Biology subjects

Bandyopadhyay, A.

Publications and source records attributed to Bandyopadhyay, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Molecularly Distinct Innexin Gap Junction Channels and Undocked Hemichannels Regulate Glia Morphology and Function in Caenorhabditis elegans

Glial cells across species orchestrate nervous system development, maintenance, and function through ionic and metabolic crosstalk. These cells express multiple gap junction and hemichannel components. However, the mechanisms by which glia communicate utilizing these channel components to support nervous system architecture and function remain poorly understood. By studying GLR glial cells in Caenorhabditis elegans, we show that individual glial cells employ distinct innexin channel configurations to support specific functional roles, utilizing discrete downstream cellular mechanisms. We find that while innexins UNC-7and UNC-9 function as gap junction channels, the innexin INX-18 operates independently to form undocked hemichannels in discrete domains within the same cells. In this combinatorial configuration, UNC-7/UNC-9 gap junctions specifically regulate Synaptobrevin/SNB-1 localization in RME neurons, as previously reported. However, both channel types function non-redundantly to maintain glial morphology by regulating the Calpain/CLP-4 and CDK-5 pathway through cell-autonomous modulation of intracellular calcium levels. In contrast, INX-18 hemichannels in GLR glia regulate high-salt-induced paralysis behavior through distinct downstream cellular mechanisms, potentially in conjunction with tyramine signaling. Altogether, our findings demonstrate a broader repertoire of innexin channel utilization at the individual glial cell level, supporting specific aspects of nervous system maintenance and functioning.

neuroscience↗

bioRxiv2026

An Improved Protocol for High-Efficiency and Cost-Effective CRISPR/Cas9-Mediated Knock-ins in C. elegans

CRISPR/Cas9-mediated homology-directed precise genome editing using long single-stranded DNA (ssDNA) donors has expanded the possibilities for generating defined genetic modifications. However, the required ssDNA donor preparation can be technically demanding and often requires extensive locus-specific sequence design. Here, we examined parameters influencing ssDNA donor-mediated genome editing and developed approaches to simplify donor preparation using {lambda}-exonuclease-mediated ssDNA generation. By evaluating donor designs across multiple genomic loci, we found that efficient genome editing can be achieved with relatively short homology regions for a range of insertion sizes. These findings provide a basis for simplifying ssDNA donor preparation and the overall gene-editing pipeline, potentially facilitating its application across species.

molecular biology↗

bioRxiv2026

A Multi-Epitope Vaccine Design for Human Pasteurellosis using Outer Membrane β-barrel Proteins of Pasteurella multocida

Pasteurella multocida is a facultative anaerobic, Gram-negative coccobacillus that causes pasteurellosis in companion animals, livestock, and poultry and poses a significant zoonotic risk to humans through bite wounds, scratches, licking, and transfer of bodily fluids. Although vaccines are available for livestock and poultry, no vaccine is currently licensed for human use. In this study, we systematically identified and characterized 29 outer membrane {beta}-barrel (OMBB) proteins in P. multocida Past9 proteome and classified them into functional categories, including TonB-dependent receptors, porins, autotransporters, adhesins, and efflux pumps. B-cell, cytotoxic T-lymphocyte (CTL), and helper T-lymphocyte (HTL) epitopes were predicted from the identified proteins and screened based on antigenicity, non-allergenicity, and non-toxicity. Epitopes conserved across eight human-infecting P. multocida strains and located within the extracellular loop (ECL) region were incorporated into a multi-epitope vaccine (MEV) construct. The designed MEV was predicted to be antigenic, non-allergenic, and soluble. Its tertiary structural model was iteratively refined and validated. Molecular docking with human toll-like receptors 4/2 (TLR4/TLR2) predicted stable interactions, further supported by 100 ns molecular dynamics simulations. Immune simulation of the MEV construct predicted a strong simulated immune response. Furthermore, codon optimization and in silico cloning supported the feasibility of recombinant MEV expression in E. coli. The construct was benchmarked against OmpH, a well-known antigenic protein and exhibited broadly comparable predicted immune responses and receptor-binding energetics. This study proposes a designed MEV candidate against human pasteurellosis and highlights OMBB proteins as potential immunogenic targets for vaccine development.

bioinformatics↗

bioRxiv2026

Design of a Multi-epitope Vaccine Against Human Glanders Targeting Outer Membrane β-barrel Proteins of Burkholderia mallei

