Biology subjects

Kumar, A.

Publications and source records attributed to Kumar, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Refactored intracellular domains enhance CAR expression and function

Chimeric antigen receptor (CAR) intracellular domains (ICDs) couple antigen recognition to T-cell activation. Here we worked with AI agents to refactor CAR ICDs, proposing designs and refining them as measurements accumulated over three rounds. This approach yielded KRGR-{zeta}1-40, a 45-amino-acid ICD less than a third the length of the clinical 4-1BB-CD3{zeta} (BB{zeta}) ICD. Compared with BB{zeta}, KRGR-{zeta}1-40 increased CAR surface expression and antigen-driven IFN-{gamma} release, without detectable antigen-independent IFN-{gamma} release. KRGR-{zeta}1-40 extended CAR expression and target-cell killing after transient mRNA delivery, supported the expression of four CARs from a shared mRNA dose, and enhanced B-cell depletion after lentiviral and lipid-nanoparticle delivery. These results show that iterative agent-guided design with experimental feedback can yield compact ICDs that enhance CAR expression and sustain CAR T-cell function.

bioengineering↗

bioRxiv2026

Topoisomerase IIβ Kinase: A protein processed from HIV-1 envelope by cysteine proteases

Topoisomerase II{beta} (TopoII{beta}) is activated by TopoII{beta} kinase (TopoII{beta}K) during HIV-1 infection. This study demonstrates that TopoII{beta}K is produced within 15-30 minutes post-infection and is conserved across the major HIV-1 subtypes (A, B, C, and D), although its formation kinetics vary among subtypes. Immunological mapping using specific anti-gp120 antibodies suggests that TopoII{beta}K is derived from the N-terminal region of gp120. Notably, TopoII{beta}K co-localizes with endolysosomes shortly after viral entry. Inhibition of cysteine proteases using E64d and Z-FY-CHO abrogates TopoII{beta}K production in both HIV-1-infected and envelope-transfected cells. These findings suggest that TopoII{beta}K is a conserved polypeptide product of gp120 processing within endolysosomes during the early stages of HIV-1 infection, highlighting a potentially conserved mechanism that regulates early viral infection across major HIV-1 subtypes.

molecular biology↗

bioRxiv2026

HIV-1 Integrase C-terminal Domain Mutations Regulate ALLINI Sensitivity

HIV-1 integrase (IN) performs two essential functions in virus replication: catalyzing the integration of reverse-transcribed viral cDNA into host DNA and promoting proper virion maturation. The latter is mediated by IN binding to the genomic RNA (gRNA) through basic residues in its C-terminal domain (CTD), thereby ensuring correct repackaging of the gRNA inside the remodeled capsid lattice. Allosteric integrase inhibitors (ALLINIs) bind within a V-shaped pocket at the interface of the catalytic core domain (CCD) of IN dimers, promote the engagement of the CTDs from adjacent dimers, and thereby occlude gRNA binding. However, virological evidence supporting a role for the IN-CTD in ALLINI mechanism of action is lacking, as mutations in CTD residues implicated in ALLINI-induced multimerization typically yield non-infectious viruses. Here we demonstrate that IN-CTD mutants bearing compensatory substitutions that restore infectivity exhibit markedly altered sensitivity to ALLINIs. Specifically, we show that R269A/K273A substitutions within the IN-CTD confer ~100-fold enhanced sensitivity to ALLINIs. This correlates with a loss of IN-gRNA binding at low ALLINI concentrations, while exerting minimal effects on IN multimerization in vitro and in virions. In contrast, we found that the IN I268T substitution which arose as a suppressor mutation of I268A confers partial resistance to ALLINI-induced inhibition of gRNA binding. Collectively, these findings provide the first direct genetic evidence that CTD residues, despite being distal to the primary drug binding site at the CCD dimer, modulate sensitivity to ALLINIs and highlights IN-gRNA binding as a central target of ALLINIs.

