Biology subjects

Garg, P.

Publications and source records attributed to Garg, P..

At least 19 recordsLinked to original sources

bioRxiv2026

Evolutionarily Conserved Plasmodium falciparum HSP70-3 Enhances the Immunogenicity of the Established Malaria Vaccine Antigen Circumsporozoite Protein through Cross-Presentation

Background: Clinically approved P. falciparum circumsporozoite protein (PfCSP)-based pre-erythrocytic stage malaria vaccines suffer from sub-optimal efficacy and rapidly waning memory. By decoding these limitations, we fast-tracked vaccine optimization by correcting CSPs fragile immunogenicity using a parasite-derived chaperone. Unlike other orthologues of P. falciparum heat shock protein 70 (PfHSP70), substrate-binding domain (SBD) of PfHSP70-3 is an evolutionarily conserved internal mitochondrial matrix chaperone. We demonstrate that PfHSP70-3-SBD hydrophobic region complexes with PfCSP and targets host CD91 on antigen-presenting cells to trigger cross-presentation, resulting in enhanced immune responses to PfCSP. Methods: In silico analyses predicted interactions among PfHSP70-3-SBD, PfCSP, and CD91 receptor. Recombinant PfHSP70-3-SBD and PfCSP proteins were purified and assessed for in vitro interaction using ELISA and far-western dot blot assays. BALB/c mice were immunized with alum-formulated PfCSP, with or without PfHSP70-3-SBD. PfCSP-specific IgG and IFN-{gamma} expression were evaluated by ELISA and RT-qPCR, respectively. Functional activity was assessed using transgenic Plasmodium berghei expressing PfCSP (Pb/Pf CSP), by sporozoite neutralization assay. Findings: PfHSP70-3-SBD demonstrated a specific interaction with PfCSP. The complex primarily engages CD91 receptor cluster IV, providing a potential mechanistic basis for antigen cross-presentation. Incorporation of PfHSP70-3-SBD enhanced PfCSP-specific total IgG responses and increased IFN-gamma expression in BALB/c mice, indicating improved immunogenicity upon co-formulation. Notably, immune sera neutralized Pb/Pf CSP sporozoites, delaying blood-stage patency until day 12 in 40% of C57BL/6 mice, with no detectable parasitemia in surviving mice for up to 2 months, indicating sustained efficacy against the sporozoite stage. Interpretation: Molecular chaperones, including HSPs, are currently under intense investigation for their immune-potentiating activity, leading to next-generation vaccine development. By utilizing chaperonic domains derived directly from the pathogenic agent itself to serve an essential adjuvant function, this approach reduces reliance on heterologous adjuvants while enhancing antigen-specific immunity. Funding: This work was supported by funding from Indian Council of Medical Research (IIRPSG-2024-01-03940). Keywords: Substrate-binding domain, PfHSP70-3, Adjuvant, Cross-presentation, Pre-erythrocytic stage, Plasmodium falciparum

microbiology↗

bioRxiv2026

Single-Cell Omics to Uncover Cell-Type-Specific Gene Regulatory Networks Driving Mucilage Production in Specialized Roots

Aerial roots of certain maize (Zea mays) and sorghum (Sorghum bicolor) genotypes secrete a carbohydrate-rich mucilage that sustains nitrogen-fixing bacteria, and the underground roots of most plants secrete mucilage at their tips. In both settings, border cells released from the root cap produce this mucilage, yet the genes controlling their formation remain largely unknown. We sequenced the RNA of about 23,000 nuclei from mucilage-secreting roots of maize, sorghum, and Medicago truncatula, sampled before and after secretion. Each species yielded 10 to 15 clusters, which we annotated with published markers and with new markers that we validated by in situ hybridization in the two cereals and by promoter-reporter fusions in M. truncatula. We then inferred modular and cell type-specific gene regulatory networks. Transcriptional programs diverged across the three species except in the quiescent center and the meristem, and in the clusters containing border cells, where a shared set of regulators emerged. Silencing three of five candidate regulators reduced border cell number, while silencing a partially overlapping set of three regulators reduced root diameter in M. truncatula. Silencing the C2H2 zinc finger protein MtrunA17_Chr1g0148421 also reduced border cell viability. A maize insertion mutant of bzip89 released fewer border cells than the wild type. These networks offer entry points for tuning mucilage production in cereal and legume crops.