Burkholderia mallei, a facultative intracellular Gram-negative pathogen, is the causative agent of glanders that primarily affects solipeds and is sporadically transmitted to humans. Current interventions mainly rely on antibiotics; however, increasing antimicrobial resistance and the lack of a licensed vaccine further complicate disease management. Surface exposed outer membrane {beta}-barrel (OMBB) proteins serve as excellent targets for vaccine development. In the present study, a consensus-based computational framework was employed on the B. mallei turkey2 proteome that identified 59 OMBB proteins - including porins, TonB receptors, autotransporters, and efflux components. These OMBB proteins were leveraged to predict B- and T-cell epitopes which were manually curated, and mapped onto the corresponding protein models to identify surface-exposed epitopes with direct accessibility to the host immune cells. These epitopes were linked together to construct a multi-epitope vaccine (MEV) that was predicted to be antigenic, and soluble upon overexpression. The tertiary structure of the MEV was generated which was used for molecular docking with TLR4 and TLR2. Molecular dynamics simulation and flexibility analysis confirmed the structural stability of the MEV-TLR4/TLR2 complexes. In-silico immune simulation showed the capability of MEV to induce a strong immune response. Codon optimization and in-silico cloning were performed to evaluate its efficient expression in the E. coli host. The findings suggest that surface exposed OMBB proteins can serve as promising antigenic candidates for designing an MEV construct.

bioinformatics↗

bioRxiv2026

Spike antibodies targeting GRP78 predispose to cardiovascular complications compared to Dengue

One major aftermath of COVID-19 pandemic is cardiovascular consequences. SARS-CoV-2 binds to ACE2 and downregulates vasodilation. Dengue favors hypotension by weakening endothelial glycocalyx leading to plasma leakage. C1q levels, immune complexes (ICs), and proteomic profiles in serum samples from 52 COVID-19 and 19 pre-pandemic Dengue cases were studied. Unlike Dengue, COVID-19 serums showed elevated coagulation proteins promoting vaso-occlusion and peripheral artery diseases. The stress-induced chaperone and atherosclerosis marker, GRP78 (gene/ protein) was found upregulated upon SARS-CoV-2 spike expression in cardiac/ lung cell lines. Elevated GRP78 levels were also observed in serum samples from COVID-19-diagnosed individuals and subjects with myocardial infarction (MI) in post COVID-era. Surprisingly, spike antibodies (Abs) showed cross-binding to GRP78 and possibly contributed to the observed higher-level ICs in COVID-19 serums (cardiovascular embolism?). Co-localization studies showed that spike Abs (analogous to pro-atherosclerotic GRP78 auto-Abs) could directly bind to upregulated cellular GRP78 (type II hypersensitivity?). Both pathways could worsen vascular injury and atherosclerosis, leading to cardiac complications in COVID-19 cases with narrowed vessels.

microbiology↗

bioRxiv2026

Deciphering the Molecular Structure of the Type III Secretion System in Chlamydia trachomatis for Structure-Based Therapeutic Targeting

Chlamydia trachomatis is an obligate intracellular pathogen that causes sexually transmitted infections and trachoma. Its persistent forms show reduced antibiotic susceptibility, creating a need for new antivirulence strategies. The C. trachomatis injectisome or type III Secretion System (T3SS) injects effector proteins into host cells, yet the molecular structure of the complete apparatus is unknown. Here, we identify and validate all 13 T3SS constituent proteins using TXSSScan and reverse-BLAST analysis, and model and assemble them into the complete T3SS apparatus using an integrated computational pipeline. Template-based modeling revealed a conserved architecture despite low sequence identity (18-46%), and the assemblies were validated stereochemically and energetically against homolog controls. Targeting the oligomerization interface of the CdsN ATPase through structure-based virtual screening of the e-Drug3D and IMPPAT libraries, followed by ADMET filtering, including predicted membrane permeability for intracellular targeting, identified three repurposable FDA-approved candidates: M Roflumilast, Elacestrant, and Tecovirimat. MM-GBSA calculations and molecular dynamics simulations confirmed stable complexes with two adjacent CdsN monomers. Elacestrant showed the most favorable binding free energy. These results provide the first molecular-level map of the C. trachomatis T3SS and a rational basis for antivirulence drug repurposing. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=93 SRC="FIGDIR/small/723290v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@6893c8org.highwire.dtl.DTLVardef@1be5eddorg.highwire.dtl.DTLVardef@1462b25org.highwire.dtl.DTLVardef@1baadab_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2026