microbiology↗

bioRxiv2026

A note on the responsiveness of neurons

The time constant of biological neurons is typically around 5-20ms. This raises the question of how a system made of slow devices can generate and respond to faster signals. A potential solution is a population of uncorrelated neurons that, as a whole, can respond arbitrarily fast despite individual slow neurons. The response speed of a neuron or a population of neurons is often defined as how quickly neurons increase or decrease their firing rate given a change in the input, similar to a reaction time. Less emphasis is placed on the recovery time - the duration to transition and settle in a stationary state again in response to the change in the input. Here, we investigate the reaction time and the recovery time of leaky integrate-and-fire neurons given a change in, either the mean input drive, or input fluctuation level. In general, neurons react faster and fire quicker the larger the change in the input. Moreover, the closer the membrane potential is to the threshold, the faster the reaction. We observe that neurons react faster to increased input fluctuations than to stronger mean drive. Complementarily, the recovery time is systematically longer the larger the input change. Also, the closer the neurons' potentials are to the threshold, the longer it takes the neuron to recover. Most interestingly, the baseline firing of the neuron strongly determines whether recovery is faster when the change happens in the mean drive or the fluctuation level. This observation suggests that communication via the mean drive and the fluctuation level could target different aspects of the overall response - quick reaction time and the recovery speed - based on the initial baseline activity.

neuroscience↗

bioRxiv2026

Pharmacologic inhibition of OLIG2 by CT-179 disrupts mitotic progression and enhances standard-of-care chemoradiotherapy in glioblastoma

Background: Glioblastoma (GBM) remains uniformly fatal. The basic helix-loop-helix transcription factor OLIG2 is universally expressed in diffuse glioma, sustains the tumor-propagating stem-like fraction, and opposes p53-mediated genotoxic responses, but has historically been considered undruggable. We characterized CT-179, a brain-penetrant small-molecule OLIG2 inhibitor, as a candidate therapeutic for GBM. Methods: Target engagement was assessed by cellular thermal shift assay, differential scanning fluorimetry, crosslinking, and electrophoretic mobility shift assay. Anti-tumor activity was evaluated across eleven patient-derived GBM cell lines, in zebrafish oligodendrocyte precursor cell (OPC)-derived tumors, and orthotopic and subcutaneous GBM models, including combinations with radiotherapy plus temozolomide and with selective Bcl-2 family inhibitors. Results: CT-179 engaged OLIG2 and altered the conformation of dimerized OLIG2 on chromatin without preventing dimerization or fully displacing OLIG2 from DNA. CT-179 reduced OLIG2-positive OPCs and OPC-derived tumors in zebrafish and inhibited proliferation across all eleven GBM lines (IC50 < 360 nM), depleting the stem-like fraction and inducing apoptosis through mitotic catastrophe, mitotic slippage, or a hybrid response. RNA-seq across three patient-derived glioma stem cell lines identified {beta}-tubulin gene isoforms as the most strongly CT-179-down-regulated transcripts at the early time point and showed correlated {beta}-tubulin and OLIG2 expression in TCGA glioblastoma specimens. Combination with the Mcl-1 inhibitor S63845 or the Bcl-2/Bcl-xL/Bcl-w inhibitor ABT-737 produced greater-than-additive cytotoxicity. Oral CT-179 crossed the blood-brain barrier and extended median survival as a single agent across two independent adult orthotopic patient-derived backgrounds (GBM8 and G06); intraperitoneal CT-179 was associated with reduced intratumoral OLIG2 and Ki-67 staining in GBM8 orthotopic xenografts. Combination of CT-179 with radiotherapy and temozolomide eliminated all GBM8 and 7 of 8 GBM4 subcutaneous xenografts without weight loss and significantly extended survival when combined with radiotherapy in MGMT-unmethylated GBM6 orthotopic xenografts (CT-179 administered intraperitoneally). Conclusions: CT-179 is a brain-penetrant inhibitor of an established GBM driver with single-agent and combination activity across several orthotopic models, supporting clinical development for GBM and translational evaluation in other OLIG2-dependent CNS malignancies.

cancer biology↗

bioRxiv2026

Sphingosine-1-Phosphate Restrains Foamy Macrophage Formation and Modulates Inflammation in Multidrug-Resistant Tuberculosis