plant biology↗

bioRxiv2026

Integrative Cross-Cohort Meta-Analysis Reveals a Conserved Dysbiotic Signature of Streptococcaceae and Lachnospiraceae in Multiple Sclerosis

BackgroundMultiple Sclerosis (MS) is a chronic autoimmune disorder characterized by inflammation and demyelination in central nervous system (CNS). Although increasing evidence suggests that gut microbial dysbiosis contributes to MS pathogenesis through the microbiota-gut-brain axis, reproducible microbial signatures associated with disease progression across independent clinical cohorts remain incompletely characterized. ObjectiveThis study aimed to identify conserved gut microbial alterations associated with Multiple Sclerosis by integrating publicly available human gut microbiome datasets and characterizing disease-associated microbial signatures linked to immune dysregulation. DesignHuman gut metagenomic 16S rRNA sequencing data from MS patients and healthy controls obtained from publicly available repositories (NCBI, Bioproject). Raw sequencing reads were processed using a standardized microbiome analysis workflow, including quality control, denoising, taxonomic assignment, phylogenetic reconstruction, diversity analyses, and differential abundance testing. Microbial community structure was evaluated using alpha- and beta-diversity analyses, while statistically significant differences between study groups were assessed using PERMANOVA, Kruskal-Wallis, and ANCOM to identify disease-associated bacterial taxa. ResultsIntegration of independent cohorts revealed consistent alterations in the gut microbial composition of MS patients compared with healthy controls. Significant reductions in microbial diversity and distinct microbial community structures were observed in MS. Differential abundance analysis demonstrated enrichment of the pro-inflammatory family Streptococcaceae, whereas beneficial short-chain fatty acid-producing taxa, particularly Lachnospiraceae, were significantly depleted in MS patients. These conserved microbial alterations indicate disruption of immune-regulatory bacterial communities and support the involvement of gut microbial dysbiosis in MS-associated neuroinflammation. ConclusionThis study identifies a reproducible gut microbial dysbiosis signature associated with Multiple Sclerosis, characterized by expansion of pro-inflammatory bacterial taxa and depletion of beneficial SCFA-producing microorganisms. These findings strengthen the evidence supporting the microbiota-gut-brain axis in MS pathogenesis and highlight microbial community signatures that may contribute to future biomarker development and microbiome- based therapeutic strategies.

microbiology↗

bioRxiv2026

Ancient human genomes from Ladakh reveal Tibetan, South Asian, and Central Asian admixture over the last three millennia

The trans-Himalayan region of Ladakh occupies a strategic position at the crossroads of South Asia, the Tibetan Plateau, and Central Asia, with archaeological evidence pointing to long-term cultural exchanges across these regions. However, the human genetic history of Ladakh remains largely unexplored. We generated paleogenomic data from seven individuals recovered from two sites in Western Ladakh - the Old Lady Spider Cave and Hanu - of which six are dated to 531-585 CE and one to the 19th century CE. The older individuals share substantial genetic ancestry with Tibetan groups but also harbor major contributions from two additional sources: one corresponding to the currently-oldest observation of the Ancestral North Indian genetic component that characterizes several present-day populations in North India and Pakistan, and another related to ancient Central Asian groups, with admixture events occurring between [~]2,100-2,500 years ago. In contrast, the later individual falls within a previously described ancient northern Himalayan genetic cline based on [~]1,100-1,300-year-old individuals from Himachal Pradesh, with ancestries related to ancient Tibetan and Steppe-related sources. Stable isotope analysis suggests that these individuals were local to Ladakh in late life and practiced an agro-pastoralist subsistence. Our study establishes that Ladakhs central role in Eurasian economic and socio-cultural networks was shaped by dynamic and sustained gene flow linking high-altitude Himalayan groups with both lowland South Asia and Inner Asia.

genetics↗

bioRxiv2025

Structural Studies of Nedicistrovirus IRES-Driven, Initiation Factor-independent Translation Shed Light on Key Steps of Eukaryotic Translation Elongation

