Biology subjects

Parveen, N.

Publications and source records attributed to Parveen, N..

10 recordsLinked to original sources

bioRxiv2026

Marine sulfated glycans inhibit the binding of Borrelia burgdorferi adhesins to heparin/GAGs

Recognition of host glycosaminoglycans (GAGs) is critical for the adhesion and colonization of Borrelia burgdorferi, but the structural requirements for these interactions remain incompletely understood. Using surface plasmon resonance (SPR), we characterized the heparin-binding properties of three Borrelia adhesins, i.e. Bgp, DbpA-A9, and DbpB-B31 and evaluated inhibitors comprising heparin oligosaccharides, selectively desulfated heparins, mammalian GAGs, marine sulfated glycans, pentosan polysulfate (PPS), and mucopolysaccharide polysulfate (MPS). All three adhesins bound directly to heparin, with equilibrium dissociation constants (KD) of 2.18 nM for Bgp and 36.4 nM for DbpB-B31. Competition assays identified N-sulfation and 6-O-sulfation as major determinants of binding, whereas 2-O-sulfation had a lower effect. Bgp showed chain-length dependence over dp4-dp20, whereas DbpB-B31 preferentially recognized longer oligosaccharides, particularly those of dp12 or greater. Chondroitin sulfate E also exhibited strong inhibitory activity, indicating that recognition was not restricted to heparin/heparan sulfate backbones. Among the tested sulfated glycans, fucosylated chondroitin sulfate (HfFucCS) from Holothuria floridana was the most potent inhibitor of Bgp, with an IC50 of approximately 4 ng/mL, whereas PPS was the strongest inhibitor of DbpB-B31, followed by MPS. These findings reveal protein-specific sulfated-glycan recognition and identify promising candidates for disrupting B. burgdorferi-host GAG interactions.

biochemistry↗

bioRxiv2026

Mosquito-directed PROTACs to block malaria transmission

The mosquito stage of the Plasmodium falciparum life cycle is an attractive target for intervention since it is crucial for the sexual reproduction and transmission of parasites to human host. Mosquito determinants crucial for parasite infection and growth pose as lucrative targets for transmission blockers. Owing to the fact that p38 MAPK has role in immune response and vector competence, we have evaluated the potential of PROTAC molecule (NR-7h) to degrade Anopheles stephensi p38 MAPK (Asp38 MAPK), a conserved serine/threonine kinase involved in stress reactions, midgut homeostasis, and parasite survival. PROTAC-mediated degradation of Asp38 MAPK led to the disrupted development of the parasite, suggesting its crucial function in vector competence. Furthermore, NR-7h-treated mosquitoes showed higher expression of immune genes such Rel-2, TEP1, APL1, and NOS, suggesting that p38 MAPK regulates host immunity in a way that promotes parasite persistence. PROTAC-mediated degradation of target proteins, provides a more persistent and resistance-proof therapeutic effect than traditional kinase inhibitors. Our findings establish PROTACs as a novel vector-targeted strategy for the development of endectocides to limit malaria transmission.

molecular biology↗

bioRxiv2026

Treponema pallidum TprD and TprK Are Adhesins and Promote Spirochetes Opsonophagocytosis

