Biology subjects

Naik, A.

Publications and source records attributed to Naik, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Proteolytic control of the SARS-CoV-2 furin cleavage site defines the phenotypic evolution from the pandemic to endemic state

The successive emergence of SARS-CoV-2 variants with altered tissue tropism has progressively decoupled transmissibility from lower respiratory tract pathogenicity. Omicron lineages transmit exceptionally well whilst limiting severe lung disease. Here, we demonstrate that this phenotype has evolved through two temporally distinct tissue-specific proteolytic controls at the Spike (S) furin cleavage site (FCS). At the virion level, cell-type-dependent furin-mediated FCS hyper-cleavage depletes S in lung but not nasal epithelial cells. At the infected cell membrane during cell-cell spread, TMPRSS2 mediates S FCS cleavage but is negatively regulated in the presence of ACE2. Tissue-specific solute carriers SLC6A19 and SLC6A20 sequester ACE2, thereby relieving inhibition of TMPRSS2 and enabling S FCS cleavage during cell-cell spread. Critically, whilst all SARS-CoV-2 lineages benefit from this latter pathway, Omicron variants have evolved exclusive dependence on it, a strategic consolidation that focuses S proteolytic activation to TMPRSS2 alone. The combined outcomes of S protein regulation across viral and cell membranes reveal how tissue-specific proteolytic optimisation drives Omicron's transmission fitness advantage in the upper respiratory tract but at the cost of heavy attenuation in the lower respiratory tract.

microbiology↗

bioRxiv2026

Distributed resonant coupling enables high efficiency power transfer to mm-scale bioelectronics

Miniaturized implantable bioelectronics offer breakthrough potential in disease treatment, biomarker monitoring, and physiological sensing. However, wireless power transfer (WPT) remains a central limitation for millimeter-scale devices, as scaling down the receiver size rapidly decreases coupling efficiency due to tissue attenuation, low quality-factors, reduced mutual inductance, and limited tolerance to spatial and angular displacement. Here, we introduce distributed resonant coupling (DRC), a 3-coil WPT paradigm which enables the receiver (Rx) to actively participate in a strongly coupled resonance, transforming the Rx from a passive energy harvester into an active participant in a strongly coupled regime. By co-designing transmitters (Tx), resonators (Rs), and mm-scale receiver coils (Rx) as fully coupled systems, DRC exhibits simulated maximum power levels of 66%, measured received power transfer efficiencies of [~]56%, and end-to-end DC power transfer efficiencies of 42%, delivering >420 mW to loads at 1 W of transmitted power while maintaining robust performance across a range of practically relevant orientations and tissue media. Systematic theoretical and experimental efforts establish core design rules for DRC systems enabling operation without specialized tuning integrated circuits or components. To illustrate the capabilities of DRC in practical applications in vivo, we demonstrate three technologies that capture a broad application space in bioelectronics: implant localization, rapid wireless battery charging, and ultraminiaturized drug delivery devices compatible with particulate drug formulations. Taken together, these results suggest operational capabilities across a wide range of angular tolerances (up to 60{degrees}), tissue depths and dielectric and scattering media.

bioengineering↗

bioRxiv2026

Extending structural surfaceomics to identify aberrant conformations of tumor surface proteins as potential immunotherapy targets

The complement of tumor cell surface proteins, or "surfaceome", is a rich source of potential immunotherapy targets. To move beyond expression-based target discovery, we previously described "structural surfaceomics," combining crosslinking mass spectrometry (XL-MS) with surface protein biotinylation to identify conformation-selective targets. In our prior work, we applied this method to a single model of acute myeloid leukemia (AML), identifying active integrin beta-2 as a promising target. Here, we expand structural surfaceomics to identify additional immunotherapy targets and surface protein biology across additional models of AML, multiple myeloma, and prostate cancer, as well as donor peripheral blood mononuclear cells. Utilizing these models and different chemical crosslinkers, we compile an extensive database of 5,209 crosslinks. We characterize both shared and unique crosslink-based features, identifying 1,612 disease model-specific crosslinks, including 212 potentially defining tumor-specific conformations based on distance constraint violations relative to AlphaFold predictions. We further implement a suite of emerging modeling tools to predict tumor-specific protein structures. We probe crosslinking patterns suggesting multiple myeloma-specific CD48 and AML-specific integrin 1/{beta}4 heterodimer conformations. This work establishes a resource for cancer structural biology by implementation of structural surfaceomics. Our findings also point toward more realistic protein design models, potentially enabling systematic detection of targetable cancer-specific epitopes for next-generation immunotherapies.

cancer biology↗

bioRxiv2025

Ceramide transfer protein regulates G-protein coupled phospholipase signalling in Drosophila photoreceptors.