We utilized the Nedicistrovirus (NediV) intergenic region (IGR) IRES-mediated, initiation factor-independent translation initiation system and determined high-resolution structures of 80S ribosome complexes with the NediV IRES in various functional states, including binary complexes, aminoacyl-tRNA-bound complexes, and complexes with elongation factor eEF2. In binary complexes, the NediV IRES primarily occupies the ribosomal P site, exhibiting conformational flexibility and engaging the ribosome at multiple interaction sites. Upon translocation, the IRES undergoes structural rearrangements, including destabilization of its PKI domain, facilitating the transition to canonical elongation. Crucially, we captured an eEF2-bound complex, along with an eEF1A-bound post-proofreading complex featuring a mismatched tRNA, the latter representing the first instance of a canonical elongation complex visualized in the presence of a natural, hydrolysable nucleotide and without the addition of any trapping agents. These findings provide a comprehensive structural overview of IGR IRES-mediated translation initiation and its transition to elongation, revealing key mechanistic details of viral translation and proofreading.

biochemistry↗

bioRxiv2025

Transcriptome Network Biology Reveals the Neuroprotective Potential of Rosmarinic Acid Against Frontotemporal Dementia

Frontotemporal dementia (FTD) is a neurodegenerative disorder characterised by impaired behaviour and language. It affects individuals between 45 and 65 years of age. The present study aims to identify potential phytochemicals against target receptor of FTD. Transcriptome data analysis guided computer-aided drug design (CADD) was conducted to investigate molecular interactions between active phytochemicals present in Indian spices and differentially expressed genes. In present study, samples for patients with FTD were obtained from GEO dataset [GSE92340]. The samples were pre-processed followed by differential gene expression analysis using DESeq2. The patient sample was analyzed for 1882 upregulated genes. Hub gene out of DEGs was identified using CytoHubba Plugin of Cytoscape based on eleven parameters (Degree, Edge Percolated Component (EPC), Maximum Neighbourhood Component (MNC), Density of Maximum Neighbourhood Component (DMNC), Maximal Clique Centrality (MCC), Bottleneck, EcCentricity, Closeness, Radiality, Betweenness, and Stress). HDAC1 gene was identified as hub gene and key target in case of FTD. CADD pipeline was used to identify phytochemicals against HDAC1 gene. A list of eighty-one phytochemicals found in Indian spices was analyzed for their drug likeness abilities using molinspiration and toxicity prediction using ProTox-II, which identified twenty-three phytochemicals as safe drug-like compounds. PatchDock was employed to examine interactions between phytochemical compounds and HDAC1. Molecular interactions revealed rosmarinic acid derived from rosemary (Salvia rosmarinus) as the most effective drug for FTD having lowest ACE value of -395 kcal/mol. In conclusion, this study demonstrates essential role of HDAC1 in pathogenesis of FTD and demonstrates the neuroprotective potential of rosmarinic acid against FTD.

bioinformatics↗

bioRxiv2024

Evaluation of Immunopharmacological Efficacy of Live Leishmania donovani Overexpressing Ld_ζ1domain for Protection Against Experimental Human Visceral Leishmaniasis