Structural Characterization of the Type IV Secretion System in Brucella melitensis for Virtual Screening-Based Therapeutic Targeting

Brucellosis is a globally important zoonotic disease caused by Brucella melitensis, the most virulent and clinically significant species affecting both humans and livestock. Unlike many Gram-negative pathogens, B. melitensis, a facultative intracellular pathogen, lacks conventional virulence factors and instead relies on specialized systems such as the Type IV Secretion System (T4SS) for secretion of effector proteins. In this study, an integrated computational pipeline was implemented to identify, model, and assemble the T4SS components, encoded by virB operon, from the complete B. melitensis proteome. Template-based modeling strategies were employed to generate structures of T4SS subcomplexes, referencing crystallographic data from E. coli T4SS. Structural superposition with E. coli homologs revealed highly conserved architecture despite only 30-50% sequence identity. Stereochemical validation confirmed high model quality and favorable interactions among most VirB protein pairs. Membrane insertion analysis of the membrane-embedded assemblies further corroborated the spatial orientation of the modeled T4SS. Potential of T4SS as a drug target was explored by targeting dimeric interface of VirB11 ATPase to disrupt protein-protein interactions that could disarm the pathogen. Virtual screening of compounds from DrugBank database revealed compounds with docking score [≤] -7.0 kcal/mol that were screened based on ADMET properties, yielding three promising candidates - Ezetimibe (Drug Id: DB00973), Chlordiazepoxide (Drug Id: DB00475), and Alloin (Drug Id: DB15477). MM-GBSA analysis estimated favorable binding free energies for these compounds and molecular dynamics simulation for 200 ns further confirmed the protein-ligand interaction stability. Collectively, these findings provide new insights into the architecture of B. melitensis T4SS and identify three potential drug molecules targeting T4SS. This supports FDA - approved drug repurposing as an effective strategy for anti-virulence therapy against Brucellosis. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=103 SRC="FIGDIR/small/706537v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@847c4borg.highwire.dtl.DTLVardef@1fc2551org.highwire.dtl.DTLVardef@f62a7corg.highwire.dtl.DTLVardef@15f3468_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2025

Molecular Configuration, Regulation and Function of Heterochannel Electrical Synapses

Neuron-specific expression of particular gap junction channel components defines the configuration and functional properties of electrical synapses. However, how a neuron utilises multiple, simultaneously expressed channel proteins - connexins or innexins -to make meaningful connections with distinct synaptic partners remains largely unknown. Using the posterior mechanosensory circuit in C. elegans, we discovered that individual electrical synapses can be formed by clustering together molecularly distinct gap junction channel-types made of three different innexin proteins, INX-1, UNC-7, and UNC-9. In this previously unknown configuration, which we term as heterochannel synapses, molecularly distinct gap junction channel types functionally collaborate to regulate posterior touch sensory behaviour, enhancing functional robustness. We show that the synaptic trafficking of the molecularly different channel types within a heterochannel synapse is independently regulated by discrete and conserved kinesin motor proteins, while distinct molecular pathways involving channel-specific retrograde kinesins regulate their turnover. These independent, channel-specific regulations also make individual synapse-level alterations in the composition of heterochannel synapses possible under altered environmental conditions, providing a novel mechanism for electrical synapse plasticity. Finally, we present evidence of heterochannel electrical synapses in C. elegans locomotory circuits and in the cerebellar Purkinje neurons of zebrafish larvae. Altogether, we demonstrate a novel heterochannel organization of electrical synapses, their regulation, and functional importance, which may be a conserved feature of metazoan nervous system.

neuroscience↗

bioRxiv2025

Identification and Characterization of Outer Membrane Proteins and Membrane Spanning Protein Complexes in Brucella melitensis

