain obscure. Methods: We developed a transplant model system of matched pancreatic and liver tumors to study PDAC metastatic progression. Using this model, we identified murine PDAC cell lines with distinct liver metastatic capacities and performed multiomic profiling of matched primary pancreatic and metastatic liver tumors. Transcriptional programs associated with high and low liver tropism were defined and evaluated across tumor models and independent human PDAC datasets. Spatial and tumor-immune interaction analyses were used to characterize microenvironmental niches, immune composition, and cellular relationships within primary and metastatic tumors. Results: A high-liver-tropic transcriptional program was enriched in high liver-tropic cell lines and malignant cells within liver metastases, conserved across human PDAC datasets, and associated with inferior patient survival. High- and low-liver-tropic tumor states occupied distinct liver microenvironmental niches and exhibited different tumor-immune communication networks, accompanied by local and systemic changes in immune composition. Liver metastases also displayed features of enhanced immunosuppression, including increased proximity of CD4+ and CD8+ T cells to tumor cells and greater spatial association between regulatory T cells and exhausted CD8+ T cells. Conclusions: These findings identify conserved PDAC cell states associated with differential liver metastatic capacity and demonstrate that liver metastasis is accompanied by spatial and immunologic remodeling of the tumor microenvironment. Together, the study provides a framework for understanding how tumor-intrinsic metastatic programs interact with site-specific immune ecosystems in PDAC.

cancer biology↗

bioRxiv2026

Interplay of Cullin5-HECTD3 ubiquitin ligases regulates stability of CRAF mutant associated with hypertrophy

Proteostatic imbalance caused by CRAF (Raf1) kinase mutation alters MAPK signaling and contributes to RASopathies. CRAF proteostasis is maintained by chaperones, Hsp90 which aids in folding, maturation, and activation; whereas Hsp70 recruits ubiquitin ligase CHIP for proteasomal degradation. However, the proteostasis of disease-associated CRAF mutants remains undefined. Here, we identify HECTD3, a HECT-ubiquitin ligase in promoting degradation of hypertrophic CRAF mutants. Interestingly, increased Hsp90 binding to CRAF plays a determinant factor in degradation via HECTD3. Transient increase in Hsp90-CRAF association upon Hsp90 inhibition promote ubiquitination, whereas prolonged inhibition stabilizes mutant CRAF, indicating temporally distinct chaperone functions. Notably, HECTD3 undergoes proteosomal degradation under stress via Cullin-RING ubiquitin ligase Cul5 in a neddylation-dependent manner. Overexpression of Cul5 thus reduces HECTD3 level resulted in stabilization of CRAFD486N, while promoting degradation of wild-type CRAF. Therefore, modulation of the Cul5-HECTD3 axis alters hypertrophic signaling in cardiomyocyte cells. These results establish a hierarchical proteostasis mechanism in which Hsp90 binding primes substrate for HECTD3-mediated ubiquitination, and Cul5-controlled ligase turnover collectively determine mutant CRAF fate.

cell biology↗

bioRxiv2026

Short-term oxycodone exposure produces delayed and persistent gut microbiome disruption in mice

The gut microbiome is a critical part of host homeostasis, yet its resilience following opioid exposure remains poorly understood. While opioid-induced short-term dysbiosis is well documented, the long-term recovery dynamics following oxycodone remain unclear. This study characterized the temporal dynamics of the fecal microbiota in male C57BL/6J mice following a brief 3-day oxycodone regimen (5mg/kg, BID). 16S rRNA gene sequencing was performed at baseline, day 3, 10, 17, and 70. While acute post-treatment phases (day 3 to 10) showed subtle taxonomic shifts in Clostridium_sensu_stricto_1 and Romboutsia, significant community disruption emerged later. By day 17, beta diversity significantly differed from saline controls (P =0.002). At day 70, both alpha diversity (p=0.02) and beta diversity (P=0.007) remained significantly altered, characterized by enriched Akkermansia and Marvinbryantia alongside depleted Eubacterium_xylanophilum. These findings demonstrate that even brief oxycodone exposure triggers persistent, non-recovering dysbiosis that became detectable only after treatment cessation and persisted through day 70. This suggests that the window for microbiome recovery exceeds two months in mice (equivalent to several human years), highlighting a potential long-term risk for patients prescribed short-term opioid courses. ImportanceShort-term opioid exposure is generally assumed to cause only transient disruption of the gut microbiome. However, the duration of microbiome recovery following clinically relevant opioid treatment remains poorly defined. In this study, we show that a brief three-day course of oxycodone in mice resulted in delayed and persistent alterations in gut microbial community structure that remained detectable for at least 70 days after treatment cessation. Notably, significant divergence in microbial composition emerged weeks after exposure rather than immediately following treatment, suggesting that short-term opioid use may initiate longer-lasting remodeling of the gut microbiome than previously appreciated. These findings highlight the importance of considering extended recovery timelines when evaluating the microbiological consequences of opioid exposure.