ObjectiveTo evaluate the efficacy and immunogenicity of the zeta domain over-expressing Leishmania donovani (Ld_{zeta}1domain) as a vaccination candidate against visceral leishmaniasis (VL). MethodsIn this study, Leishmania overexpressor Ld_{zeta}1domain (OE) were transformed by electroporation using a GFP-tagged Ld_{zeta}1domain recombinant plasmid. The resulting overexpressing cells were analysed in vitro to assess their growth dynamics and infectivity. We also investigated the immune-protective potential of these overexpressor in a mouse model challenged with Leishmania donovani. The immune response, including Th1 and Th2 pathways, was thoroughly characterized using RT-PCR and ELISA assays. In addition, the study conducted a thorough evaluation of the mouses spleen and liver parasites, as well as quantitative evaluation of tissue pathological changes. ResultsLd_{zeta}1 domain (OE) parasites exhibited significantly lower viability and replication rates than WT parasites, and in vivo studies showed that mice immunized with the Ld_{zeta}1(OE) domain had lower parasite numbers than mice infected with LdWT. Spleen and liver showed significant histological changes suggestive of protection. Parasite burden in the spleen and liver of vaccinated mice were significantly reduced. The immune response showed increased IFN-{gamma} levels and lower IL-10 production, resulting in a greater IFN-{gamma}/IL-10 ratio, indicating parasite elimination. The vaccination also caused a significant IgG humoral response and increased nitric oxide production in immunized mice. ConclusionOur findings demonstrated that overexpressing the zeta toxin resulted in controlled parasite attenuation, lowering pathogenicity while retaining immunogenic features. Our work established the zeta over-expressors protective efficacy, immunogenicity, and proliferation in response to a Leishmania challenge in vitro and in vivo. This preliminary prototype study suggested that Ld_{zeta}1domain (OE) parasites may be suitable for developing an attenuated vaccine against leishmaniasis. Graphical representation O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=88 SRC="FIGDIR/small/630021v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@32dfc0org.highwire.dtl.DTLVardef@1ffe89dorg.highwire.dtl.DTLVardef@d9feorg.highwire.dtl.DTLVardef@1e7bae7_HPS_FORMAT_FIGEXP M_FIG C_FIG A schematic representation on the protective effectiveness, immunogenicity, and proliferation of the zeta over-expressor in response to Leishmania challenge in vitro and in vivo model.

pathology↗

bioRxiv2024

Transcriptomic analysis reveals vector attraction to potato virus Y is mediated through temporal regulation of TERPENE SYNTHASE 1 (TPS1)

Virus-plant dynamics change over time, influencing interactions between plants and insect vectors. However, the signaling pathways and regulators that control these temporal responses remain largely unknown. In this study, we used insect performance and preference bioassays, RNA-Seq, and genetic tools to identify underlying mechanisms mediating temporal variation in plant-virus-vector interactions. We show that settlement and fecundity of the aphid vector, Myzus persicae, is increased on potato virus Y (PVY)-infected Nicotiana benthamiana plants two weeks after inoculation but not after six weeks. RNA-Seg analysis revealed transcripts related to plant defense and amino acid biosynthesis are upregulated in response to PVY infection and down regulated in response to aphid herbivory, and these patterns changed over time. Based on this analysis we identified a sesquiterpene synthase gene, terpene synthase 1 (NbTPS1), that is upregulated early in PVY infection, but not at later infection time points. Using virus-induced gene silencing and transient overexpression in N. benthamiana we demonstrate that PVY induction of NbTPS1 is required for increased aphid attraction to PVY-infected plants in the early stages of infection. Taken together, PVY temporally regulates transcriptional pathways related to plant defense responses and volatile organic compounds that influence aphid vector performance and preference.

plant biology↗

bioRxiv2024

Passage through micro-sprayer increases functional activity-implications for activity assays in time-resolved cryo-EM

This study examines the validity of an assay that is used to report on the retainment of functional competence by ribosomes as they pass a micro-sprayer. We find a reproducible increase, rather than the expected decrease in GFP production as monitored by fluorescence, which suggests heterogeneity or partial aggregation of ribosomes in solution. An even larger increase in functional activity is observed when sonication is used, pointing to mechanical agitation as the decisive factor in both scenarios. The results have a bearing on the design and interpretation of validation experiments in time-resolved cryo-EM based on microfluidic chips.

biophysics↗

bioRxiv2024

Identifying patterns differing between high-dimensional datasets with generalized contrastive PCA

