Biology subjects

Prakash, S.

Publications and source records attributed to Prakash, S..

At least 19 recordsLinked to original sources

bioRxiv2026

A Tissue-Targeted Immunotherapy Protects Against Recurrent Ocular Herpes Infection and Disease, and Alleviates the Associated Cognitive, Behavioral, and Social Deficits in a 'Humanized' HLA Transgenic Mouse Model of UVB-Induced Reactivation

Background: Recurrent ocular herpetic disease is a leading infectious cause of corneal blindness and is associated with psychological distress, anxiety, and social withdrawal. No ocular herpes vaccine or immunotherapy currently exists. Hypothesis: A prime/pull/keep (PPK) T-cell immunotherapy, built on HSV-1 epitopes preferentially recognized by CD8+/CD4+ T cells from naturally protected asymptomatic (ASYMP) individuals, would reduce viral reactivation, shedding, recurrent corneal disease, and associated neurobehavioral deficits. Methods: Immunodominant HLA-restricted CD8+/CD4+ epitopes from HSV-1 gD/gB, tegument proteins VP11/12/VP13/14, and helicase UL9 were delivered via topical ocular AAV8 vector (prime) to latently infected HLA-A*0201/DRB1 mice, combined with chemokine CXCL11 (pull) and IL-2/IL-15 cytokines to sustain TRM cells (keep), then UVB-exposed to trigger reactivation and recurrent corneal disease. Results: PPK immunotherapy conferred robust, near-complete protection against recurrent HSV-1 disease and improved survival after UVB-induced reactivation, unlike its individual components. This protection was associated with site-specific accumulation of HSV-1-specific CD8+/CD4+ TRM cells in the cornea and trigeminal ganglia (TG), but not in the spleen, with peak retention from combined IL-2/IL-15, alongside durable central/effector memory populations. PPK-induced T cells showed enhanced polyfunctionality (cytokine production, cytotoxicity, proliferation), reduced exhaustion markers, and a TCF-1+ self-renewing phenotype. PPK also elicited robust, Th1-biased HSV-1-specific IgG responses complementing mucosal immunity, and restored spatial memory, reduced depression-like behavior, and reversed social withdrawal in mice with recurrent ocular herpes. Conclusion: PPK immunotherapy induces potent, durable antiviral TRM cells at primary sites of HSV-1 latency and reactivation, protecting against recurrent ocular herpetic disease and alleviating associated neurobehavioral deficits, supporting its translational potential in humans.

immunology↗

bioRxiv2026

A Comparative Benchmark of Biomedical Language Models for Concept Normalization from Real-World Text

Patient-reported and clinically documented narratives often contain informal, fragmented and linguistically heterogeneous expressions, complicating medical concept normalization (MCN) and frequently necessitating pre-processing before terminology mapping. Despite rapid advances in biomedical language modeling, the comparative utility of representation models for MCN and their integration with instruction-tuned LLMs in automated normalization workflows remain underexplored. In this study, we systematically benchmarked 15 general, biomedical and clinical transformer-based representation models together with 12 instruction-tuned LLMs across 12,713 instances from five established datasets: TAC2017_ADR, TwADR-L, TwiMed, CADEC and SMM4H2017. Representation models were evaluated using embedding-based semantic retrieval, whereas instruction-tuned LLMs were assessed as upstream text-correction modules. SapBERT achieved the highest Top-5 accuracy among representation models, reaching 63.8% for SNOMED CT and 58.0% for MedDRA without correction. Qwen 2 Instruct was selected as the preferred corrector on the basis of its favorable balance between Top-1 normalization performance and computational efficiency relative to substantially larger models, including Llama 3.1 Instruct (70B). Incorporation of Qwen 2 Instruct increased Top-5 accuracy to 69.4% for SNOMED CT and 63.2% for MedDRA. The resulting framework accepts heterogeneous short medical expressions without manual input pre-processing and automatically performs text refinement, semantic retrieval and terminology mapping to standardized concepts and vocabulary codes. These findings establish a benchmark-guided, scalable framework for automated medical terminology standardization.

bioinformatics↗

bioRxiv2026

Tissue-Resident and Effector-Memory Lymphocyte Recruitment to the Ocular Mucosa Requires CCL28/CCR10 Signaling During Recurrent HSV-1 Infection