Treponema pallidum subspecies pallidum (T. pallidum) causes systemic syphilis, exclusively infects humans in nature and can persist for decades in the absence of treatment despite generating robust adaptive immune responses. The T. pallidum repeat (Tpr) family of outer membrane proteins are immunogenic but have been implicated in immune evasion due to antigenic variation, indicating that Tprs are virulence factors displayed on the spirochetes surface. Long-term survival of T. pallidum is largely attributed to the sparse surface-exposed outer membrane proteins and antigenic variation exhibited by the major surface protein TprK, which undergoes phase variation. Mechanism of antigenic variation has been studied for decades; however, functions of Tprs in this extracellular pathogen have not been experimentally determined. In this study, we determined TprD and TprK location, their role in adherence and in clearance of T. pallidum by macrophages. Using our now established heterologous surrogate system and gain-in-function approach using non-adherent related spirochete, the B314 strain of Borrelia burgdorferi, we show that both TprD and TprK are surface exposed to some extent on engineered B. burgdorferi as well as on T. pallidum Nichols and SS14 strains. We further demonstrate that both proteins mediate adherence to different mammalian cells in vitro and mouse antibodies generated against TprD and TprK putative surface loops bind spirochetes and promote J774A.1 macrophages-mediated opsonophagocytosis. Thus, surface-exposed adhesins TprD and TprK of T. pallidum contribute to binding to different types of cells which likely reflect the pathogens ability to colonize different tissues, and they are also targets of opsonic antibodies. IMPORTANCESyphilis remains a major global public health challenge and is exacerbated by the rising number of cases of congenital infection and increased risk of HIV acquisition and transmission in syphilitic patients. A critical barrier to improving understanding of the molecular basis of syphilis pathogenesis owe to fragility of T. pallidum, inability to grow it in pure culture, and difficulty in generating knockout mutants in predicted virulence factors due to their possible essential role in spirochetes viability. Our findings provide experimental evidence linking Tpr proteins to host cell adherence, their ability to generate humoral immune response in the rabbit model of infection as well as in humans which could facilitate clearance by macrophages. Demonstration of TprD and TprK as the targets of opsonic antibodies here highlight their potential as protective immunogens and emphasizes importance of their inclusion in the cocktail to produce effective vaccine against syphilis.

microbiology↗

bioRxiv2025

Retrospective analysis of Pseudomonas aeruginosa clinical isolates for secreted PrpL protease: a key virulence factor associated with corneal tissue damage

BackgroundPseudomonas aeruginosa is a ubiquitous organism that adapts well in different environments. It is an opportunistic bacterial pathogen that produces a wide range of virulence factors, colonizes lungs to cause pneumonia, causes non-healing wounds especially in burn victims, and is a major culprit in destructive keratitis. It can reach the cornea through reusable, extended-use contact lenses and by contaminated eyedrops and artificial tears. Secreted proteases of P. aeruginosa together with pyocyanin metabolite, which inhibits Serine Protease Inhibitors (Serpins) activity contribute to severe tissue damage during infection. MethodsP. aeruginosa strains isolated from different clinical sites were obtained from different researchers and clinicians for this study. We examined P. aeruginosa strains and secretion defective and prpL knockout mutants in PA64481 strain for lysyl endopeptidase (PrpL, a serine protease that cleaves after a lysine residue) activity using serine protease specific D-Val-Leu-Lys-p-nitroanalide substrate. We also determined pyocyanin production in these strains. ResultsExamination of secreted milieu from P. aeruginosa showed that 41 corneal isolates had detectable lysyl endopeptidase activity associated with PrpL at levels significantly higher than by 27 non-corneal isolates. We found that PrpL is secreted by the xcp-based type II secretion system. Bacterial culture supernatants displaying higher PrpL activity and not low secreted PrpL levels disrupted corneal epithelial cell monolayers in vitro, which is consistent with a role of this protease in destructive keratitis. Many examined strains also produced high levels of pyocyanin. ConclusionsThis retrospective examination of clinical P. aeruginosa suggests that high levels of PrpL and pyocyanin-producing isolates are more prevalent among corneal isolates and could enhance tissue damage during infection. Supporting this premise, corneal epithelial cell monolayers disrupted by high PrpL-producing strains but remained intact after treatment with P. aeruginosa mutants culture supernatants that lack or have reduced secreted PrpL.

microbiology↗

bioRxiv2025

Opposing roles for iron transport systems in gallium tolerance in extraintestinal pathogenic Escherichia coli