The non-vesicular transfer of lipids between organelles at membrane contact sites (MCS) has been proposed as a key principle in the regulation of cell physiology. While several proteins with lipid transfer activity have been identified and localized to MCS, their functional significance for supporting physiology is poorly understood. Ceramide transfer protein (CERT) is one such molecule that can transfer ceramide between membranes in vitro. However, evidence for the mechanism and in vivo significance of CERT function is limited. In this study, we have analyzed the function of the only gene (dcert) encoding CERT in Drosophila. We find that loss of function alleles of dcert (dcert1), show elevated levels of short chain ceramide species along with a reduction in the levels of its metabolite phosphatidyl ethanolamine ceramide. Physiological analysis of dcert1 mutant alleles showed reduced electrical responses in the eye to light stimulation although photoreceptors did not undergo retinal degeneration, and this phenotype could be rescued by reconstitution of dcert1 with the wild type gene. The altered light response in dcert1 was associated with a reduction in the rate of phosphatidylinositol 4,5 bisphosphate (PIP2) resynthesis following light induced phospholipase C (PLC) stimulation. The reduced electrical response of dcert1 could be suppressed by reducing ceramide synthesis at the ER. Taken together, our findings suggest that ceramide synthesized at the ER and transferred to the Golgi by CERT regulates G-protein coupled phospholipase C signaling in vivo.

cell biology↗

bioRxiv2025

A Geometrically Transient Material for Bioelectronic Implants

The outstanding barrier properties of skin make it difficult to obtain reliable physiological information (especially chemical) without the use of implantable bioelectronic sensing devices to directly access the internal biology. The clinical utility of bioelectronic implants however, hinges on a key geometrical optimization problem: devices must scale-down in size to reduce surgical invasiveness while also creating enough space to integrate electronics for wireless power delivery, data exchange, and electrical/electrochemical monitoring. Here, we present a minimally invasive bioelectronic implant with a transient geometry that can be inserted subcutaneously and measures important markers, such as pH, temperature, cardiac and respiratory activity, and levels of lithium, an important element for medical applications. To produce this new class of minimally invasive and foldable implantable sensors, we developed a fabrication method that works with highly flexible substrates to enable multiple-fold miniaturization during implantation. After implantation, the implant autonomously unfolds back to its planar form for continuous wireless operation. We demonstrate the key concepts concerning implantation, operation, and removal through extensive in vitro, ex vivo, and in vivo animal experiments. Our approach allows for robust multiplexed monitoring using quick and suture-free insertion procedures and may provide a unique advantage in the transition towards personalized health profiles.

bioengineering↗

bioRxiv2025

Affinity-matured CD72-targeting Nanobody CAR T-cells Enhance Elimination of Antigen-Low B-cell Malignancies