Ocular herpes simplex virus type 1 (HSV-1) infection is a major cause of infectious corneal disease that can lead to recurrent herpetic keratitis, chronic inflammation, and vision loss. Although effective antiviral immunity is essential for controlling HSV-1 and limiting recurrent disease, the mechanisms regulating recruitment of protective lymphocytes to the ocular mucosa remain incompletely understood. CCL28 is a mucosa-associated chemokine expressed at epithelial surfaces that signals through its receptor CCR10, expressed by distinct antiviral T- and B-cell subsets. Using a mouse model of recurrent ocular HSV-1 infection, we investigated the role of the CCL28/CCR10 axis in mobilizing protective HSV-1-specific immune cells to the ocular mucosa (OM). Following infection, wild-type (WT) B6 mice showed a marked increase in CCL28 expression within the OM, coinciding with accumulation of HSV-specific effector-memory (CCR10+CD44+CD62L-) T cells and CCR10+B220+ B cells in the OM and trigeminal ganglia (TG). These cells displayed an antigen-experienced, effector-memory phenotype, suggesting that CCL28/CCR10 signaling contributes to a localized antiviral immune response in the ocular mucosa. By contrast, CCL28-deficient (CCL28(-/-)) mice: (i) were more susceptible to recurrent ocular HSV-1 infection; (ii) exhibited significantly reduced frequencies of HSV-specific effector-memory (CCR10+CD44+CD62L-) and tissue-resident memory (CCR10+CD69+CD103+) T cells, as well as CCR10+B220+ B cells, in the infected OM and TG; and (iii) showed diminished effector function (IFN-{gamma}, TNF, and Granzyme B production) in these populations, associated with increased viral burden and ocular pathology. These findings identify the CCL28/CCR10 axis as a critical regulator of antiviral B- and T-cell trafficking to the ocular mucosa, contributing to protection against recurrent herpetic eye disease and providing a mechanistic basis for targeting mucosal chemokine pathways to improve vaccine strategies against ocular herpes.

immunology↗

bioRxiv2026

Chlorogenic acid fails to confer neuroprotection in a chronic mouse model of Parkinsons disease

Parkinsons disease (PD) is a progressive neurodegenerative disorder characterized by -Synuclein (-Syn) aggregation, dopaminergic neuronal loss, and chronic neuroinflammation. Chlorogenic acid (CA), a dietary polyphenol abundant in coffee, exhibits antioxidant and anti-inflammatory properties and has shown neuroprotective effects in acute toxin-based PD models. However, its efficacy in chronic, -Syn-driven PD models remains unclear. Here, we evaluated the therapeutic potential of CA using an -Syn-based in vitro system and a chronic -Syn overexpression mouse model that recapitulates key pathological features of human PD. In vitro, CA significantly improved cell viability, reduced -Syn aggregation, and attenuated H2O2-induced apoptosis in U118 and N2a cells. In contrast, chronic oral administration of CA (100 mg/kg for 16 weeks) in C57BL/6J mice (male and female) failed to improve motor behavior, attenuate -Syn pathology, preserve nigrostriatal dopaminergic neurons, or reduce oxidative stress-associated DNA double-strand breaks in vivo. Notably, CA elicited a modest reduction in microglial and astrocytic activation in female mice, highlighting a sex-dependent immunomodulatory response. Collectively, these findings reveal a clear dissociation between robust in vitro neuroprotection and limited in vivo efficacy in a chronic -Syn-driven PD mouse model, emphasizing the importance of incorporating progressive disease paradigms and sex as a biological variable in preclinical therapeutic evaluation.

neuroscience↗

bioRxiv2026

Engineering bacterial combinatorial promoters for two-input chemical AND switching