Gallium is a promising antibacterial candidate because it displaces iron atoms inside bacterial cells but does not undergo redox cycling. It inhibits growth by disrupting essential iron-dependent processes. However, Escherichia coli are naturally less sensitive to gallium than many other bacteria, and the mechanisms that control gallium tolerance are not completely understood. We performed a genome-wide transposon sequencing (TnSeq) screen to identify genes important for the survival of an extraintestinal pathogenic E. coli isolate (M12) in gallium nitrate. The TnSeq results indicated that inactivation of enterobactin siderophore-related genes (entS, fepD, fes, fepB) enhances bacterial survival in gallium, while disrupting the ferric dicitrate transport system increases susceptibility. We validated these findings through targeted gene knockouts and gallium sensitivity experiments. Our findings suggest that enterobactin can complex with gallium for cellular uptake, but that the ferric citrate receptor FecA can discriminate between gallium citrate and iron citrate. Expression of fecA increased with gallium exposure, showing that gallium induces FecA-mediated iron uptake. Gallium also increased intracellular levels of manganese in the {Delta}fecA strain. Supplementation with iron or manganese restored growth of M12 {Delta}fecA in gallium, suggesting that gallium sensitivity is linked to both iron starvation and oxidative stress. As the ferric dicitrate transport system is an important virulence factor in several extraintestinal infection sites, our results suggest that targeting FecA may increase E. coli susceptibility to gallium while also suppressing virulence. ImportanceEscherichia coli extraintestinal infections that are resistant to traditional antibiotics are associated with more deaths than any other species. Gallium-based therapies may represent a non-antibiotic approach to treating extraintestinal pathogenic E. coli strains that affect both humans and animals. Our results are significant as they show that the enterobactin siderophore and the ferric dicitrate iron transport systems expressed by these bacteria have opposing roles in E. coli gallium sensitivity. These findings could be leveraged to enhance the efficacy of gallium therapeutics.

microbiology↗

bioRxiv2023

Particle-based phasor-FLIM-FRET resolves protein-protein interactions inside single viral particles

Fluorescence lifetime imaging microscopy (FLIM) is a popular modality to create additional contrast in fluorescence images. By carefully analyzing pixel-based nanosecond lifetime patterns, FLIM allows studying complex molecular populations. At the single molecule or single particle level, however, image series often suffer from low signal intensities per pixel, rendering it difficult to quantitatively disentangle different lifetime species, such as during FRET analysis in the presence of a significant donor-only fraction. To address this problem, we combined particle localization with phasor-based FLIM analysis. Using simulations, we first showed that an average of [~]300 photons, spread over the different pixels encompassing single fluorescing particles and without background, is enough to determine a correct phasor signature (standard deviation <5% for a 4 ns lifetime). For immobilized single- or double-labeled dsDNA molecules, we next validated that particle-based phasor-FLIM-FRET readily allows estimating fluorescence lifetimes and FRET from single molecules. Thirdly, we applied particle-based phasor-FLIM-FRET to investigate protein-protein interactions in sub diffraction HIV-1 viral particles. To do this, we first quantitatively compared the fluorescence brightness, lifetime and photostability of different popular fluorescent protein-based FRET probes when genetically fused to the HIV-1 integrase enzyme (IN) in viral particles, and conclude that eGFP, mTurquoise2 and mScarlet perform best. Finally, for viral particles co-expressing FRET-donor/acceptor labeled IN, we determined the absolute FRET efficiency of IN oligomers. Available in a convenient open-source graphical user interface, we believe that particle-based phasor-FLIM-FRET is a promising tool to provide detailed insights in samples suffering from low overall signal intensities. Why it mattersPhasor-FLIM is an extraordinarily popular tool for fluorescence lifetime imaging analysis. However, it remains susceptible for low signal intensities, operational challenges and therefore required informed users and a clear analysis understanding. In this work we developed a convenient all-graphical workflow for quantitative phasor-FLIM in heterogenous and low-signal samples and applied it to quantifying absolute FRET efficiencies from protein-protein interactions inside single viral particles. Moreover, containing a well-illustrated theoretical introduction to time-domain phasor-FLIM, our paper helps novice users to correctly implement phasor-FLIM in standard microscopy practice.

biophysics↗

bioRxiv2023

Establishment of beta cell heterogeneity via differential CpG methylation at Nnat