Brucellosis (Malta fever) is a zoonotic disease that affects both humans and animals, including cattle, sheep, and goats. Brucella melitensis is the most virulent and clinically significant species in humans. It is a Gram-negative bacterium with three groups of outer membrane proteins (OMPs): minor OMPs (Group 1), and major OMPs (Groups 2 and 3). OMPs with {beta}-barrel architecture play important roles in nutrient transport, efflux, adhesion, and membrane biogenesis. Despite their importance, the structure, function, and interaction dynamics of several B. melitensis {beta}-barrel OMPs and associated protein complexes remain mostly unexplored. In this study, we conducted a comprehensive in silico analysis to characterize known outer membrane {beta}-barrel (OMBB) proteins and identify novel OMBBs in B. melitensis 16M. Proteins were modelled using five computational tools: AlphaFold 3, ESMFold, SWISS-MODEL, RoseTTAFold, and TrRosetta. Outer-membrane insertion of the novel OMBBs was confirmed using PPM 3.0, Protein GRAVY, DREAMM, and MemProtMD_Insane. Putative functions were predicted using structure- and sequence-based annotations. Sequence variation across 46 B. melitensis strains were identified and mapped onto the structural models. OMBB-associated protein complexes - the RND (Resistance-Nodulation-Division) efflux pumps, the lipopolysaccharide transport (Lpt) complex, and the {beta}-barrel assembly machinery (BAM) complex - were modelled, and protein-protein interactions (PPIs) were analyzed to confirm thermodynamically stable assemblies. This study presents a robust in silico strategy for exploring OMP architecture and provides valuable structural insights to support the development of diagnostics, targeted therapeutics, and vaccines against B. melitensis.

bioinformatics↗

bioRxiv2025

A Mechanistic Whole Brain Model to Capture Simultaneous EEG-fMRI Data

This study introduces a novel oscillatory network model to simulate simultaneous EEG-fMRI data, addressing the reconstruction challenge that arises due to their contrasting spatiotemporal scales. Here, each brain region is modelled by two oscillator clusters - a cluster of low-frequency (LFO) and high-frequency Hopf oscillators (HFO) coupled with an innovative power-coupling rule, facilitating cross-frequency interactions. The model is trained in two stages: learning oscillators frequencies and phase relations using a biologically plausible complex-Hebbian rule in the first stage and, followed by a modified complex backpropagation for amplitude approximation, overcoming limitations of poor accuracy and computational complexity in existing models. This framework outperforms current methods in replicating empirical Functional Connectivity (FC), Functional Connectivity Dynamics (FCD), and modularity over disparate spatio-temporal scales. The correlation between the FC of fMRI and the FCs of various EEG frequency bands is reflected in the strengths of the LFO-HFO coupling. Furthermore, in-silico structural perturbation studies quantified the effect of pruning of the anatomical connectivity on spatiotemporal dynamics in terms of FC, FCD, modularity, and integration level (ISOR). The models ability to reconstruct simultaneous EEG-fMRI data showcases significant advancement in understanding the resting-state brains functionality from multimodal settings and deciphering neurological disorders in diverse spatiotemporal scales.

neuroscience↗

bioRxiv2025

Structural insights into the mechanism of C3 side chain fragmentation in cephalosporins with SME-1 class A carbapenemase

Cephalosporins remain one of the largest classes of antibiotics used clinically, with a diverse group of dual substituents at C3 and C7 positions. The fragmentation of the C-3 side chain from the C3 position after hydrolysis by Serine or metallo-{beta} lactamase leaving an exo-methylene group has been introduced in the literature recently, but the exact mechnism by which it happens remains unclear until now. Our incrystallo findings suggest that the fragmentation of the C3 side chain will depend on the C3 atom and its hybridization state. A C3 carbon with sp2 hybridization will not allow fragmentation, but a sp3 hybridized carbon will facilitate the detachment of the side chain. Furthermore, heteroatoms like Sulfur, on C3 position, bearing vacant d-orbitals participate in {pi}-cloud delocalization, making the detachment of C3 side chain energetically unfeasible. For the sp3 hybridized carbon at C3, the fragmentation of the bond was independent of the C3 atom. We observed fragmentation of the C3-C3 bond for all three heteroatoms, Nitrogen, Oxygen, and sulfur, at the C3 position. We validated our crystallographic findings with mass spectrometry to confirm the fragmentation of the C3 side chain in sp3 carbon and the unfragmented side chain in sp2 carbon and sulfur at C3 position. QM/MM simulations on three types of enzyme-cephalosporin complexes with different C3 side chains confirm the mechanism of fragmentation. Beyond revealing C3 side chain fragmentation in cephalosporins, useful for inhibitor and diagnostic design, our QM/MM methods are useful on incrystallo complexes to show the enzyme catalysis.