The high abundance of lipid loaded foamy macrophages in the granulomatous lesions is a key hallmark of advanced pulmonary tuberculosis. These macrophages serve as lipid-rich intracellular hiding niches that provides Mycobacterium tuberculosis (M.tb) survival and persistence during prolonged anti-tubercular treatment. Persistent infection and dysregulated inflammatory responses further complicate disease control, particularly in multidrug-resistant tuberculosis (MDR-TB), highlighting the need for host-directed therapeutic strategies. Our previous preclinical study demonstrated the therapeutic potential of the host sphingolipid sphingosine-1-phosphate (S1P) in reducing the mycobacterial burden in M.tb-infected animals, accompanied by enhanced pulmonary macrophage activation. To explore its translational potential, we investigated whether S1P modulates foamy macrophage formation in CD14 monocyte-derived macrophages from healthy donors and patients with MDR-TB. Notably, macrophages derived from patients with MDR-TB exhibited greater intracellular lipid droplet accumulation than those derived from healthy donors. This phenotype was further characterized by increased nitric oxide production and quantitative RT-PCR analysis of genes associated with immune regulation and lipid metabolism, including APOE, LDLR1, CLEC4E, CLEC7A, GP96, IL4, and IL13, which suggested a Th2-associated immunological bias in foamy macrophages. Collectively, these findings highlight the potential role of host sphingolipids in modulating foamy macrophage phenotypes and associated immunometabolic dysregulation, supporting further investigation of S1P as a host-directed therapeutic strategy against MDR-TB.

immunology↗

bioRxiv2026

Enabling Spatially Resolved Gene Expression Analysis in a Multilayer Paper-Based In Vitro Tumor Model

Spatial heterogeneity is a defining feature of solid tumors, yet molecular analysis of three-dimensional (3D) in vitro tumor models is often performed as a bulk measurement, potentially obscuring position-dependent cellular responses. Here, we establish an analytical workflow for spatially resolved gene-expression analysis in a multilayer paper-based 3D tumor model by addressing the challenge of RNA recovery from cell-laden paper-Matrigel scaffolds. Using MDA-MB-231 breast cancer cells, multiple RNA extraction strategies were systematically evaluated across conventional culture, Matrigel, paper, and composite paper-Matrigel environments. Conventional silica-column extraction yielded only 3.39 {+/-} 0.93 ng/L RNA from the composite scaffold, whereas TRIzol-chloroform extraction increased recovery to 201.13 {+/-} 38.32 ng/L. The optimized workflow subsequently enabled independent RNA recovery from all eight layers of a multi-layer paper-based in vitro tumor, with concentrations ranging from 113.06 {+/-} 5.00 to 245.26 {+/-} 15.76 ng/L and purity suitable for downstream qRT-PCR analysis. Layer-resolved profiling of untreated cultures revealed distinct spatial patterns in HIF1A, HIF2A, VEGFA, and GLUT4 expression across the multilayer construct. To evaluate the workflow under pharmacological perturbation, cultures were exposed to 5 M doxorubicin (DOX) for 48 h. Relative cell viability exhibited a non-monotonic spatial profile, whereas DOX-induced HIF1A and VEGFA expression generally became more pronounced toward the deeper layers when each treated layer was compared with its corresponding untreated control. Collectively, these results establish a workflow for converting individually addressable regions of a multilayer 3D tumor model into spatially resolved transcriptional measurements, enabling analysis of both baseline spatial heterogeneity and position-dependent molecular responses to pharmacological perturbation.

bioengineering↗

bioRxiv2026

Trust, but Verify: Epistemic Uncertainty-Gated Pseudo-Labeling for Safer Semi-Supervised Malaria Screening