BackgroundChimeric antigen receptor (CAR) T-cell therapies are highly efficacious for several different hematologic cancers. However, for most CAR T targets it is observed that low surface antigen density on tumors can significantly reduce therapeutic efficacy. Here, we explore this dynamic in the context of CD72, a surface antigen we recently found as a promising target for refractory B-cell cancers, but for which CD72 low antigen density can lead to therapeutic resistance in preclinical models. MethodsPrimary samples were accessed via institutional review board-approved protocols. Affinity-matured and humanized nanobody clones were previously described in Temple et al.1 CAR T-cells were generated via lentiviral transduction. In vitro cytotoxicity assays were performed using luciferase-labeled cell lines. In vivo studies were performed using cell line- or patient-derived xenografts implanted in NOD scid gamma (NSG) mice. ResultsWe first confirmed ubiquitous CD72 expression across a range of primary B-cell non-Hodgkin lymphomas. We further found that after resistance to CD19-directed therapies, across both B-cell acute lymphoblastic leukemia (B-ALL) models and primary tumor samples, surface CD72 expression was largely preserved while CD22 expression was significantly diminished. Affinity maturation of a nanobody targeting CD72, when incorporated into chimeric antigen receptor (CAR) T-cells, led to more effective elimination in vitro of isogenic models of CD72 low-expressing tumors. These results suggested that nanobody-based CAR T-cells (nanoCARs) may exhibit a similar relationship between binder affinity, antigen expression, and efficacy as previously demonstrated only for scFv-based CAR T-cells. Surprisingly, however, this significantly improved in vitro efficacy only translated to modest in vivo survival benefit. As a parallel strategy to enhance CAR T function, we found that the small molecule bryostatin could also significantly increase CD72 surface antigen density on B-cell malignancy models. Structural modeling and biochemical analysis identified critical residues improving CD72 antigen recognition of our lead affinity-matured nanobody. ConclusionsTogether, these findings support affinity-matured CD72 nanoCARs as a potential immunotherapy product for CD19-refractory B-cell cancers. Our results also suggest that for B-ALL in particular, CD72 may be a preferable second-line immunotherapy target over CD22. What is already known on this topicPrevious work using single chain variable fragment (scFv) based CAR Ts has suggested that improving affinity for target antigen could potentially help mitigate tumor resistance mediated by antigen downregulation, or baseline low antigen density. However, it is unknown whether this same dynamic holds for CAR T-cells that utilize different antigen recognition elements, such as nanobodies. What this study addsHere we show that affinity maturation of nanobody-based CAR T-cells (nanoCARs) targeting CD72 can improve their in vitro efficacy versus CD72-low tumors; however, in vivo efficacy differences are more modest. Furthermore, we show that for refractory B-cell malignancies, surface CD72 appears preserved after CD19 resistance even in situations where CD22 is strongly downregulated. How this study might affect research, practice or policyCD72 warrants further investigation as a preferred immunotherapy target in the context of CD19-refractory B-cell cancers, though nanobody affinity maturation is not a universal solution to the challenge of low tumor surface antigen density.

immunology↗

bioRxiv2025

Anti-tumor effects of a novel cell penetrating peptide-based therapeutic approach to target Lactate Dehydrogenase C (LDHC) in triple negative breast cancer.

BackgroundLactate Dehydrogenase C (LDHC) is a promising candidate for therapeutic targeting thanks to its highly tumor-specific expression, immunogenicity, and pro-tumorigenic functions. Aberrant LDHC expression is associated with poor clinical outcomes in multiple cancers, including breast cancer. However, no specific LDHC inhibitors are currently available, highlighting the need for novel strategies to selectively target LDHC in tumor cells. This study explores the anti-tumor potential of cell-penetrating peptides (CPPs) to target LDHC in triple negative breast cancer (TNBC). MethodsFour CPPs were evaluated for their ability to deliver LDHC siRNA to tumor cells, including the positively charged 10R peptide (10R) and three bifunctional peptides containing the integrin v{beta}3 recognition motif Arg-Gly-Asp (RGD): 10R-RGD, cyclicRGD-10R (cRGD-10R), and internalizing RGD-10R (iRGD-10R). We characterized the physicochemical properties of all CPP:siRNA complexes, and determined their serum stability, cytotoxicity, cellular uptake, and LDHC silencing efficiency in vitro. The anti-tumor effects and cytotoxicity of cRGD-10R:siRNA and iRGD-10R:siRNA complexes were further assessed in a TNBC xenograft zebrafish model. ResultsAll four CPPs formed stable nanocomplexes with favorable safety profiles. The 10R-RGD and cRGD-10R peptides demonstrated the most efficient LDHC knockdown, reduced the clonogenic ability of TNBC cells and enhanced their treatment response to the chemotherapeutic drug olaparib in vitro. Treatment of TNBC xenograft zebrafish with 10R-RGD:siRNA and cRGD-10R:siRNA complexes significantly reduced tumor burden without inducing major toxicity. Conclusion Our findings demonstrate that CPP-based siRNA delivery provides a novel and safe approach to target LDHC, either as a monotherapy or in combination with common anti-cancer drugs, to enhance treatment outcomes.

cancer biology↗

bioRxiv2025

Role of IL10 signaling in the circadian control of host response to Influenza infection