Engineering compact bacterial promoters that integrate two chemical inputs would simplify biosensors, synthetic circuits, and dynamic metabolic control, but such promoters remain difficult to build because a second operator can increase partial-state leak, reduce the fully induced state, and create sequence- or reporter-context effects. Here we engineered 12 compact Marionette-derived two-input promoter architectures in Escherichia coli by combining defined template scaffolds with added operators, then quantified 19 reporter-specific four-state truth tables under a predefined operational criterion for chemical-input AND behaviour. Single-input template-promoter controls in matched reporter contexts helped separate weak parent-scaffold output from effects introduced by the second operator. Nine curated architectures passed in at least one reporter, and selected constructs delivered high-utility AND responses, including [11]/max-off separation up to >1,367. The full pass-fail dataset showed that a statistically higher [11] state does not necessarily create a useful switch. Practical utility depended mainly on suppressing [10] and [01] leakage, matching the template scaffold to the inserted operator, and controlling long-operator orientation and local sequence context. Reciprocal architectures behaved differently, and 4 of 7 dual-reporter architectures changed operational classification with reporter context. Thus, this work delivers a compact set of candidate Marionette-compatible two-input promoter parts and a source-traceable design framework for engineering bacterial chemical-input AND switches.

synthetic biology↗

bioRxiv2026

A genetically encoded local learning rule enables physical learning in engineered bacteria

Training physical neural networks directly in matter remains difficult because most platforms do not store and update weights within the same physical substrate. Here we show that engineered Escherichia coli can implement a genetically encoded local learning rule acting on persistent biological memory. We introduce memregulons: bacterial memory elements in which coupled plasmids store an analogue weight as the relative abundance of P1 within the P1+P2 plasmid pool. Environmental inputs activate local promoters, and a shared sublethal kanamycin signal converts promoter activity into differential growth that lowers the stored P1 fraction. In single strains, flow-cytometry trajectories across distinct promoters support the predicted dependence of weight change on learning-channel activity and standing population variance. At the single-cell level, repeated negative learning reshapes the stored weight distribution as it approaches the lower boundary. In mixed populations and co-cultures, one shared negative learning signal selectively rewrites active memregulons, enabling externally routed supervised tic-tac-toe lessons. We then generalise the architecture across orthogonal chemical inputs and combinatorial promoters, and use experimentally measured updates in hybrid analyses of winner-take-all and nonlinear-classifier tasks. These results establish physical learning in living matter through local negative updates of genetically encoded analogue weights.

synthetic biology↗

bioRxiv2026

Discovery and characterization of small molecule inhibitors of CBL-B that act as intramolecular glue to enhance T-cell anti-tumor activity

CBL-B is a RING-type E3 ubiquitin ligase that acts as a critical negative regulator of T-cell activation. It promotes T-cell anergy and suppresses immune responses through ubiquitin-mediated control of signaling proteins at the immunological synapse. T cells deficient in CBL-B activity lose their dependence on CD28 co-stimulation, exhibit heightened activation and increased cytokine production, and fail to re-establish anergy. In addition, mice deficient in CBL-B activity reject tumors. Together, this cellular mechanism and in vivo phenotype suggest inhibition of CBL-B may be a viable immuno-oncology therapeutic strategy. Here, we report the rational design and execution of a high-throughput screen (HTS) to identify small molecule inhibitors of CBL-B. This campaign led to the discovery of a scaffold that inhibits CBL-B E3 ligase activity with micromolar potency. Structural characterization revealed an intramolecular glue mechanism, in which the compound stabilizes the closed state of CBL-B, preventing phosphorylation of a tyrosine residue that is critical for activation and E2 binding. Iterative structure-activity optimization yielded compounds with nanomolar activity that enhanced T-cell activation and cytokine secretion in primary human T cells and suppressed tumor growth in a syngeneic colorectal mouse model. Together, these studies validate the biological rationale for pharmacological CBL-B inhibition and enabled the de novo discovery of intramolecular CBL-B glue inhibitors. This work culminated in the identification of NX-1607, a first-in-class oral CBL-B inhibitor now in clinical development for cancer immunotherapy.

immunology↗

bioRxiv2026

Synovial lining expressed mechanosensor PIEZO1 drives inflammation-permissive macrophage phenotypes and joint inflammation