Background. Deep learning could make malaria microscopy more accessible in low-resource settings, but training it usually needs thousands of expert-labeled images. Semi-supervised learning (SSL) reduces this need by also learning from unlabeled images. However, methods such as FixMatch accept pseudo-labels using a fixed confidence threshold, and lowering that threshold to use more data can teach the model to make confident mistakes. These silent failures are dangerous because they are never flagged for expert review. Methods. We added an uncertainty check to SSL pseudo-labeling. Each unlabeled image is passed through the model ten times with Monte Carlo dropout active; its pseudo-label is used only if the normalized predictive entropy of the averaged prediction is below 0.2, and it is then weighted by a sigmoid function of model confidence. We trained a ResNet-18 on the public Malaria Cell Images dataset (27,558 cell images; 70:10:20 split) using labels for only 20% of the training images. Over six random seeds, we compared the method with a supervised baseline and with FixMatch at thresholds of 0.95 and 0.80, measuring accuracy, AUC and silent failure rate (SFR), the share of predictions that are wrong despite confidence above 0.95. Results. Compared with FixMatch at a threshold of 0.80, our method lowered the mean SFR from 2.28% to 1.52%, a 33.2% relative reduction (paired t-test, t = 4.29, p = 0.0078), and the SFR was lower in all six runs. Accuracy (96.6% vs. 96.4%) and AUC (0.991 for both) were unchanged. The mean SFR was also below that of FixMatch at 0.95 (1.97%) and of the supervised baseline (2.01%). Conclusions. Checking epistemic uncertainty before accepting pseudo-labels reduced confident errors in semi-supervised malaria cell classification without lowering accuracy. Patient-level and external validation on whole-slide images are still needed before clinical use.

bioinformatics↗

bioRxiv2026

Functional dissection of ultraconserved non-coding regions present near the Hoxd complex in vertebrates

Ultraconserved non-coding elements (UCEs) exhibit near-perfect sequence conservation across vertebrate genomes, implying severe evolutionary constraint and essential biological functions. Near the vertebrate Hoxd complex, a master regulator of axial patterning and limb morphogenesis, three Conserved Regions (CR1, CR2, and CR3), each longer than 200 bp, show exceptional sequence conservation and drive robust regulatory activity in transient reporter assays. To investigate their endogenous in vivo functions, we used CRISPR-Cas9 genome editing to generate a series of deletions of varying sizes spanning CR2 and CR3, individually and in combination, within hoxd complex in zebrafish. Screening thousands of founder embryos revealed a clear size and conservation-dependent relationship between deletion extent and viability: complete deletion of core CR2, CR3, or the combined CR2/3 loci could not be transmitted through the germline, whereas the smaller and least-conserved deletion, a 626-bp subregion of CR2, was the only allele to yield a stable, homozygous-viable line. Our results indicate that the core ultraconserved sequences are developmentally indispensable, whereas flanking, less conserved sequences can be removed without consequence. Detailed phenotypic and molecular characterization of the mutant embryos revealed normal primary and secondary body axis formation, intact somite segmentation, and unperturbed craniofacial and pectoral fin development. Expression of hoxd complex genes showed no significant change in mutant embryos by RNA-ISH or RT-qPCR, although RT-qPCR revealed a consistent, modest upward trend at 24 and 48 hpf, raising the possibility that the deleted subregion normally contributes a weak repression of overall hoxd complex genes transcription. While this study reveals some functional aspects of ultraconserved elements, much more remains to be uncovered.

developmental biology↗

bioRxiv2026

A zebrafish ribosomopathy links Ddx49-dependent ribosome biogenesis to differential failure of population expansion among pigment cell lineages

Defects in ribosome biogenesis cause surprisingly cell-type specific pathologies. Why some cells are more vulnerable than others remains unresolved. Here, we show that a zebrafish pigment pattern mutant, sisley, results from a viable missense allele of ddx49, encoding a DEAD-box RNA helicase required for small ribosomal subunit biogenesis and early survival. In mutants, yellow/orange xanthophores fail to proliferate during development of the adult stripe pattern, whereas closely related melanophores are affected only mildly. The mutant protein has reduced helicase but increased ATPase activity and, consistent with impaired ribosome biogenesis in vivo, mutants are deficient in 18S rRNA relative to its precursor. Remarkably, a similar deficiency in 18S rRNA relative to precursor rRNA distinguishes xanthophores from melanophores even in the wild-type, suggesting an inherent difference in rRNA processing that leaves xanthophores vulnerable to Ddx49 dysfunction. Both xanthophore deficiency and a male-biased sex ratio of the sisley mutant were rescued by mutation of tp53. Our findings suggest how baseline differences in ribosome production shape cell-type specific outcomes of mutations affecting ribosome biogenesis, and provide an adult-viable model for Ddx49 function in vivo.