We have previously demonstrated that the circadian clock regulates the host response to influenza A virus (IAV) infection, conferring a time-of-day-specific protection -infection at dawn resulted in a threefold increase in survival and reduced immunopathology compared to infection at dusk. While IL10 is well-known for its immunoregulatory function, its role in IAV remains unclear, with studies reporting both protective and detrimental effects. Given the diurnal rhythmicity of IL-10 receptor (Il10ra) expression in the lung, we investigated the contribution of IL-10 signaling to time-of-day-specific IAV protection. We found that blocking IL10 signaling abrogated the time of day protection, leading to increased immunopathology characterized by enhanced lymphocyte infiltration and global immune activation (transcriptomic analysis). Interestingly, while later, IL-10R blockade also eliminated the time-of-day difference in IAV outcomes, it improved the outcome of dusk-infected mice. Furthermore, natural killer (NK) cell depletion suppressed IL-10 levels in bronchoalveolar lavage, suggesting a role for NK cells in regulating IL-10 signaling. In conclusion, incorporating the circadian context has not only clarified the IL-10 role in IAV infection but also underscored the pivotal influence of circadian regulation on immune responses.

immunology↗

bioRxiv2025

Sources of non-uniform coverage in short-read RNA-Seq data

The origin of several normal cellular functions and related abnormalities can be traced back to RNA splicing. As such, RNA splicing is currently the focus of a vast array of studies. To quantify the transcriptome, short-read RNA-Seq remains the standard assay. The primary technical artifact of RNA-Seq library prep, which severely interferes with analysis, is extreme non-uniformity in coverage across transcripts. This non-uniformity is present in both bulk and single-cell RNA-Seq and is observed even when the sample contains only full-length transcripts. This issue dramatically affects the accuracy of isoform-level quantification of multi-isoform genes. Understanding the sources of this non-uniformity is critical to developing improved protocols and analysis methods. Here, we explore eight potential sources of non-uniformity. We demonstrate that it cannot be explained by one factor alone. We performed targeted experiments to investigate the effect of fragment length, PCR ramp rate, and ribosomal depletion. We assessed existing data sets with varying sample quality, PCR cycle number, reverse transcriptase, and technical or biological replicates. We found evidence that interference of reverse transcription by secondary structure is unlikely to be the major contributing factor, that rRNA pull-down methods do not cause non-uniformity, that PCR ramp rate does not substantially impact non-uniformity, and that shorter fragments do not reduce non-uniformity. All these findings contradict prior publications or recommendations.

bioinformatics↗

bioRxiv2024

Longitudinal cellular and humoral immune responses following Covid-19 BNT162b2-mRNA-based booster vaccination of craft and manual workers in Qatar.

IntroductionIn March 2020, the rapid spread of SARS-CoV-2 prompted global vaccination campaigns to mitigate COVID-19 disease severity and mortality. The 2-dose BNT162b2- mRNA vaccine effectively reduced infection and mortality rates, however, waning vaccine effectiveness necessitated the introduction of a third vaccine dose or booster. To assess the magnitude and longevity of booster-induced immunity, we conducted a longitudinal study of SARS-CoV-2 specific cellular and humoral immune responses among Qatars vulnerable craft and manual worker community. We also investigated the impact of prior naturally acquired immunity on booster vaccination efficacy. MethodsSeventy healthy participants were enrolled in the study, of whom half had prior SARS-CoV-2 infection. Blood samples were collected before and after booster vaccination to evaluate immune responses through SARS- CoV-2 specific ELISpots, IgG ELISA, neutralization assays, and flow cytometric immunophenotyping ResultsT cell analysis revealed increased Th1 cellular responses, marked by enhanced IFN-{gamma} release, in recently infected participants, which was further enhanced by booster vaccination for up to 6-months. Furthermore, booster vaccination stimulated cytotoxic T cell responses in infection-naive participants, characterized by granzyme B production. Both natural SARS-CoV-2 infection and booster vaccination induced robust and durable SARS-CoV-2 specific humoral immune responses, with high neutralizing antibody levels. Prior natural infection was also linked to an increased number of class- switched B cells prior to booster vaccination. ConclusionThese findings underscore the importance of booster vaccination in enhancing anti-viral immunity across both infection-naive and previously infected individuals, enhancing distinct arms of the anti-viral immune response and prolonging naturally acquired immunity.

immunology↗

bioRxiv2024

Immunomodulatory effects of tumor Lactate Dehydrogenase C (LDHC) in breast cancer