High-dimensional data have become ubiquitous in the biological sciences, and it is often desirable to compare two datasets collected under different experimental conditions to extract low-dimensional patterns enriched in one condition. However, traditional dimensionality reduction techniques cannot accomplish this because they operate on only one dataset. Contrastive principal component analysis (cPCA) has been proposed to address this problem, but it has seen little adoption because it requires tuning a hyperparameter resulting in multiple solutions, with no way of knowing which is correct. Moreover, cPCA uses foreground and background conditions that are treated differently, making it ill-suited to compare two experimental conditions symmetrically. Here we describe the development of generalized contrastive PCA (gcPCA), a flexible hyperparameter-free approach that solves these problems. We first provide analyses explaining why cPCA requires a hyperparameter and how gcPCA avoids this requirement. We then describe an open-source gcPCA toolbox containing Python and MATLAB implementations of several variants of gcPCA tailored for different scenarios. Finally, we demonstrate the utility of gcPCA in analyzing diverse high-dimensional biological data, revealing unsupervised detection of hippocampal replay in neurophysiological recordings and heterogeneity of type II diabetes in single-cell RNA sequencing data. As a fast, robust, and easy-to-use comparison method, gcPCA provides a valuable resource facilitating the analysis of diverse high-dimensional datasets to gain new insights into complex biological phenomena.

neuroscience↗

bioRxiv2024

Reproductive success of inbred strain MV31 of the ctenophore Mnemiopsis leidyi in a self-sustaining inland laboratory culture system

Ctenophores are an attractive lineage for studying animal evolution due to their early divergence from other metazoans. Among Ctenophora, Mnemiopsis leidyi is a model system for developmental, cellular, molecular genetic, and evolutionary studies. Until recently, many of these studies were conducted on wild-caught animals, limiting access to researchers on the coast. Here we present significant advancements towards culturing M. leidyi in laboratories without coastal access, enabling its wider use as an experimental and genetic model system. We detail updated feeding regimes that take advantage of co-culturing Brachionus rotifers with Apocyclops copepods, and quantify the reproductive output of our M. leidyi lab strain on this diet. Our updated feeding regime maintains reproductive fitness comparable to wild-caught individuals. Importantly, we have eliminated the logistical complexities and costs of regularly feeding live larval fish to M. leidyi. Our updated protocols make it feasible to maintain continuous ctenophore cultures independent of access to both coastal populations of wild M. leidyi and larval fish culturing facilities.

zoology↗

bioRxiv2023

Mixed signatures for subcritical dynamics in rodent hippocampus during sleep and awake epochs

Neuronal dynamics such as brain criticality have recently been attributed to optimal information processing. Brain criticality attempts to elucidate the collective dynamics of a large number of neurons. It posits that the brain operates near critical to the critical point, although the field is rife with controversies and contrasting evidence. Similar computational capacities are observed during sharp wave ripples in the hippocampus prompting the need to correlate their dynamics. In the current study, the measures of avalanche criticality including neuronal avalanches, branching process, crackling noise relation, and deviation from criticality coefficient and Hurst exponents for long-range temporal correlations in rodent hippocampus during sharp wave ripples are reported. The evidence for mixed subcritical to critical dynamics in the hippocampus and minimal difference between ripple and no ripple times across measured metrics was found. The evidence demonstrates heterogeneity in signatures of criticality among animals and brain areas, indicating the presence of broad-range neuronal dynamics.

neuroscience↗

bioRxiv2023

Computational Modeling of Hyperpolarizing Astrocytic Influence on Cortical Up-Down State Transitions

The Up-Down dynamics seen in cortical structures during non-rapid-eye-movement (NREM) sleep, anesthetized states, and quiet wakefulness is the spontaneous alternation between phases of heightened firing activity (referred to as the Up state) and periods of neuronal inactivity (termed the Down state) within neural networks. By leveraging bistable dynamics imposed by a depolarising astrocyte population, in the current paper, we introduced a hyperpolarising astrocyte population to an existing model of Up-Down dynamics to account for biological relevance. We created a computational rate model that includes populations of depolarizing and hyperpolarizing astrocytes and neurons. To optimize model parameters, we used the Elementary Effects (EE) test. It was followed by linear stability analysis to locate bistable regimes in the parameter hyperspace. The addition of hyperpolarizing gliotransmission perturbed model dynamics, indicating its sensitivity to qualitatively differing architectures. We then identified a bistable regime within the dynamics spectrum. According to the EE test, the strength of cell population coupling is a low-sensitivity parameter, possibly due to neuroplastic changes. We also found that the threshold of excitatory cell populations and the strength of adaptation are high-sensitivity parameters, whereas the threshold of inhibitory cell populations is low-sensitivity. Our model enables the possibility of testing biologically relevant theories of hyperpolarizing gliotransmission, where data remains scant.