Aims/hypothesisBeta cells within the pancreatic islet represent a heterogenous population wherein individual sub-groups of cells make distinct contributions to the overall control of insulin secretion. These include a subpopulation of highly-connected hub cells, important for the propagation of intercellular Ca2+ waves. Functional subpopulations have also been demonstrated in human beta cells, with an altered subtype distribution apparent in type 2 diabetes. At present, the molecular mechanisms through which beta cell hierarchy is established are poorly understood. Changes at the level of the epigenome provide one such possibility which we explore here by focussing on the imprinted gene neuronatin (Nnat), which is required for normal insulin synthesis and secretion. MethodsSingle cell RNA-seq datasets were examined using Seurat 4.0 and ClusterProfiler running under R. Transgenic mice expressing eGFP under the control of the Nnat enhancer/promoter regions were generated for fluorescence-activated cell (FAC) sorting of beta cells and downstream analysis of CpG methylation by bisulphite and RNA sequencing, respectively. Animals deleted for the de novo methyltransferase, DNMT3A from the pancreatic progenitor stage were used to explore control of promoter methylation. Proteomics was performed using affinity purification mass spectrometry and Ca2+ dynamics explored by rapid confocal imaging of Cal-520 and Cal-590. Insulin secretion was measured using Homogeneous Time Resolved Fluorescence Imaging. ResultsNnat mRNA was differentially expressed in a discrete beta cell population in a developmental stage- and DNA methylation (DNMT3A)-dependent manner. Thus, pseudo-time analysis of embryonic data sets demonstrated the early establishment of Nnat-positive and negative subpopulations during embryogenesis. NNAT expression is also restricted to a subset of beta cells across the human islet that is maintained throughout adult life. NNAT+ beta cells also displayed a discrete transcriptome at adult stages, representing a sub-population specialised for insulin production, reminiscent of recently-described "{beta}HI" cells and were diminished in db/db mice. Hub cells were less abundant in the NNAT+ population, consistent with epigenetic control of this functional specialization. Conclusions/interpretationThese findings demonstrate that differential DNA methylation at Nnat represents a novel means through which beta cell heterogeneity is established during development. We therefore hypothesise that changes in methylation at this locus may thus contribute to a loss of beta cell hierarchy and connectivity, potentially contributing to defective insulin secretion in some forms of diabetes. Research in contextWhat is already known about this subject? - Neuronatin (Nnat/NNAT) is an imprinted gene in humans and mice and is required for glucose-stimulated insulin secretion in vivo - Pancreatic beta cells are functionally heterogeneous with specific highly-connected subpopulations known to coordinate islet wide Ca2+ dynamics - Functional subpopulations have been described in human beta cells and their distribution is altered in type 2 diabetes What is the key question? - Does NNAT mark a discrete subpopulation of functional beta cells and which epigenetic pathways coordinate its formation and maintenance? What are the new findings? - A subpopulation of NNAT+ beta cells is established prior to the first week of postnatal life in mice via de novo DNA methylation at the Nnat promoter - NNAT+ beta cells are transcriptionally highly differentiated and appear to be functionally specialised for insulin production, possibly corresponding to recently-described "{beta}HI" and "CD63hi" beta cells. NNAT is expressed in a subset of beta cells across the human islet, and its deficiency in human beta cells diminishes glucose-stimulated insulin secretion - NNAT+ cells are likelier to belong to the population of follower, rather than hub cells, consistent with a role in insulin production rather than glucose detection How might this impact on clinical practice in the foreseeable future? - Epigenome-modifying compounds may provide a way of enhancing beta cell function and the ensemble behaviour of the islet to stimulate insulin secretion

cell biology↗

bioRxiv2022

DNA methylation Dependent Restriction of Tyrosine Hydroxylase Contributes to Pancreatic β-cell Heterogeneity