BackgroundImmunotherapy has significantly improved outcomes for cancer patients; however, its clinical benefits vary among patients and its effectiveness across breast cancer subtypes remains uncertain. To enhance its efficacy, it is important to gain more insight into tumor-intrinsic immunomodulatory factors that could be used as therapeutic targets. We previously identified Lactate Dehydrogenase C (LDHC) to be a promising anti-cancer target due to its role in regulating cancer cell genomic integrity. In this study, we investigated the effects of tumor LDHC expression on immune responses. MethodsTIMER AND TIDE deconvolution methods were used to investigate the relationship between tumor LDHC expression, immune cell infiltration and T cell dysfunction. Multiplex cytokine assays and flow cytometry analyses of breast cancer cell monocultures, and direct and indirect cancer cell-immune cell co-culture models were performed to assess the effect of LDHC knockdown on the secretion of inflammatory mediators and the expression of immune checkpoint molecules. T cell activity was determined by IFN-{gamma} ELISPot assays and 7-AAD viability flow cytometry of cancer cells in direct co-culture. ResultsTIMER and TIDE analyses revealed that tumor LDHC expression is associated with T cell dysfunction in breast cancer and worse post-immunotherapy survival in melanoma. Depletion of LDHC in three breast cancer cell lines (MDA-MB-468, BT-549, HCC-1954) enhanced T cell activation and cytolytic function (4-hour direct co-culture). Analysis of cancer cell monocultures revealed an increase in secreted pro-inflammatory cytokines (IFN-{gamma}, GM- CSF, MCP-1, CXCL1), a decrease in immunosuppressive factors (IL-6, Gal-9) and a reduction in tumor cell surface PD-L1 expression following LDHC knockdown. Using 72-hour direct co- cultures with LDHC-silenced cancer cells, we observed a decrease in tumor-promoting cytokines (IL-1{beta}, IL-4 and IL-6) and an increase in the tumor-inhibiting cytokine CXCL1. Furthermore, LDHC knockdown reduced the number of CD8+ T cells expressing PD-1 and CTLA-4, as well as the cell surface expression of CTLA-4, TIGIT, TIM3, and VISTA. ConclusionsOur findings suggest that targeting LDHC may improve anti-tumor immune responses by modulating the secretion of pro- and anti-tumorigenic cytokines and impairing immune checkpoint signaling. Further studies are needed to elucidate the molecular mechanisms by which LDHC modulates these responses in breast cancer.

cancer biology↗

bioRxiv2024

The accessory domains of Phospholipase D regulate its localization and activity in Drosophila photoreceptors

Phospholipase D (PLD) is activated during the response of animal cells to a large number of extracellular signals including growth factors, hormones and other extracellular stimuli. The product of PLD activity, phosphatidic acid (PA) is thought to act as a signalling molecule in cells. Since both the substrate, phosphatidylcholine and product phosphatidic acid (PA) are membrane anchored lipids, the precise localization and activation of PLD at specific locations in cells is critical. Here we report that that the precise localization of Drosophila PLD (dPLD) in photoreceptors is critical to maintain the structure of these cells during illumination. This localization of dPLD is dependent on its N-terminal PH domain; deletion of the PH domain (dPLD{Delta}PH) results in mis localization of the protein, loss of functional activity and dPLD{Delta}PHis unable to rescue the phenotypes of dPLD loss of function. By contrast, deletion of the PX domain (dPLD{Delta}PX) does not impact the localization of dPLD but enhances the functional activity of the protein. Thus the PH and PX domains of dPLD regulate its localization and activity respectively.

cell biology↗

bioRxiv2024

Handy divisions: Hand-specific specialization of prehensile control in bimanual tasks

When hammering a nail, why do right-handers wield the hammer in the right hand? The dynamic dominance theory suggests a somewhat surprising answer. The two hands are specialized for different types of tasks: the dominant for manipulating objects, and the non-dominant for stabilizing objects. Right-handers wield the moving object with their right hand to leverage the skills of both hands. Functional specialization in hand use is often illustrated using examples of object manipulation. However, the dynamic dominance theory is supported by wrist kinematics rather than object manipulation data. Therefore, our goal was to determine whether this theory extends to object manipulation. We hypothesized that hand-specific differences will be evident in the kinematics of hand-held objects and in the control of grip forces. Right-handed individuals held two instrumented objects that were coupled by a spring. They moved one object while stabilizing the other object in various bimanual tasks. They performed motions of varying difficulty by tracking predictable or unpredictable targets. The two hands switched roles (stabilization vs movement) in various experimental blocks. The changing spring length perturbed both objects. We quantified the movement performance by measuring the objects positions, and grip force control by measuring grip-load coupling in the moving hand and mean grip force in the stabilizing hand. The right hand produced more accurate object movement, along with stronger grip-load coupling, indicating superior predictive control of the right hand. In contrast, the left hand stabilized the object better and exerted a higher grip force, indicating superior impedance control of the left hand. Task difficulty had a weak effect on grip-load coupling during object movement and no effect on mean grip force during object stabilization. These results suggest that dynamic dominance extends to object manipulation, though the weak effect of task difficulty on grip characteristics warrants further investigation.