microbiology↗

bioRxiv2026

Elevated conformational dynamics makes ACKR3 activation-prone and G protein-incompetent

The atypical receptor ACKR3 works together with the canonical chemokine receptor CXCR4 to drive cell migration along gradients of their shared agonist CXCL12. CXCR4 promotes chemotaxis by activating canonical G protein pathways and recruiting {beta}-arrestins. ACKR3 indirectly regulates CXCR4-mediated chemotaxis by scavenging CXCL12. Unlike canonical chemokine receptors, ACKR3 does not couple to G proteins and instead is 100% biased towards {beta}-arrestins. CXCR4 activation by CXCL12 is exquisitely sensitive to subtle changes in both receptor and ligand. By contrast, ACKR3 is activation-prone: it recruits {beta}-arrestins in response to many ligands and is much less sensitive to mutations, suggesting distinct activation mechanisms compared to CXCR4. To explore the basis of these differences, we compared the dynamics of ACKR3 and CXCR4 complexes with chemokines using molecular dynamic (MD) simulations. Ten-microsecond atomistic MD simulations revealed that CXCR4 adopts a stable active state when bound to WT CXCL12 but transitions to an inactive state when in complex with the antagonist variant, [P2G]CXCL12. By comparison, ACKR3 exhibits a variable transmembrane (TM) 6 state distribution and persistently "active" TM7 when complexed with either WT CXCL12 or [P2G]CXCL12, the latter retaining substantial agonistic activity at ACKR3. We further identified ligand-mediated residue interaction networks in the TM core that regulate TM6 and TM7 activation in CXCR4 but are absent or disrupted in ACKR3, resulting in less constrained receptor dynamics. These findings were validated by BRET-based assays with CXCL12 and ACKR3 mutants. Together, the data suggests that the unique conformational dynamics of ACKR3 govern its activation propensity, its ligand promiscuity, and its atypical effector coupling.

pharmacology and toxicology↗

bioRxiv2026

Connecting Cryo-EM and Crystallographic Views of RNA Folding through Ionic Conditions and Structural Flexibility

Discrepancies between biomolecular structures resolved by cryo-electron microscopy (cryo-EM) and X-ray crystallography (XRD) often arise from differences in ionic conditions and construct design, yet their mechanistic impact on RNA folding remains unclear. In the SARS-CoV-2 frameshifting stimulatory element, cryo-EM and XRD structures reveal distinct pseudoknot conformations--a bent and a coaxially stacked state--complicating its structure-function relationship. Here, combining all-atom explicit-solvent simulation results with a structure-based electrostatic model, we show that Mg{superscript 2} ions drive transitions between these states by stabilizing long-range tertiary interactions and modulating local dynamical coupling involving the slippery site and stem 3. Energy landscape analysis reveals distinct folding pathways, while deletion of the slippery segment in crystallographic constructs alters intermediates and produces pathways inconsistent with single-molecule optical tweezer experiments. This study demonstrates how condition-dependent experiments encode complementary interaction-level information and how physics-based computational approaches integrate these to yield a coherent, mechanistic picture of RNA folding. TOC GRAPHICS O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/722415v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@1a7c324org.highwire.dtl.DTLVardef@fcabceorg.highwire.dtl.DTLVardef@736704org.highwire.dtl.DTLVardef@7061e6_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2026