neuroscience↗

bioRxiv2023

Host inducible-HSP70A1A is an irresistible drug target to combat SARS-CoV-2 infection and pathogenesis

One of the fundamental mechanisms developed by the host to contain the highly infectious and rapidly proliferating SARS coronavirus is elevation of body temperature, a natural fallout of which is Heat Shock Protein (HSP) over-expression. Here, for the first time, we demonstrate that the SARS-CoV-2 virus exploits the host Hsp70 chaperone for its entry and propagation and blocking it can combat the infection. SARS-CoV-2 infection as well as febrile temperature enhanced Hsp70 overexpression in host Vero E6 cells. In turn, Hsp70 overexpression elevated the host cell autophagic response that is a prerequisite for viral propagation. Suppressive and prophylactic treatment of Vero E6 cells with HSP70 inhibitor PES-Cl, a small molecule derivative of Pifithrin , abrogated viral infection more potently than the currently used drug Remdesivir by suppressing host HSP70 and autophagic response. In conclusion, our study not only provides a fundamental insight into the role of host Hsp70 in SARS-CoV-2 pathogenesis, it paves the way for the development of potent and irresistible anti-viral therapeutics.

microbiology↗

bioRxiv2023

Predicting the feasibility of targeting a conserved region on the S2 domain of the SARS-CoV-2 spike protein

The efficacy of vaccines against the SARS-CoV-2 virus significantly declines with the emergence of mutant strains, prompting investigation into the feasibility of targeting highly conserved but often cryptic regions on the S2 domain of spike protein. Using tools from molecular dynamics, we find that this conserved S2 epitope located in the central helices below the receptor binding domains is unlikely to be exposed by dynamic fluctuations without any external facilitating factors, in spite of previous computational evidence suggesting transient exposure of this region. Furthermore, glycans inhibit opening dynamics, and thus stabilize spike in addition to immunologically shielding the protein surface, again in contrast to previous computational findings. Though the S2 epitope region examined here is central to large scale conformational changes during viral entry, free energy landscape analysis obtained using the path coordinate formalism reveals no inherent "loaded spring" effect, suggesting that a vaccine immunogen would tend to present the epitope in a pre-fusion-like conformation and may be effective in neutralization. These findings contribute to a deeper understanding of the dynamic origins of the function of the spike protein, as well as further characterizing the feasibility of the S2 epitope as a therapeutic target.

biophysics↗

bioRxiv2023

Genomic perspectives of SARS CoV-2 in liver disease patients with its clinical correlation: A single centre retrospective study

BackgroundSevere Acute Respiratory Syndrome Coronavirus-2 (SARS CoV-2), is a causative agent of current global pandemic of Coronavirus disease-19 (COVID-19). Due to propagated outbreak and global vaccination drive an immense immunological selection pressure has been exerted on SARS CoV-2 leading to evolution of new variants. This study was performed to compare the mutational and clinical profile of liver disease patients infected with different variants of SARS CoV-2. MethodologyThis was a single-centre, retrospective, cohort study in which clinicogenomic analysis of liver disease (LD) patients infected with SARS CoV-2 was performed. Complete demographic and clinical details were retrieved from Hospital Information System (HIS). QC-threshold passed FASTA files containing sequences from COVID-19 patients (n=174) were compared with a reference genome of SARS-CoV-2 isolate named Wuhan-Hu-1 (NCBI Reference Sequence: NC_045512.2) for mutational analysis. ResultsOut of 232 finally analysed patients 137 (59.1%) were LD-CoV (+) and 95 (40.9%) were LD-CoV(-). LD patients with comorbidities were affected more with COVID-19 (p=0.002). On comparing the outcome in the terms of mortality, LD-CoV (+) had 2.29 times (OR 2.29, CI 95%, 1.25-4.29) higher of odds of succumbing to COVID-19 (p=0.006). Multivariate regression analysis revealed, abdominal distention (p=0.05), severe COVID-19 pneumonia (p=0.046) and the change in serum bilirubin levels (p=0.005) as well as Alkaline phosphatase (ALP) levels (p=0.003) to have an association with adverse outcome in LD patients with COVID-19. In Delta (22%) and Omicron (48%) groups, Spike gene harboured maximum mutations. On comparing the mutations between LD-CoV(+/D) and LD-CoV(+/O) a total of nine genes had more mutations in LD-CoV(+/O) whereas three genes had more mutations in LD-CoV(+/D). ConclusionWe concluded that LD patients are more susceptible to COVID-19 as compared to a healthy adult with associated adverse clinical outcomes in terms of mortality and morbidity. Therefore this special group should be given priority while devising and introducing new vaccination and vaccination policies. The infection with different variants did not result in different outcome in our group of patients.