Synovial tissue-resident macrophages regulate immune homeostasis within the joint, but can adopt an inflammation-permissive phenotype that promotes immune cell infiltration in rheumatoid arthritis (RA). Understanding the factors that drive this phenotypic switch may help prevent the localisation of inflammation in the joints of individuals at-risk of RA. We identified the mechanosensitive ion channel PIEZO1 as a potential regulator of lining-layer synovial tissue macrophage (STM) function. PIEZO1 was highly expressed in homeostatic, tissue resident TREM2pos lining-layer STMs and in its pathogenic chemokine producing TREM2low phenotype that characterises the hyperplastic lining-layer in active RA. Intra-articular injection of a PIEZO1 agonist in mice induced neutrophil and monocyte infiltration, whereas inhibition of PIEZO1 signalling restored the protective macrophage phenotype. Thus, mechanosensing via PIEZO1 is a defining feature of the joint lining-layer, and its aberrant activation by mechanical stress may lead to the localisation of inflammation within the joint, facilitating a transition from asymptomatic autoimmunity of at-risk RA to clinical disease.

immunology↗

bioRxiv2025

Filling knowledge gaps on vulnerable electric ray Narcine timlei (Elasmobranchii: Torpediniformes: Narcinidae) through ecological and biological studies

Narcine timlei (Bloch & Schneider, 1801), commonly known as the brown numbfish, recently moved from Data Deficient to Vulnerable in IUCN Red List. However, the biological information on the species is still lacking. In the present study, we provide an integrative assessment of populational dynamics on N. timlei. Morphological and molecular analyses ascertain the species identity as N. timlei. A total of 331 individuals were examined, comprising 156 males and 175 females, including 13 gravid, bringing an overall sex ratio of 1:1.21. We observed the highest specimens during the post-monsoon season (33.5%), and minimum during summer (18%). Length-weight relationships indicated negative allometric growth (b=2.78). Most specimens fall under the 17-20 cm size class, and a few exceeded 23 cm. Pregnant females carried 2 to 9 embryos. A total of 69 embryos from 13 females were obtained. Embryo length and weight were 5.18 {+/-} 0.22 cm and 2.02 {+/-} 0.36 g, respectively. No correlation was observed between maternal length and fecundity (R2 = 0.0036). Male to female embryo ratio was skewed to 1:1.3. The average length of electric organs in adults was 4.30 {+/-} 0.47 cm. Mean electrocyte count for adults was 117.55 {+/-} 14.48, while in embryos it was 123.36 {+/-} 15.9, indicating early developmental establishment of electrocyte structure in N. timlei. We strongly recommend long term species-specific monitoring and management measures to ensure conservation of this ecologically significant but often overlooked batoid.

zoology↗

bioRxiv2025

Development of an EPR-based methodology to study protein-lipid interaction

The interaction of protein with other biomolecules is central to all cellular processes. In particular, protein-lipid interactions play an essential role in regulating soluble and membrane protein function, structure, and dynamics. However, probing these interactions remains challenging due to the complexity and heterogeneity of membranes. Various methods have been developed to characterize protein-membrane interaction, each presenting advantages and limitations. This study presents a robust methodology based on continuous-wave Electron Paramagnetic Resonance (CW-EPR) spectroscopy to characterize protein-membrane interactions. We focused on the protein Tau, an intrinsically disordered protein associated with neurodegenerative diseases. We show that the interaction of labelled Tau with lipids gives rise to a very distinct lineshape, which can be used to quantify the fraction of bound protein. This allows to obtain the apparent binding mode and affinity through titration experiments. In addition, we show that a single measurement provides the absolute concentration of free and bound protein. We argue that this information, which is rarely obtained by other methods providing relative signals, is very useful for mechanistic studies. Furthermore, using mathematical modeling, we developed a minimal-data approach and demonstrated that a single EPR measurement can be used to derive binding constants. The approach is applied to the Taumembrane interaction occurring in different conditions affecting the binding behavior. The presented methodology is expected to be applicable to other proteins. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/672991v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@19cf0c5org.highwire.dtl.DTLVardef@833209org.highwire.dtl.DTLVardef@149fd8corg.highwire.dtl.DTLVardef@a4f50a_HPS_FORMAT_FIGEXP M_FIG C_FIG TOC Figure. Schematic representation of EPR-based analysis of Tau-membrane interactions using spin-labeled Tau and POPS vesicles

biophysics↗

bioRxiv2025

CXCL13/CXCR5 Chemokine Axis Promotes CXCR5+CD19+ B-Cell and Follicular/Effector CXCR5+CD4+ T-Cell Responses in the Lungs Associated with Protection from Severe and Fatal COVID-19 Following Infection with Pathogenic SARS-CoV-2 Delta Variant