developmental biology↗

bioRxiv2026

Methane oxidation genes are only present in bioaerosols at sites with high methane concentrations

Airborne microorganisms connect Earth's habitats, yet a central question in microbial ecology remains: do airborne communities simply reflect what is released from the surfaces below, or does local environmental context shape them? Methane oxidation offers a tractable test, because methane monooxygenase genes are restricted to defined bacterial lineages and methane is directly measurable. Pairing aerosol and underlying soil or water metagenomes across a 35-fold methane range, we show that these genes were present in every soil and water sample, yet detected in aerosols only above the landfill and dairy farm, and not above the three wetlands. The soil did not predict the air; instead, gene detection tracked local methane availability. Aerosol cell concentrations spanned 1,000 to 1,000,000 cells per cubic metre, and bacterial communities tracked their sources whereas eukaryotes were enriched in spore-dispersing fungi. Predicting what airborne microbiomes carry will therefore require accounting for local environmental context, not passive dispersal alone.

microbiology↗

bioRxiv2026

Endothelial CD36 links arteriolar remodeling to metabolic and immune alterations in Alzheimer's disease

Alzheimer's disease (AD) is accompanied by profound cerebrovascular dysfunction, yet it remains unclear whether endothelial abnormalities reflect only vascular injury or altered regulatory states connecting vascular remodeling with metabolic and immune function. We integrated human brain transcriptomic and histopathological data with complementary mouse genetic and human iPSC-derived endothelial models to examine a CD36-centered endothelial-state framework in AD. Analysis of the Seattle Alzheimer's Disease Brain Cell Atlas identified three CD36-expressing endothelial populations; intermediate-CD36 Endo 2 predominated in both reference and AD samples, although the relative subset distributions differed. In hippocampal and cortical tissue from six AD patients, endothelial CD36 expression was observed in structurally abnormal arterioles with disrupted mural organization. In complementary mouse studies, endothelial PKD1 deficiency was accompanied by increased vascular CD36 and cerebrovascular abnormalities, whereas endothelial CD36 deletion altered lipid-metabolic and innate-immune-associated transcriptional programs, including AD-relevant genes and pathways, under metabolic-inflammatory stress. To assess whether distinct human endothelial backgrounds remained pharmacologically responsive, we treated control and CRISPR-engineered PSEN1 E276A iPSC-derived ECs with GS4898, a small molecule previously linked to arterial specification. Both maintained endothelial identity but displayed different baseline transcriptional profiles and shared as well as cell-line-dependent responses to GS4898. Together, our study identifies a CD36-associated vascular phenotype in human AD, demonstrates CD36-dependent regulation of brain endothelial metabolic-immune transcriptional programs in vivo, and reveals endothelial transcriptional responsiveness to pharmacologic perturbation. The observed endothelial transcriptional plasticity may provide a basis for developing state-selective therapeutic strategies targeting cerebrovascular dysfunction in AD.

neuroscience↗

bioRxiv2026

Mutation in Ralstonia pseudosolanacearum F1C1 cytochrome p450 homologue impairs virulence through root but not through shoot of tomato plants