biochemistry↗

bioRxiv2025

Design of a Multi-Epitope Vaccine using β-barrel Outer Membrane Proteins Identified in Chlamydia trachomatis

Chlamydia trachomatis is an obligate intracellular Gram-negative pathogen responsible for causing sexually transmitted infections (STIs) and trachoma. Current interventions, including screening and antibiotics, are limited due to the widespread nature of asymptomatic infections, and the absence of licensed vaccine exacerbates the challenge. In this study, we predicted outer membrane {beta}-barrel (OMBB) proteins and designed a multi-epitope vaccine (MEV) construct using identified proteins. We employed a consensus-based computational framework on the C. trachomatis D/UW-3/CX proteome and identified 17 OMBB proteins, including well-known Pmp family members and MOMP. Eight OMBB proteins were computationally characterized, which showed significant structural homology with known outer membrane proteins from other bacteria. Sequence-based annotation tools were used to determine their putative functions. B-cell and T-cell epitopes were predicted from the selected proteins. The MEV construct was designed using four cytotoxic T lymphocyte (CTL) epitopes and 29 helper T lymphocyte (HTL) epitopes predicted from six OMBB proteins, which were conserved across 106 C. trachomatis serovars. The vaccine was supplemented at the N-terminus with Cholera enterotoxin subunit B and PADRE sequence to enhance its immunogenicity. The MEV construct of 780 amino acids was antigenic, non-allergenic, non-toxic, and soluble. Secondary structure analysis revealed 95% random coils. The 3D structural model of MEV was generated and validated, confirming its structural reliability. Molecular docking between MEV and Toll-like receptor 4 (TLR4) revealed strong and stable binding interactions, supporting its potential to elicit a strong immune response. This study highlights OMBB proteins as promising immunogenic targets and presents a computationally designed MEV candidate for C. trachomatis infection.

bioinformatics↗

bioRxiv2025

Investigation of the Cyanothece nitrogenase cluster in Synechocystis: a blueprint for engineering nitrogen fixing photoautotrophs

The nitrogenase gene cluster of unicellular diazotrophic cyanobacteria such as Cyanothece is frequently selected by nature for nitrogen fixing partnerships with eukaryotic phototrophs. The essential cluster components that confer an advantage in such partnerships remain under explored. To use this cluster for the development of synthetic, phototrophic nitrogen-fixing systems, a thorough and systematic analysis of its constituent genes is necessary. An initial effort to assess the possibility of engineering this cluster into non-diazotrophic phototrophs led to the generation of a Synechocystis 6803 strain with significant nitrogenase activity. In the current study, a refactoring approach was taken to determine the dispensability of the non-structural genes in the cluster and to define a minimal gene set for constructing a functional nitrogenase for phototrophs. Using a bottom-up strategy, the nif genes from Cyanothece 51142 were re-organized to form new operons. The genes were then seamlessly removed to determine their essentiality in the nitrogen fixation process. We demonstrate that besides the structural genes nifHDK, nifBSUENPVZTXW as well as hesAB are important for optimal nitrogenase function in a phototroph. We also show that optimal expression of these genes is crucial for efficient nitrogenase activity. Our findings provide a solid foundation for generating synthetic systems that will facilitate solar powered conversion of atmospheric nitrogen into nitrogen rich compounds, a stride towards a greener world. IMPORTANCEIntegrating nitrogen fixation genes into various photosynthetic organisms is an exciting strategy for converting atmospheric nitrogen into nitrogen rich products in a green and energy efficient way. In order to facilitate this process, it is essential that we understand the fundamentals of the functioning of a prokaryotic nitrogen fixing machinery in a non-diazotrophic, photoautotrophic cell. This study examines a nitrogenase gene cluster that has been naturally selected on multiple occasions for a nitrogen fixing partnership by eukaryotic photoautotrophs and provides a basic blueprint for designing a photosynthetic organism with nitrogen fixing ability.

synthetic biology↗

bioRxiv2025

Identification and Characterization of Novel Outer Membrane Proteins of Brachyspira pilosicoli