High cRel-expressing germinal center B cells favor precursor plasma cell generation

Germinal centers (GCs) are the site of antibody affinity maturation by the process of somatic hypermutation (SHM) and produce long-lived plasma cells (PCs). GC B cells circulate between two distinct zones: the light zone (LZ) and the dark zone (DZ). It has been demonstrated that the transcription factor NF{kappa}B cRel is required for B cell proliferation and GC B cell maintenance. cRel-deleted GC B cells show reduced SHM and affinity maturation. In contrast, transgenic overexpression of cRel does not affect SHM and results in little or no increase in affinity maturation. Therefore, the function of cRel in regulating SHM and GC B cell-derived PC generation remains unknown. To understand the function of cRel in GC B cell selection, we have used cRel fluorescence reporter mice, which provide insight into cRel function in GC B cells at the level of natural expression variation. We found that about 6-9% of LZ cells increased cRel expression compared to DZ cells, and high cRel-expressing LZ cells showed increased SHM and increased expression of IRF4 and cyclin D2, though cMyc expression remained similar. Combining single-cell RNA-Seq and flow cytometry, we revealed that high cRel-expressing LZ cells are enriched for precursor PCs (pre-PCs), not precursor memory B cells (pre-MBCs). Our findings provide insight into the physiologic function of cRel in regulating GC B cell output.

immunology↗

bioRxiv2026

Repurposed COMT Inhibitors Tolcapone and Entacapone Selectively Suppress Aggregation and Seeding of P301 Mutant TAU in Human Neuronal Models

Background and PurposePathogenic aggregation and propagation of seed-competent TAU assemblies drive tauopathies. MAPT P301 mutations accelerate aggregation and enhance seed competence, yet pharmacological strategies selectively targeting these pathogenic species remain limited. We investigated whether the clinically approved catechol-O-methyltransferase inhibitors tolcapone (TOL) and entacapone (ENT) preferentially modulate mutant TAU aggregation and seeding. Experimental ApproachTOL and ENT effects on TAU aggregation were evaluated via cell-free assays, surface plasmon resonance (SPR), and in silico docking. Functional consequences of compound-modified fibrils were assessed in mutant TAU-expressing SH-SY5Y cells. Translational relevance was examined in human induced pluripotent stem cell (hiPSC)-derived neurons exposed to pathogenic K18 fibrils, followed by post-seeding compound treatment. Key ResultsBoth compounds dose-dependently inhibited TAU aggregation, exhibiting greater potency, stronger SPR binding affinities, and more favorable computed interaction energies for P301S mutant versus wild-type TAU. Fibrils formed with TOL or ENT induced less downstream TAU oligomerization and phosphorylation in SH-SY5Y cells, with TOL showing superior protection. In hiPSC-derived neurons, post-seeding treatment with either compound decreased fibril-induced, sarkosyl-insoluble TAU aggregation and phosphorylation without overt cytotoxicity. Conclusion and ImplicationsTOL and ENT preferentially inhibit the aggregation and seeding of pathogenic P301 mutant TAU. This supports mutation-focused pharmacological strategies and highlights catechol scaffolds as viable starting points for the development of disease-modifying therapeutics. Future research must determine the precise interaction mechanisms with aggregation intermediates and evaluate in vivo efficacy in animal models.