Chemokines play an important role in shaping lung innate and adaptive immunity to pulmonary infections and diseases. However, the role of CXC ligand 13 (CXCL13), a chemokine homeostatically produced by various lung cell types, in the protection from SARS-CoV-2 infection and disease remains controversial. Some studies reported that asymptomatic patients who survived severe COVID-19 had CXCL13-dominated mucosal immune responses in the lungs early during infection. In contrast, other studies reported that a high level of CXCL13 was associated with severity and mortality in COVID-19 patients. In this study, to determine the direct role of CXCL13 in SARS-CoV-2 infection and disease, we generated CXCL13-/-K18-hACE2 mice, that are both transgenic for ACE2 and deficient in CXCL13 and compared their infection and COVID-19-like disease symptoms with those in wild-type K18-hACE2 transgenic mouse littermates following intranasal inoculation with the pathogenic SARS-CoV-2 delta variant (B.1.617.2). Compared to age- and gender-matched SARS-CoV-2 infected wild-type K18-hACE2 mice, SARS-CoV-2 infected CXCL13-/-K18-hACE2 deficient mice exhibited (i) higher viral load in the lungs; (ii) severe COVID-19-like lung pathology; (iii) exacerbated weight loss; (iv) increased mortality. The apparent severe COVID-19-like symptoms in CXCL13-/-K18-hACE2 deficient mice were associated with: (i) significantly lower frequencies of functional lung-resident C-X-C chemokine receptor 5+ (CXCR5)+CD19+ B cells, follicular CXCR5+CD4+ helper T cells (Tfh cells), and IFN-{psi}+TNF-+GzmB+Ki67+effector CD4+ Th1 cells; and (ii) a significant reduction in the levels of SARS-CoV-2-Spike specific Th1 associated IgG1 and IgG2b antibody isotypes. These findings corroborate previous human reports suggesting a critical role of the CXCL13/CXCR5 chemokine axis in the protective B- and T-cell mucosal immunity to SARS-CoV-2 infection and disease, offering a potential new immunotherapeutic target for treatment.

immunology↗

bioRxiv2025

M6A reader protein YTHDF3 regulates cardiomyocyte death and atrophy by modulating the alternative splicing program

BackgroundThe functional impact of m6A modifications on RNA is governed by reader proteins that read the m6A marks and process the RNA accordingly. Recent studies have highlighted the importance of m6A reader proteins in the heart. However, the function of a reader protein Ythdf3 in the heart remains completely unknown. ObjectiveWe aim to uncover the function of Ythdf3 in the hearts. Methods& ResultsHere, we show that Ythdf3 is indispensable for the heart, and its knockdown leads to cardiac cachexia evident by cardiomyocyte atrophy, death, and less dense myofibrils visualized by electron microscopy. We also found downregulation of Ythdf3 in response to doxorubicin, and its overexpression rescues doxorubicin-induced cardiomyocyte atrophy and death. Contrary to primarily cytoplasmic localization, we found that Ythdf3 localizes in the cardiomyocyte nucleus. Furthermore, using co-immunoprecipitation coupled with LC-MS/MS, we show that Ythdf3 interacts with splicing proteins like DDX5 and HNRNPU and regulates alternative splicing in the heart. Mechanistically, we found that Ythdf3 regulates the splicing of CaMKII{delta}, and its knockdown leads to an increase in CaMKII{delta}A and CaMKII{delta}C isoforms while a decrease in CaMKII{delta}9 isoform. ConclusionCollectively, Ythdf3 knockdown in the adult heart leads to cardiac cachexia due to the alteration of the splicing program.

cell biology↗

bioRxiv2025

A Pan-Beta-Coronavirus Vaccine Bearing Conserved and Asymptomatic B- and T-Cell Epitopes Protect Against Highly Pathogenic Delta and Highly Transmissible Omicron SARS-CoV-2 Variants of Concern.