Ralstonia pseudosolanacearum causes a lethal bacterial wilt disease in many plants. The pathogen enters through roots and systemically colonizes the xylem tissue, resulting in wilting and eventual death of the host. Using transposon mutagenesis followed by screening in tomato seedlings, we identified a mutant exhibiting virulence deficiency by root inoculation, but not by leaf inoculation. The mutation was recreated in the wild-type genome via natural transformation to prove the linkage of virulence deficiency with the transposon insertion. Whole genome sequencing revealed that the transposon insertion was in a cyp450 homologue, encoding cytochrome P450 monooxygenase. Two independent targeted mutations were generated in the cyp450 gene, and both the mutants recapitulated the root-specific virulence deficiency. The mutants rarely cause disease in mature plants following soil drenching but remained fully virulent upon petiole pricking inoculation. Complementation with the full length cyp450 gene amplified from the wild-type strain F1C1 restored virulence in the mutant. Compared to the wild-type F1C1, the cyp450 mutants exhibited greater sensitivity to tomato root extracts. HR-LCMS of tomato root extracts revealed the occurrence of several antimicrobial compounds. Cytochrome P450 enzymes are known to mediate the degradation or biosynthesis of diverse metabolites in bacteria. Considering the virulence difference due to the mode of inoculation, our findings indicate that cyp450 contributes to tissue-specific adaptation of the pathogen during infection has opened an avenue for integrating tissue metabolomics with bacterial wilt susceptibility research.

pathology↗

bioRxiv2026

A domesticated prophage enzyme links bacterial metabolism, episymbiosis and phage fitness

Prophages are increasingly recognized as drivers of bacterial adaptation, yet how domesticated prophage genes shape microbial symbiosis remains poorly understood. Here, we identify GDPDXhp1, a glycerophosphodiester phosphodiesterase encoded by prophage Xhp1 of Schaalia odontolytica XH001, as a metabolic regulator linking bacterial physiology, episymbiosis and phage fitness. Association with the epibiotic bacterium Nanosynbacter lyticus TM7x strongly induced gdpdXhp1 expression and triggered GDPDXhp1-dependent lipid droplet accumulation in XH001. GDPDXhp1 orchestrated glycerophospholipid remodeling, membrane biophysical properties, respiration and cell-surface glycan composition in XH001, thereby promoting TM7x episymbiosis. Remarkably, GDPDXhp1 was also required for productive Xhp1 infection by remodeling susceptible bacterial host envelope structures to facilitate phage-host interactions. Together, our findings reveal a domesticated prophage enzyme that contributes to bacterial metabolism, interspecies symbiosis and viral reproduction, highlighting prophage as an overlooked source of metabolic innovation in microbial symbiosis.

microbiology↗

bioRxiv2026

A minimally modified AAV9 variant (AAV9fyhr) for enhanced CNS gene delivery by intra-CSF administration.

Adeno-associated virus (AAV) mediated gene therapies continue to show promise for the treatment of genetic diseases. However, delivery and safety still represent significant prohibitive barriers restricting translation of these delivery modalities to humans. We describe a novel AAV9 variant, AAV9fyhr or FYHR, identified from screening a controlled randomization point mutation AAV9 library in mice and nonhuman primates followed by directed evolution of in vivo CNS-derived libraries screened in human iPSC differentiated cortical neurons. Capsid variants of interest were identified by coupling long read sequencing and computational structure modeling. Structure-function analysis of our lead candidate reveal that two targeted amino acid changes are sufficient to result in beneficial pleiotropic effects across viral stability, cell surface attachment, and gene delivery. We developed a novel multiplexed barcoding reporter system, enabling a time and cost-effective option to gather spatially resolved single cell capsid validation data in mice and nonhuman primates. When administered intrathecally, AAV9fyhr exhibits 2-to-6-fold higher gene delivery across several tissues in the CNS while minimally altering the tropism to peripheral tissues in comparison to AAV9. These findings demonstrate the potential of whole capsid engineering for the rational development of translational next generation AAV capsids.

bioengineering↗

bioRxiv2026

Zinc preserves translational proteostasis to counteract aging and proteotoxicity in Caenorhabditis elegans