neuroscience↗

bioRxiv2026

An Endocytic Checkpoint Controls Macrophage PD-1 Function and Immunotherapy Fate

Responses to PD1 blockade span durable tumor control to hyperprogressive disease (HPD), yet innate immune mechanisms governing these extremes remain undefined. Here we integrate a macrophage systems atlas (>12,500 transcriptomes) with single-cell profiles from >1,000 anti-PD1 treated patients, to identify CCDC88A (GIV) as a macrophage-intrinsic determinant of durable response versus HPD. GIV loss increases PD1 surface retention, suppresses phagocytosis, and accelerates tumor growth across murine models, human macrophages, and patient-derived organoids. Myeloid-specific GIV deletion converts PD1 blockade from tumor-restraining to tumor-accelerating by reprogramming macrophages toward HPD-like states. Mechanistically, GIV engages a conserved TIR-like [TILL] motif within the PD1 cytoplasmic tail to drive dynamin-dependent endocytosis, coupling innate immune signaling logic to checkpoint receptor trafficking. Pharmacologic disruption of this axis phenocopies GIV loss, revealing an endocytic vulnerability that undermines checkpoint efficacy and triggers accelerated growth at relapse. These findings define PD1 routing, rather than ligand-binding, as a macrophage-encoded checkpoint governing antitumor immunity. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/718292v1_ufig1.gif" ALT="Figure 1"> View larger version (65K): org.highwire.dtl.DTLVardef@a8f83aorg.highwire.dtl.DTLVardef@140dc7dorg.highwire.dtl.DTLVardef@14bc366org.highwire.dtl.DTLVardef@1096b8a_HPS_FORMAT_FIGEXP M_FIG GRAPHIC ABSTRACT C_FIG In briefMullick et al. show that PD1 trafficking in tumor associated macrophages (TAMs), controlled by the endocytic adaptor GIV/Girdin, determines the durability of immunotherapy. Disrupting this pathway traps PD1 at the surface, converts checkpoint blockade into hyperprogressive disease, and abolishes therapeutic benefit, revealing PD1 routing as a macrophage-encoded checkpoint of immunotherapy outcome. HighlightsO_LIGIV defines macrophage states that stratify durable response vs hyperprogressive disease C_LIO_LIGIV limits surface checkpoint accumulation by facilitating PD1 endocytosis C_LIO_LILoss of GIV traps PD1 at the membrane and drives hyperprogressive disease C_LIO_LIEndocytic blockade phenocopies GIV loss and abolishes checkpoint efficacy C_LIO_LIPsychotropic drugs disrupt PD1 trafficking and accelerate tumor growth C_LI

cancer biology↗

bioRxiv2026

Contemporaneity of the past in stochastic intergenerational homeostasis

What recurring patterns of behaviors are hidden in the stochastic intergenerational dynamics of individual bacterial cells in different environments? Embracing the inherently stochastic nature of the homeostasis process, first we reconceptualize homeostasis as representing a "standing pattern of variation in trajectory space" of a naturally occurring adaptive complex system (rather than the simple "set-point" of a self-regulating apparatus), then delineate two mechanistically distinct potential routes to achieving homeostasis (either elastic, reflexive, memory-free adaptation or plastic, reflective, memoryful adaptation) and show that both schemes are simultaneously utilized during multigenerational stochastic growth and division of an individual cell. From experimental data we identify an intergenerational scaling law which directly yields the exact stochastic map governing stochastic intergenerational cell size homeostasis of individual bacterial cells. Its broad applicability across bacterial species, growth conditions, and microenvironments suggests that the organizational motif representing the nature of coupling of growth to division is effectively the same in all of these scenarios, despite apparent differences in actualization through molecular circuitry. The precise parameters characterizing the intergenerational scaling law vary from condition to condition and provide early hints of two tradeoffs: precision-speed and precision-energy.

biophysics↗

bioRxiv2026

Deep-Plant: a supervised foundation model for plant regulatory genomics

Large-scale sequence-to-function deep learning models have demonstrated unparalleled ability to model biological sequences and have revolutionized the field of regulatory genomics. However, the majority of such efforts have centered on human and mammalian systems, leaving plant regulatory genomics comparatively underexplored. To address this gap, we introduce DO_SCPLOWEEPC_SCPLOW-PO_SCPLOWLANTC_SCPLOW, a supervised foundation model trained to predict chromatin state directly from genomic sequence. In contrast to large language models, which are trained in a selfsupervised manner using sequence alone, our model is trained to predict chromatin state across tissues and conditions. Training the model on a large collection of genome-wide experiments including DNA accessibility, transcription factor binding, and histone modifications, provides it with added biological context beyond the sequence itself. We demonstrate that the resulting model is an effective platform for developing accurate models of regulatory activity relevant to gene expression and active enhancers, exhibiting large improvements in speed, accuracy, and interpretability over the complementary approach of fine-tuning DNA language models. DO_SCPLOWEEPC_SCPLOW-PO_SCPLOWLANTC_SCPLOW models are available in Arabidopsis and rice, and work well as a building block for sequence modeling in related species such as corn. Together, these results establish supervised, chromatin-informed foundation models as a practical and effective paradigm for regulatory sequence modeling in plants.