SARS CoV-2 continues to evolve into new viral variants due to mutation, primarily in the Spike protein. Existing Spike-based vaccines are less effective because these variants can be more transmissible and evade vaccine-induced immunity. By targeting more conserved, Spike, and non-Spike, viral antigens using both arms of the adaptive immune system, i.e. B and T cells, we aim to reduce the reliance on neutralizing antibodies and avoid potential mismatches between the COVID-19 vaccines and circulating virus strains. In this way, enhanced immune memory function and broad-spectrum protection against existing and evolving virus variants can be attained. We have developed a mRNA-LNP-based multi-epitope vaccine incorporating conserved CD8+ T-cell, CD4+ T-cell, and B-cell epitopes. These conserved epitopes were selected as being highly recognized by B- and T-cells from unvaccinated asymptomatic COVID-19 patients. To evaluate the effectiveness of this multi-epitope "asymptomatic" vaccine, we utilized a novel triple transgenic h-ACE-2-HLA-A2/DR mouse model to enable the assessment of human T cell epitopes. Key observations include induction of: (i) robust protection against infection and disease caused by SARS-CoV-2 Delta (B.1.617.2) and Omicron (XBB.1.5) variants, as measured by reduced weight loss, virus replication, and lung pathology; (ii) strong antibody responses,; and (iii) potent SARS-CoV-2 epitope-specific IFN-{gamma}-producing CD4+/CD8+ T cells and Follicular helper CD4+ T (TFH) cells. These data support the strategy of targeting B-cells and T-cells directed toward highly conserved and "asymptomatic" epitopes, from both structural and non-structural viral protein antigens, to generate a broad-spectrum protective immunity to minimize disease impact across multiple SARS-CoV-2 variants.

immunology↗

bioRxiv2025

WRN and WRNIP1 ATPases impose high fidelity on translesion synthesis by Y-family DNA polymerases

Y-family DNA polymerases (Pols) are intrinsically highly error-prone; yet they conduct predominantly error-free translesion synthesis (TLS) in normal human cells. In response to DNA damage, Y-family Pols assemble and function together with WRN, WRNIP1, and Rev1 in TLS. Among these proteins, WRN possesses a 3[->]5 exonuclease activity and an ATPase/3[->]5 DNA helicase activity, and WRNIP1 has a DNA-dependent ATPase activity. In a previous study, we identified a role of WRN 3[->]5 exonuclease activity in the high in vivo fidelity of TLS by Y-family Pols. Here we provide evidence for a crucial role of WRN and WRNIP1 ATPase activities in raising the fidelity of TLS by these Pols. Defects in WRN and WRNIP1 ATPase activities cause a diversity of nucleotide (nt) misincorporations opposite DNA lesions by Y-family Pols, implicating an unprecedented role of these activities in restraining nt misincorporations, which they could accomplish by tightening the active site of the TLS Pol. Altogether, the combined actions of WRN and WRNIP1 ATPases in preventing nt misincorporations and of WRN exonuclease in removing misinserted nts confer such an enormous rise in the fidelity of Y-family Pols that they perform error-free TLS - essential for genome stability and cellular homeostasis.

genetics↗

bioRxiv2024

Spike mRNA Vaccine Encapsulated in a Lipid Nanoparticle Composed of Phospholipid 1,2-Dioleoyl-sn-Glycero-3-Phosphoethanolamine Induced Potent B- and T-cell Responses Associated with Protection against COVID-19 in Hamsters

Lipid nanoparticles (LNPs) have recently emerged as one of the most advanced vehicle platforms for efficient in vivo delivery of nucleoside-modified mRNA vaccine, particularly for COVID-19. LNPs comprise four different lipids: ionizable lipids, helper or neutral lipids, cholesterol, and lipids attached to polyethylene glycol (PEG). Studies on using the mRNA-LNP platform for vaccines have largely focused on the nucleic acid cargo with less attention to the LNP vehicle. While the LNPs protect mRNA from degradation and efficiently deliver the mRNA to antigen-presenting cells the effect of lipid composition and biophysical properties on the immunogenic and protective mRNA vaccine remain to be fully elucidated. In the present study, we used SARS-CoV-2 Spike-mRNA as a prototype vaccine, to study the effect of 4 different of LNPs with various lipid compositions. We demonstrate that when the same Spike-mRNA was delivered in the LNP4 formulation based on phospholipid 1,2-dioleoyl-sn-glycero-3- Phosphoethanolamine it outperformed the immunogenicity and protective efficacy of three LNPs (LNP1, LNP2, and LNP3) that are based on different lipids. Compared to other three LNPs, the LNP4: (i) enhanced phenotypic and functional maturation of dendritic cells; (ii) induced strong T-cell responses, (iii) increased secretion of proinflammatory, pro-follicular T helper (Tfh) cell cytokines; (iv) induced higher neutralization IgG titers; and (v) and provided better protection against SARS-CoV-2 infection and COVID-19 in the hamster model. We discussed the potential mechanisms by which LNP which include the phospholipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine may activate protective B- and T-cell responses.