Brachyspira pilosicoli is a pathogenic, Gram-negative, spirochete bacterium that causes intestinal spirochetosis (IS) in birds, pigs, and humans and is distributed worldwide. This anaerobic intestinal bacterium colonizes the large intestine, potentially leading to colitis, diarrhoea, and decreased growth rate. Outer membrane proteins of Gram-negative bacteria play crucial roles in adhesion and host-pathogen interaction, helping the bacteria to evade the host immune system, and enhancing their virulence. However, B. pilosicoli outer membrane proteins are yet to be identified and characterized. Here, we report the computational discovery of 42 outer membrane {beta}-barrel (OMBB) proteins in B. pilosicoli proteome predicted using a consensus-based computational framework. {beta}-barrel architectures of the predicted proteins were confirmed by generating AlphaFold 3-based structural models. Structure-based functional annotation predicted putative functions for the identified OMBB proteins, including BamA homolog involved in folding and membrane insertion of OMPs, LptD homolog involved in transport of lipopolysaccharides into the OM, efflux pumps, transporters, enzymes, diffusion channels, and porins. Sequence variations across nine strains of B. pilosicoli were identified and mapped onto structural models, revealing that many of the variations were present on the surface exposed loop regions of the {beta}-barrel structures. Our in-silico analysis has identified 42 OMBB proteins, including homologs of BamA, LptD, TolC, TonB-dependent receptors, CsgG. Seven of these were identified as hypothetical proteins. Computational characterization of the predicted OMBB proteins offer insights into their potential roles in physiology, virulence, and disease pathogenesis, highlighting their potential for applications in diagnostics, vaccine development, or therapeutic interventions.

bioinformatics↗

bioRxiv2024

Age-induced BMP signaling inhibits the ICAT-mediated Wnt pathway to promote osteoarthritis.

Osteoarthritis (OA) is a highly prevalent and debilitating musculoskeletal disorder that affects billions of aging individuals worldwide, causing chronic pain, impaired mobility, and a substantial decline in quality of life, yet effective disease-modifying therapies remain unavailable. Moreover, existing experimental models, however, inadequately capture the complex, age-related factors that contribute to OA pathogenesis, limiting our understanding of underlying molecular mechanisms and hindering therapeutic development. To address this gap, we examined mouse articular cartilage (AC) during natural aging, without surgical or chemical intervention. In young AC, BMP ligand expression is restricted to the bone-cartilage junction, limiting BMP signaling despite widespread BMPR1A receptor expression, while Wnt/{beta}-catenin signaling predominates in the outer layers. With aging, BMP ligand expression expands throughout the cartilage, leading to widespread activation of BMP signaling, which induces ICAT expression, a known inhibitor of Wnt/{beta}-catenin signaling. This elevated BMP signaling triggers chondrocyte hypertrophy while concurrently suppressing Wnt/{beta}-catenin activity. Moreover, genetic or pharmacological activation of BMP signaling in young AC recapitulates the cellular and molecular features of aged cartilage, independent of chronological age. Collectively, our findings demonstrate that an age-dependent shift in BMP-Wnt signaling disrupts AC homeostasis and drives OA progression in mice. Author SummaryDespite the prevalence of osteoarthritis in aging individuals, there are no effective treatments that can stop or reverse disease progression. In this study, we investigated how joint cartilage changes during natural aging in mice. We observed that healthy, young articular cartilage exhibits robust Wnt/{beta}-catenin with minimal BMP siganling. With age, this balance shifts, with BMP signaling becoming dominant, coupled with reduced Wnt/{beta}-catenin activity. This disruption of Wnt-BMP homeostasis induces the pathogenesis of OA. Importantly, local inhibition of BMP signaling significantly alleviates the severity of the disease. Overall, these results show that age-associated changes in key signaling pathways drive OA progression and suggest potential targets for therapeutic intervention. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/616036v2_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@b2be12org.highwire.dtl.DTLVardef@dcb1f3org.highwire.dtl.DTLVardef@126fc40org.highwire.dtl.DTLVardef@7af96c_HPS_FORMAT_FIGEXP M_FIG C_FIG

pathology↗

bioRxiv2024

Genome streamlining to improve performance of a fast-growing cyanobacterium Synechococcus elongatus UTEX 2973