The progressive collapse of protein homeostasis is a defining feature of aging, yet upstream events deteriorating protein synthesis remain poorly defined. Zinc is an essential micronutrient and structural cofactor, but its role in the aging ribosomal proteome is poorly understood. Here, using Caenorhabditis elegans, we show that the whole-body zinc-to-sulfur ratio declines by ~90% between day 1 and day 13 of adulthood. Supplementing aged animals with a low dose of ZnCl2 (10 M) restored organismal zinc levels and improved healthspan, thermotolerance, and oxidative stress resistance, whereas higher doses were toxic. Quantitative proteomics revealed that structural constituents of the ribosome were the most enriched functional category altered by zinc, accompanied by robust induction of metallothionein MTL-1. Zinc-supplemented animals exhibited increased protein synthesis and a transient increase in ribosomal proteins, while 45S pre-rRNA levels remained unaffected, indicating that zinc acts on ribosome structure rather than biogenesis. Zinc supplementation coordinately reversed the age-associated decline of antioxidant, chaperone, proteasome, and energy-metabolism factors. Furthermore, zinc conferred protection in two proteotoxicity models, suppressing polyglutamine aggregation and delaying amyloid-beta-induced paralysis. Our findings establish that zinc preserves translational proteostasis, identifying late-life zinc supplementation as a translatable micronutrient intervention against age-related proteinopathies.

molecular biology↗

bioRxiv2026

Force Induces a Collective Inactivity in Dyneins to Auto-Regulate Bidirectional Transport of Vesicles

The back-and-forth (bidirectional) motion of Endosomes and Phagosomes along microtubules is fundamentally important for nutrient absorption, receptor recycling, pathogen degradation and many other downstream cellular pathways. Bidirectional motion requires opposing force from the kinesin and dynein motors, causing intermittent Tug-of-Wars (ToWs) followed by runs where winning motors drive their cargo until the next ToW. Dynein displays a counter-intuitive behaviour of catch bonding tightly to the microtubule against opposing force, but why such a response is needed remains unknown. Here we force dynein to catch-bond using an optical trap when dynein-teams are transporting phagosomes in-vitro and inside cells. An unexpected form of collective inactivity emerges, where larger dynein-teams remain inactive for longer after they have been catch-bonded. Kinesin exhibits a high probability of reversing the cargo during such periods of inactivity of dyneins. Therefore, the inactivity of dyneins allows cells to auto-regulate the effect of opposing molecular forces for the purpose of achieving bidirectional motion of endosomes/phagosomes. We propose that an inactivating conformational change is induced when dynein detaches against strain from its catch-bonded state. Collective inactivity follows because each inactive dynein exerts an entropic force on the other to delay the entire team from being activated. We capture the dynamics of this process through a statistical model of Entropically Constrained Reactivation, obtaining predictions in excellent agreement with experimental data. The emergent behaviour observed here possibly prevents a runaway dynein situation inside cells, its simple entropic origin suggesting broader relevance to catch-bonded receptor ligand interactions operating under conditions of cellular crowding.

cell biology↗

bioRxiv2026

Myeloid Ecotropic Viral Integration Site 1 Promotes Vascular Smooth Muscle Differentiation through Activation of Myocardin

Myocardin (MYOCD) is the principal trigger-switch for vascular smooth muscle cell (VSMC) growth arrest and the cyto-contractile program of differentiation. While much is known about MYOCD-dependent VSMC differentiation, limited information exists as to its transcriptional regulation. Here, we report moderate mRNA expression of Myeloid Ecotropic viral Integration Site 1 (MEIS1) in both human and mouse VSMCs. MEIS1 mRNA is reduced in parallel with myocardin mRNA and protein under conditions of VSMC de-differentiation. Loss- and gain-of-Meis1 expression studies in cultured human VSMCs and the mouse carotid artery support MEIS1 as an activator of myocardin and VSMC differentiation. CRISPR-mediated knockout of the endogenous Meis1 gene in vitro and elicited lower levels of VSMC contractile markers and an increase in growth rate. Similarly, Femoral artery wire injury, in mice lacking Meis1 showed and increased of neointima formation. Mechanistically, we identified a conserved MEIS1 response element in the proximal promoter of myocardin, that appears to contribute to the promoter activity, suggesting a potential regulatory interaction. These results suggest a previously unrecognized role for the homeodomain protein MEIS1 in the activation of myocardin and the quiescent, VSMC differentiated state.

molecular biology↗