immunology↗

bioRxiv2024

Tissue-Targeted Prime/Pull/Keep Therapeutic Herpes Simplex Virus Vaccine Protect Against Recurrent Ocular Herpes Diseases in HLA-A*0201 Transgenic Rabbits

Herpes Simplex Virus type 1 (HSV-1) continues to be one of the most prevalent viral infections globally, with approximately 3.72 billion individuals affected worldwide. A clinical herpes vaccine is still lacking. In the present study, a novel prime/pull/keep vaccine was tested in Human Leukocyte Antigen transgenic rabbit model of ocular herpes (HLA-A*0201 Tg rabbit). Ten asymptomatic (ASYMP) CD8+ T cell peptide epitopes and 3 CD4+ T-cell epitopes were selected from the HSV-1 glycoproteins D and B (gD and gB), viral tegument proteins (VP11/12 and VP13/14) and the DNA replication binding helicase (UL9), all preferentially recognized by CD8+ and CD4+ T-cells from "naturally protected" HSV-1-seropositive healthy ASYMP individuals (who never had recurrent corneal herpetic disease). HLA Tg rabbits were ocularly infected with HSV-1 then during latency at day 30 post-infection, the rabbits were ocularly vaccinated with a recombinant neurotropic AAV8 vector (107GC/ eye) encoding for the 10 CD8+ T cell peptide and 4 CD4+ T cells peptide (prime), T cell-attracting CXCL-11 (Pull) and T-cell keeping IL-2/IL-15 cytokines (keep). The rabbits were followed up for eye disease and viral loads in tears for 28 days. The frequency, function and protective efficacy of HSV-specific CD8+ T cells induced by the prime/pull/keep vaccine were assessed in the trigeminal ganglia (TG), cornea, Spleen, and peripheral blood. Compared to the mock group (unvaccinated), the peptides/CXCL11/IL-2/IL-15 vaccine generated frequent resident CD8+ T cells that infiltrated the TG. CD8+ T cells mobilization and retention into TG of prime/pull/keep vaccinated rabbits was associated with a significant reduction in corneal herpes infection and disease after an ocular HSV-1 challenge (McKrae). These findings draw attention to the novel prime/pull/keep therapeutic vaccine strategy to mobilize and retain anti-viral T cells into tissues protecting them against herpetic infection and disease. IMPORTANCEThere is an urgent need for a vaccine against widespread human herpes simplex virus infections. The present study demonstrates that immunization of humanized HLA-A*0201 transgenic rabbits with CD8+ and CD4+ T-cell epitope peptides (prime)/ CXCL11 (pull)/ IL-2/IL-15 (Keep) AAV8-based vaccine triggered mobilization ad retention of HSV-1-specific CD8+ T cells locally in the cornea and TG, the sites of acute and latent herpes infections. Mobilization and retention of antiviral CD8+ T cells into cornea and TG of HSV-1 infected rabbits that received the prime/pull/keep vaccine was associated with protection against ocular herpes infection and disease. These results highlight the importance of the prime/pull/keep vaccine strategy to bolster the number and function of protective CD8+ T cells within infected tissues.

immunology↗

bioRxiv2024

Dynamics of Spike-Specific Neutralizing Antibodies Across Five-Year Emerging SARS-CoV-2 Variants of Concern Reveal Conserved Epitopes that Protect Against Severe COVID-19