The molecular and functional heterogeneity of pancreatic {beta}-cells is well recognized. Pancreatic islets harbor a small subset of {beta}-cells that co-express Tyrosine Hydroxylase (TH), an enzyme involved in synthesis of catecholamines that repress insulin secretion. Restriction of this sub-population within islets is essential for appropriate insulin secretion. However, the distinguishing characteristics of this subpopulation and the mechanisms that restrict TH expression in {beta}-cells are not known. Here, we define the specific molecular and metabolic characteristics of the TH+ {beta}-cells and show that TH expression in {beta}-cells is restricted by DNA methylation patterning during {beta}-cell lineage specification. Ablation of de novo DNA methyltransferase Dnmt3a in the pancreatic- and endocrine-progenitor lineages results in a dramatic increase in the proportion of TH+ {beta}-cells, while {beta}-cell specific ablation of Dnmt3a has no effect on this sub-population. We demonstrate that maintenance of Th promoter DNA methylation patterns is essential for its continued restriction in postnatal {beta}-cells, and that loss of DNA methylation dysregulates TH expression in {beta}-cells in response to chronic overnutrition, contributing to impairment of {beta}-cell identity. These data highlight the essential requirement of DNA methylation patterning in regulating endocrine cell fates, and reveal a novel role of DNA methylation in {beta}-cell heterogeneity.

developmental biology↗

bioRxiv2022

Coordination between ECM and cell-cell adhesion regulates the development of islet aggregation, architecture, and functional maturation

Pancreatic islets are 3-dimensional cell aggregates consisting of unique cellular composition, cell-to-cell contacts, and interactions with blood vessels. Cell aggregation is essential for islet endocrine function; however, it remains unclear how developing islets establish aggregation. By combining genetic animal models, imaging tools, and gene expression profiling, we demonstrate that islet aggregation is regulated by extracellular matrix signaling and cell-cell adhesion. Islet endocrine cell-specific inactivation of extracellular matrix receptor Integrin {beta}1 disrupted blood vessel interactions but promoted cell-cell adhesion and the formation of larger islets. In contrast, ablation of cell-cell adhesion molecule -Catenin promoted blood vessel interactions yet compromised islet clustering. Simultaneous removal of Integrin {beta}1 and -Catenin disrupts islet aggregation and the endocrine cell maturation process, demonstrating that establishment of islet aggregates is essential for functional maturation. Our study provides new insights into understanding the fundamental self-organizing mechanism for islet aggregation, architecture, and functional maturation. HIGHLIGHTSO_LIIslet vascularization and aggregation are regulated via ECM-Itgb1 signaling C_LIO_LIECM-Itgb1 signaling negatively controls islet aggregation via regulation of cell-cell adhesion during development C_LIO_LICell-cell adhesion negatively regulates the interaction of endocrine cell-vasculature in islets C_LIO_LIDifferential cell adhesion regulates the establishment of islet architecture C_LIO_LIEndocrine functional maturation depends on islet aggregation regulated by the coordination of ECM-Itgb1 signaling and cell-cell adhesion C_LI

developmental biology↗

bioRxiv2019

A novel antibody targeting ICOS increases intratumoural cytotoxic to regulatory T cell ratio and induces tumour regression.

The immunosuppressive tumour microenvironment constitutes a significant hurdle to the response to immune checkpoint inhibitors. Both soluble factors and specialised immune cells such as regulatory T cells (TReg) are key components of active intratumoural immunosuppression. Previous studies have shown that Inducible Co-Stimulatory receptor (ICOS) is highly expressed in the tumour microenvironment, especially on TReg, suggesting that it represents a relevant target for preferential depletion of these cells. Here, we used immune profiling of samples from tumour bearing mice and cancer patients to characterise the expression of ICOS in different tissues and solid tumours. By immunizing an Icos knockout transgenic mouse line expressing antibodies with human variable domains, we selected a fully human IgG1 antibody called KY1044 that binds ICOS from different species. Using KY1044, we demonstrated that we can exploit the differential expression of ICOS on T cell subtypes to modify the tumour microenvironment and thereby improve the anti-tumour immune response. We showed that KY1044 induces sustained depletion of ICOShigh TReg cells in mouse tumours and depletion of ICOShigh T cells in the blood of non-human primates, but was also associated with secretion of pro-inflammatory cytokines from ICOSlow TEFF cells. Altogether, KY1044 improved the intratumoural TEFF:TReg ratio and increased activation of TEFF cells, resulting in monotherapy efficacy or in synergistic combinatorial efficacy when administered with the immune checkpoint blocker anti-PD-L1. In summary, our data demonstrate that targeting ICOS with KY1044 can favourably alter the intratumoural immune contexture, promoting an anti-tumour response.

cancer biology↗