genomics↗

bioRxiv2026

A NOD2-Encoded Toggle Switch Resolves the Host-Microbe Battle Over Cyclic AMP Control

Pathogens hijack macrophages by triggering pathological cAMP surges that block phagolysosomal killing--a defect mirrored in phagocytes from refractory colitis. We identify a host-encoded, pathogen-specific surge-protector comprised of a three-protein toggle: The innate sensor NOD2 binds and masks an evolutionarily conserved motif in GIV that activates trimeric-GTPase Gi, enforcing a biphasic surge-to-plunge cAMP-program: early, NOD2*GIV assembly permits a brief, tolerogenic cAMP rise, whereas subsequent GIV*Gi engagement collapses cAMP to drive phagolysosomal fusion and microbial clearance. Structural, biochemical, and ultrastructural analyses reveal how molecular toggling imposes precise spatial and temporal control. Pharmacogenomic perturbations pinpoint cAMP-PKA hyperactivation as the defining lesion in GIV-deficient macrophages. Functional studies in primary macrophages and human gut organoid co-cultures show that toggling the NOD2*GIV*Gi-axis is necessary and sufficient to convert tolerant macrophages into microbicidal machines that preserve mucosal barrier integrity. These findings uncover a druggable cAMP-control pathway with therapeutic promise in colitis. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/715116v1_ufig1.gif" ALT="Figure 1"> View larger version (70K): org.highwire.dtl.DTLVardef@53cd40org.highwire.dtl.DTLVardef@1c83c82org.highwire.dtl.DTLVardef@fcbc58org.highwire.dtl.DTLVardef@b14f8e_HPS_FORMAT_FIGEXP M_FIG C_FIG eTOC BlurbPathogens hijack macrophages by inducing cAMP surges that help them evade clearance. Anandachar et al. identify a host "toggle switch" in which NOD2 and G proteins compete for GIV, driving a rapid and robust surge-to-plunge transition in cAMP. This temporal switch limits tolerogenic signaling, restores microbial clearance and barrier integrity, and unveils a targetable host pathway in infection and IBD. HighlightsO_LIPathogens exploit cAMP surges in macrophages to block phagolysosomal killing of microbes C_LIO_LIGIV acts as a molecular "toggle" linking NOD2 sensing to Gi-mediated cAMP control C_LIO_LIStructural and mutagenesis studies reveal mutually exclusive binding of NOD2 and Gi to GIV C_LIO_LIPharmacogenomic perturbations pinpoint PKA, not EPAC, as the critical downstream effector C_LIO_LIOrganoid co-cultures show NOD2-GIV-PKA crosstalk safeguards microbial clearance and gut barrier integrity C_LI

immunology↗

bioRxiv2026

The pyruvate branch point controls lymphoid cancer cell dissemination

Cancer cell dissemination critically determines clinical prognosis, yet metabolic dependencies and corresponding therapeutic targets during spread of lymphoid malignancies remain poorly understood. Here we show that the pyruvate branch point operates as a metabolic checkpoint for lymphoid cancer cell migration and disease dissemination through mitochondrial ROS (mROS)/HIF-1a signaling. Isolation of highly migratory mROShi cells led us to identify selective metabolic requirements of malignant lymphocyte migration and disease dissemination. Highly migratory cells show a reprogrammed metabolic profile characterized by increased glucose uptake and reduced glucose-carbon entry into the TCA cycle. Reprogramming of the TCA cycle with downregulation of citrate synthase provide the mechanistic basis for decreased pyruvate oxidation leading to increased migration and disease dissemination through mROS/HIF-1a signaling. Our findings connect central carbon metabolism and migratory capacity of lymphoid cancer cells and identify the pyruvate branch point as a metabolic switch and potential therapeutic target in lymphoid cancer cell dissemination.