neuroscience↗

bioRxiv2024

Enabling Packaging Integration of Paper-based Electrical Gas Sensors for the Monitoring of Spoilage in Fresh Spinach

Gas sensors present an alternative to traditional off-package food quality assessment, due to their high sensitivity and fast response without the need of sample pre-treatment. The safe integration of gas sensors into packaging without compromising sensitivity, response rate, and stability, however, remains a challenge. Such packaging integration of spoilage sensors is crucial towards preventing food waste and transitioning towards more sustainable supply chains. Here, we demonstrate a wide-ranging solution to enable the use of gas sensors for the continuous monitoring of food spoilage, building upon our previous work on Paper-based Electrical Gas Sensors (PEGS). By comparing various materials commonly used in the food industry, we analyze the optimal membrane to encapsulate PEGS for packaging integration. Focusing on spinach as a high-value crop, we assess the feasibility of PEGS to monitor the gases released during its spoilage at low and room temperatures. Finally, we integrate the sensors with wireless communication and batteryless electronics, creating a user-friendly system to evaluate the spoilage of spinach, operated by a smartphone via Near Field Communication (NFC). The work reported here provides an alternative approach that surpasses traditional on-site and in-line monitoring, ensuring comprehensive monitoring of food shelf life.

bioengineering↗

bioRxiv2024

Glomerular Elasticity and Gene Expression Patterns Define Two Phases of Alport Nephropathy

Alport syndrome (AS), caused by COL4A3,4,5 mutations, leads to progressive glomerular disease and eventual kidney failure. In Col43-/- mice (C57BL/6 background), we found that increased glomerular capillary deformability (reduced Youngs modulus, E) appears 2-3 months before detectable proteinuria or elevated serum creatinine. This early change indicates that podocyte injury precedes traditional clinical markers of disease and corresponds to reduced podocyte adhesion and loss. Bulk and podocyte-enriched RNA-seq data, obtained from deconvoluting bulk RNA sequencing data, showed that starting at 4 months, endoplasmic reticulum (ER) stress and the unfolded protein response (UPR) steadily rise while extracellular matrix remodeling, inflammation, epithelial-mesenchymal transition, and maladaptive repair begin. By 7 months, pathology shifts toward widespread parenchymal fibrosis, interleukin, cytokine, and chemokine signaling, cytoskeleton disruption, metabolic failure, and podocyte dedifferentiation. Notably, administration of the chemical chaperone Tauro-Urso-Deoxcycholic Acid (TUDCA) from weaning reduced ER stress, preserved glomerular stiffness, minimized podocyte detachment and loss, normalized inflammatory, injury, and fibrotic gene expression, and halved proteinuria and serum creatinine at 7 months, preserving kidney structure. Differentially expressed podocyte enriched genes from 4-month Col43-/- (vs WT) mice were mapped to human orthologs in the NEPTUNE cohort. Four genes (CRB2, GPC6, NKD1, STX11) were associated with End Stage Renal Disease or 40% loss of eGFR (ESRD40). Collectively, our results identify these four genes and ER stress/UPR as key and treatable drivers of podocyte injury and disease progression in Alport syndrome, well before overt proteinuria, and highlight UPR activation and several genes as promising targets for disease-modifying therapies.

cell biology↗

bioRxiv2023

Challenges in the discovery of tumor-specific alternative splicing-derived cell-surface antigens in glioma