Since early 2020, several SARS-CoV-2 variants of concern (VOCs) continue to emerge, evading waning antibody mediated immunity produced by the current Spike-alone based COVID-19 vaccines. This caused a prolonged and persistent COVID-19 pandemic that is going to enter its fifth year. Thus, the need remains for innovative next generation vaccines that would incorporate protective Spike-derived B-cell epitopes that resist immune evasion. Towards that goal, in this study we (i) Screened the sequences of Spike among many VOCs and identified conserved and non-conserved linear B-cell epitopes; (ii) Compared titers and neutralization antibodies specific to these conserved and non-conserved B-cell epitopes from serum of symptomatic and asymptomatic COVID-19 patients that were exposed to multiple VOCs across the 5-year COVID-19 pandemic, and (iii) Compared protective efficacy of conserved versus non-conserved B-cell epitopes against the most pathogenic Delta variant in a "humanized" ACE-2/HLA transgenic mouse model. We found robust conserved B-cell epitope-specific antibody titers and neutralization in sera from asymptomatic COVID-19 patients. In contrast, sera from symptomatic patients contained weaker antibody responses specific to conserved B-cell epitopes. A multi-epitope COVID-19 vaccine that incorporated the conserved B-cell epitopes, but not the non-conserved B-cell epitopes, significantly protected the ACE2/HLA transgenic mice against infection and COVID-19 like symptoms caused by the Delta variant. These findings underscore the importance of conserved B-cell epitopes in generating robust protective immunity against severe COVID-19 symptoms caused by various VOCs, providing valuable insights for the development of broad-spectrum next generation Coronavirus vaccines capable of conferring cross-variant protective immunity. IMPORTANCEA persistent COVID-19 pandemic continues to evolve because of a continued emergence of SARS-CoV-2 variants of concern (VOCs) that escape the antibodies induced by the current Spike-alone COVID-19 vaccines. Identifying and characterizing the protective and non-protective Spike-derived B-cell epitopes that resist immune-evasion is a paramount for the development of broad-spectrum next generation Coronavirus vaccines. The present study identified Spike-derived conserved B cell epitopes that (i) are targeted by consistent and strong antibody responses in asymptomatic COVID-19 patients across the 5-year pandemic regardless of VOCs; and (ii) provided strong protection in humanized" ACE2/HLA transgenic mice against infection and COVID-19 like symptoms caused by the most pathogenic Delta variant. The findings have the potential to inform the design of next generation Coronavirus vaccines capable of conferring cross-variant protective immunity. TWEETProtective SARS-CoV-2 Conserved Linear B Cell Epitopes Identified from Spike Protein.

immunology↗

bioRxiv2024

Neuronal lipid droplets play a conserved and sex-biased role in maintaining whole-body energy homeostasis

ABSTRACTLipids are essential for neuron development and physiology. Yet, the central hubs that coordinate lipid supply and demand in neurons remain unclear. Here, we combine invertebrate and vertebrate models to establish the presence and functional significance of neuronal lipid droplets (LD) in vivo. We find that LD are normally present in neurons in a non-uniform distribution across the brain, and demonstrate triglyceride metabolism enzymes and lipid droplet-associated proteins control neuronal LD formation through both canonical and recently-discovered pathways. Appropriate LD regulation in neurons has conserved and male-biased effects on whole-body energy homeostasis across flies and mice, specifically neurons that couple environmental cues with energy homeostasis. Mechanistically, LD-derived lipids support neuron function by providing phospholipids to sustain mitochondrial and endoplasmic reticulum homeostasis. Together, our work identifies a conserved role for LD as the organelle that coordinates lipid management in neurons, with implications for our understanding of mechanisms that preserve neuronal lipid homeostasis and function in health and disease. HIGHLIGHTSO_LILipid droplets (LD) normally form in neurons across species Neuronal LD are regulated by a conserved gene network C_LIO_LINeuronal LD regulation plays a conserved and sex-biased role in maintaining energy homeostasis C_LIO_LILD regulation supports ER and mitochondrial function in hunger-activated neurons C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/613929v1_ufig1.gif" ALT="Figure 1"> View larger version (54K): org.highwire.dtl.DTLVardef@1936e92org.highwire.dtl.DTLVardef@40478aorg.highwire.dtl.DTLVardef@18d5faorg.highwire.dtl.DTLVardef@882ee9_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