cancer biology↗

bioRxiv2026

Archaic translation initiation factor eIF5B supports KSHV late lytic replication and viral oncogenesis by mimicking a hypoxic cellular landscape

Kaposis Sarcoma (KS) Herpesvirus (KSHV) is the etiological agent of KS, an AIDS-defining illness. Recent studies have demonstrated that even during normoxic conditions, KSHV facilitates replication by modulating hypoxia-inducible factors (HIFs), creating a hypoxia-like environment that promotes cellular transformation. KSHV lytic viral genes are favored for protein synthesis during infection by upregulating HIF2 and utilizing the hypoxic eIF4E2 translation initiation complex in oxygen-replete conditions. This translation initiation plasticity (TRIP) links viral replication strategies to angiogenic signaling and oncogenicity. However, the molecular basis of this plasticity remains poorly understood. This study reveals that viral inhibition of eukaryotic initiation factor 2 (eIF2) induces the use of another alternative initiation factor, eIF5B, which canonically mediates delivery of initiator methionine-tRNAiMet during hypoxia. We demonstrate through ribosome density fractionation that eIF2 accumulates in translationally inactive monosome fractions while eIF5B redistributes its translation activity toward polysomes during lytic replication. Progressive dependence on eIF5B during KSHV infection was illustrated by impaired late lytic gene expression, diminished virion production, and altered polysome profiles following eIF5B knockdown. Transcriptomic analyses further reveal that the mRNA landscape in KSHV-infected cells depleted of eIF5B mirrors that of uninfected hypoxic cells. Moreover, silencing of eIF5B in a natural infection model reduces both VEGF secretion and anchorage-independent growth--two hallmarks of KSHV viral oncogenicity. These results demonstrate that eIF5B functions as an essential component of alternative translation initiation machinery activated in response to eIF2 inactivation during KSHV lytic replication, emphasizing its key role in viral pathogenesis and its potential as a novel therapeutic target. IMPORTANCEKaposis Sarcoma Herpesvirus (KSHV) is a human cancer virus that causes severe malignancies in immunocompromised individuals, including the blood vessel cancer Kaposis Sarcoma and a highly aggressive body-cavity lymphoma. Although cells mount a response to infection by shutting down global protein synthesis, how viruses like KSHV continue making viral proteins when this cellular machinery is shutdown remains unclear. This study uncovers that KSHV exploits an ancient cellular factor typically used for protein production under low-oxygen conditions. We identify a previously unrecognized role for the translation factor, eIF5B, in supporting KSHV replication and select production of viral proteins during infection. Our work further demonstrates that KSHVs utilization of eIF5B contributes to a hypoxia-like environment during infection, ultimately contributing to cancer-promoting changes within the cell. These findings highlight a new strategy for tumor-virus reprogramming of host cells, opening an avenue for novel therapeutic targets to eliminate virus-associated cancers.

microbiology↗

bioRxiv2026

A functional annotation based integration of different similarity measures for gene expressions

Genes with similar expression profiles often exhibit similar functional properties. An "integrated similarity score" (ISS) is developed by combining different expression similarity measures through weights, obtained using biological information, for improving gene similarity. The expression similarity measures are converted to the common framework of positive predictive value using functional annotation. A fitness function, called "fitness function using functional annotation of genes" (FFFAG), is also developed by minimizing the difference between functional similarity value and the ISS. The FFFAG is used to determine the weight combination of different similarity measures in ISS. In addition, an existing similarity measure, called TMJ (integrated similarity measure by multiplying Triangle and Jaccard similarity), is also modified to incorporate biological knowledge involving functional annotation. The results demonstrate that ISS is superior to individual similarity measure to find similar gene pairs. Further, the ISS predicts the functional categories of 40 unclassified yeast genes at p-value cutoff of 10-10 from 12 clusters. The associated code is accessible at http://www.isical.ac.in/[~]shubhra/ISS.html.

bioinformatics↗