BackgroundDespite advancements in cancer immunotherapy, solid tumors remain formidable challenges. In glioma, profound inter-and intra-tumoral heterogeneity of antigen landscape hampers therapeutic development. Therefore, it is critical to consider alternative sources to expand the repertoire of targetable (neo-)antigens and improve therapeutic outcomes. Accumulating evidence suggests that tumor-specific alternative splicing (AS) could be an untapped reservoir of neoantigens. ResultsIn this study, we investigated tumor-specific AS events in glioma, focusing on those predicted to generate major histocompatibility complex (MHC)-presentation-independent, cell-surface neoantigens that could be targeted by antibodies and chimeric antigen receptor (CAR)-T cells. We systematically analyzed bulk RNA-sequencing datasets comparing 429 tumor samples (from The Cancer Genome Atlas [TCGA]) and 9,166 normal tissue samples (from the Genotype-Tissue Expression project [GTEx]), and identified 13 AS events in 7 genes predicted to be expressed in more than 10% of the patients, including PTPRZ1 and BCAN, which were corroborated by an external RNA-sequencing dataset. Subsequently, we validated our predictions and elucidated the complexity of the isoforms using full-length transcript amplicon sequencing on patient-derived glioblastoma cells. However, analyses of the RNA-sequencing datasets of spatially mapped and longitudinally collected clinical tumor samples unveiled remarkable spatiotemporal heterogeneity of the candidate AS events. Furthermore, proteomics analysis did not reveal any peptide spectra matching the putative neoantigens. ConclusionsOur investigation illustrated the diverse characteristics of the tumor-specific AS events and the challenges of antigen exploration due to their notable spatiotemporal heterogeneity and elusive nature at the protein levels. Redirecting future efforts toward intracellular, MHC-presented antigens could offer a more viable avenue.

cancer biology↗

bioRxiv2023

Klebsiella Pneumoniae turns more virulent under flow stresses in capillary like microchannels

Fluidic habitats are very common to bacterial life, however, very little is known about the effect of the flow stresses on the virulence of the bacteria. In the present work, we conduct microfluidic experiments to understand the consequence of stresses generated by flowing fluid on the bacterial morphology and virulence. We consider Klebsiella pneumoniae (KP), an ESKAPE pathogen as the model bacteria that are responsible for blood stream infections like bacteremia apart from pneumonia, urinary tract infections and more. We generate four different stress conditions by changing the flow rate and channel geometry subsequently altering the shear rate and stressing time ({tau}). We observe significant changes in the structural aspects of the stressed bacteria. With an increase in stressing parameters, the viability of the bacterial sample deteriorated. Most importantly, these stressed samples proliferate much more than unstressed samples inside the RAW264.7 murine macrophages. The results shed light on the complex relationship between flow stresses and bacterial virulence. Furthermore, we challenge the bacterial samples with ciprofloxacin to see how they behave under different stress conditions. The present study can be extended to model deadly diseases like bacteremia using organ-on-a-chip technology and help understand bacterial pathogenicity under realistic environments. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=111 SRC="FIGDIR/small/558194v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@1111ea1org.highwire.dtl.DTLVardef@f20494org.highwire.dtl.DTLVardef@10ac86dorg.highwire.dtl.DTLVardef@ec6eb1_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOFigure:C_FLOATNO A schematic representation of the present work. Figure created with BioRender (www.biorender.com) C_FIG

biophysics↗

bioRxiv2023

Parallel, Continuous Monitoring and Quantification of Programmed Cell Death in Plant Tissue

The accurate quantification of hypersensitive response (HR) programmed cell death is imperative for understanding plant defense mechanisms and developing disease-resistant crop varieties. In this study, we report an accelerated phenotyping platform for the continuous-time, rapid and quantitative assessment of HR: Parallel Automated Spectroscopy Tool for Electrolyte Leakage (PASTEL). Compared to traditional HR assays, PASTEL significantly improves temporal resolution and has high sensitivity, facilitating the detection of microscopic levels of cell death. We validated PASTEL by transiently expressing the effector protein AVRblb2 in transgenic lines of the model plant Nicotiana benthamiana (expressing the corresponding resistance protein Rpi-blb2) to reliably induce HR. We were able to detect cell death at microscopic intensities, where leaf tissue appeared healthy to the naked eye one week after infiltration. PASTEL produces large amounts of frequency domain impedance data captured continuously (sub-seconds to minutes). Using this data, we developed a supervised machine learning models for classification of HR. We were able to classify input data (inclusive of our entire tested concentration range) as HR-positive or negative with 84.1% mean accuracy (F1 score = 0.75) at 1 hour and with 87.8% mean accuracy (F1 score = 0.81) at 22 hours. With PASTEL and the ML models produced in this work, it is possible to phenotype disease resistance in plants in hours instead of days to weeks.

plant biology↗