Cyanobacteria are photosynthetic organisms that have garnered significant recognition as potential hosts for sustainable bioproduction. However, their complex regulatory networks pose significant challenges to major metabolic engineering efforts, thereby limiting their feasibility as production hosts. Genome streamlining has been demonstrated to be a successful approach for improving productivity and fitness in heterotrophs but is yet to be explored to its full potential in phototrophs. Here we present the systematic reduction of the genome of the cyanobacterium exhibiting the fastest exponential growth, Synechococcus elongatus UTEX 2973. This work, the first of its kind in a photoautotroph, involved an iterative process using state-of-the-art genome-editing technology guided by experimental analysis and computational tools. CRISPR/Cas3 enabled large, progressive deletions of predicted dispensable regions and aided in the identification of essential genes. The large deletions were combined to obtain a strain with 55 kb genome reduction. The strains with streamlined genome showed improvement in growth (up to 23%) and productivity (by 22.7%) as compared to the WT. This streamlining strategy not only has the potential to develop cyanobacterial strains with improved growth and productivity traits but can also facilitate a better understanding of their genome to phenome relationships. ImportanceGenome streamlining is an evolutionary strategy used by natural living systems to dispense unnecessary genes from their genome as a mechanism to adapt and evolve. While this strategy has been successfully borrowed to develop synthetic heterotrophic microbial systems with desired phenotype, it has not been extensively explored in photoautotrophs. Genome streamlining strategy incorporates both computational predictions to identify the dispensable regions and experimental validation using genome editing tool and in this study we have employed a modified strategy with the goal to minimize the genome size to an extent that allows optimal cellular fitness under specified conditions. Our strategy has explored a novel genome-editing tool in photoautotrophs which, unlike other existing tools, enables large, spontaneous optimal deletions from the genome. Our findings demonstrate the effectiveness of this modified strategy in obtaining strains with streamlined genome, exhibiting improved fitness and productivity.

synthetic biology↗

bioRxiv2023

Suppressive might of a few: T follicular regulatory cells impede auto-reactivity despite being outnumbered in the germinal centres

The selection of high-affinity B cells and the production of high-affinity antibodies are mediated by T follicular helper cells (Tfhs) within germinal centres (GCs). Therein, somatic hypermutation and selection enhance B cell affinity but risk the emergence of self-reactive B cell clones. Despite being outnumbered compared to their helper counterpart, the ablation of T follicular regulatory cells (Tfrs) results in enhanced dissemination of self-reactive antibody-secreting cells (ASCs). The specific mechanisms by which Tfrs exert their regulatory action on self-reactive B cells are largely unknown. We developed computer simulations to investigate how Tfrs regulate either selection or differentiation of B cells to prevent auto-reactivity. We observed that Tfr-induced apoptosis of self-reactive B cells during the selection phase impedes self-reactivity with physiological Tfr numbers, especially when Tfrs can access centrocyte-enriched GC areas. While this aided in selecting non-self-reactive B cells by restraining competition, higher Tfr numbers distracted non-self-reactive B cells from receiving survival signals from Tfhs. Thus, the location and number of Tfrs must be regulated to circumvent such Tfr distraction and avoid disrupting GC evolution. In contrast, when Tfrs regulate differentiation of selected centrocytes by promoting recycling to the dark zone phenotype of self-reactive GC resident pre-plasma cells (GCPCs), higher Tfr numbers were required to impede the circulation of self-reactive ASCs (s-ASCs). On the other hand, Tfr-engagement with GCPCs and subsequent apoptosis of s-ASCs can control self-reactivity with low Tfr numbers, but does not confer selection advantage to non-self-reactive B cells. The simulations predict that to restrict auto-reactivity, natural redemption of self-reactive B cells is insufficient and that Tfrs should increase the mutation probability of self-reactive B cells.

immunology↗

bioRxiv2023

Emergence of distributed working memory in a human brain network model

Working memory is a fundamental cognitive function which allows to transiently store and manipulate relevant information in memory. While it has been traditionally linked to activity in specific prefrontal cortical areas, recent electrophysiological and imaging evidence has shown co-occurrent activities in different brain regions during working memory. To dissect the mechanisms behind the emergence of such distributed working memory activity in the human brain, we built and analyzed a detailed, data-constrained model of the human brain combining multiple sources of large-scale brain data. The model reveals the emergence of distributed working memory patterns which rely on long-range synaptic projections, differing from classical models where working memory emerges due to local recurrent connectivity. When compared to existing evidence regarding areas linked to working memory, our model explains almost 80% of the observed variability across brain areas. Our results predict important differences between the distributed working memory in human and non-human primates, and propose a potential solution for the debate regarding the role of early visual areas in working memory encoding. This work provides a novel approach to simulate the human brain which combines the richness of large-scale brain dynamics with the insight of mechanistic models of cognitive functions.

neuroscience↗