microbiology↗

bioRxiv2023

Cryo-EM reveals the mechanism of DNA compaction by Mycobacterium smegmatis Dps2

DNA-binding protein under starvation (Dps), is a miniature ferritin complex which plays a vital role in protecting bacterial DNA during starvation for maintaining the integrity of bacteria from hostile conditions. Mycobacterium smegmatis is one such bacteria that express MsDps2, which binds DNA to protect it under oxidative and nutritional stress conditions. Several approaches, including cryo-electron tomography (Cryo-ET), were implemented to identify the structure of the Dps protein that is bound to DNA. However, none of the structures of the Dps-DNA complex was resolved to high resolution to be able to identify the DNA binding residues. In this study, we implemented various biochemical and biophysical studies to characterize the DNA protein interactions of Dps protein. We employed single-particle cryo-EM-based structural analysis of MsDps2-DNA and identify that the region close to N-terminal confers the DNA binding property. Based on cryo-EM data, we performed mutations of several arginine residues proximal to DNA binding region, which dramatically reduced the MsDps2-DNA interaction. In addition, we demonstrated the proposed model for DNA compaction during lattice formation. We also pinpointed arginine residues, which are responsible for DNA binding in lattice arrangement of MsDps2. We performed single-molecule imaging experiments of MsDps2-DNA interactions that corroborate well with our structural studies.

biophysics↗

bioRxiv2022

Ancestral SARS-CoV-2 driven antibody repertoire diversity in an unvaccinated individual correlates with expanded neutralization breadth

Understanding the quality of immune repertoire triggered during natural infection can provide vital clues that form the basis for development of humoral immune response in some individuals capable of broadly neutralizing pan SARS-CoV-2 variants. We assessed the diversity of neutralizing antibody responses developed in an unvaccinated individual infected with ancestral SARS-CoV-2 by examining the ability of the distinct B cell germline-derived monoclonal antibodies (mAbs) in neutralizing known and currently circulating Omicron variants by pseudovirus and authentic virus neutralization assays. The ability of the antibodies developed post vaccination in neutralizing Omicron variants was compared to that obtained at baseline of the same individual and to those obtained from Omicron breakthrough infected individuals by pseudovirus neutralization assay. Broadly SARS-CoV-2 neutralizing mAbs representing unique B cell lineages with non-overlapping epitope specificities isolated from a single donor varied in their ability to neutralize Omicron variants. Plasma antibodies developed post vaccination from this individual demonstrated neutralization of Omicron BA.1, BA.2 and BA.4 with increased magnitude and found to be comparable with those obtained from other vaccinated individuals who were infected with ancestral SARS-CoV-2. Development of B cell repertoire capable of producing antibodies with distinct affinity and specificities for the antigen immediately after infection capable of eliciting broadly neutralizing antibodies offers highest probability in protecting against evolving SARS-CoV-2 variants. ImportanceDevelopment of robust neutralizing antibodies in SARS-CoV-2 convalescent individuals is known, however varies at population level. We isolated monoclonal antibodies from an individual infected with ancestral SARS-CoV-2 in early 2020 that not only varied in their B cell lineage origin but also varied in their capability and potency to neutralize all the known VOC and currently circulating Omicron variants. This indicated establishment of unique lineages that contributed in forming B cell repertoire in this particular individual immediately following infection giving rise to diverse antibody responses that could compensate each other in providing broadly neutralizing polyclonal antibody response. Individuals who were able to produce such potent polyclonal antibody responses after infection have a higher chance of being protected from evolving SARS-CoV-2 variants.

immunology↗