Biology subjects

Khalil, A.

Publications and source records attributed to Khalil, A..

At least 19 recordsLinked to original sources

bioRxiv2026

Discovery of the rosalexin pathway expands the modular network of maize diterpenoid chemical defenses.

The evolutionary expansion of specialized metabolism has shaped the ability of plants to adapt to combined pathogen, pest, and other environmental pressures. For instance, the duplication and divergence of ancestral gibberellin pathway genes have given rise to specialized kauralexin and dolabralexin diterpenoids in maize (Zea mays) that serve as core components of disease resistance and stress adaptation. Here, we describe the biosynthesis and elicited production of rosalexins as a previously unrecognized component of the maize chemical defense network. By integrating genomics-enabled gene discovery, combinatorial enzyme assays, and AI-assisted enzyme mechanistic studies we show that maize rosalexin biosynthesis proceeds via a distinct 5-rosanol scaffold formed by the pairwise activity of two diterpene synthases, ZmTPS38/CPS2/AN2 and ZmTPS42/KSL1, recruited from gibberellin metabolism. Further oxygenation by the promiscuous P450 enzyme, ZmCYP71Z18, yields epoxyrosanol that, in turn, can undergo epoxide ring opening to form trihydroxyrosanol. Epoxyrosanol, but not 5-rosanol or trihydroxyrosanol, display strong inhibitory activity on fungal pathogen growth in vitro, highlighting the contribution of the epoxide group to antibiotic efficacy. Large variation in rosalexin presence and abundance exists across maize genotypes due to expansive ZmTPS42/KSL1 gene sequence variation and pseudogenization. Transcriptomics and targeted metabolomics demonstrated the pathogen-elicited accumulation of rosalexins in maize lines featuring functional ZmTPS42/KSL1 genes. However, no dominant pathogen resistance phenotype was observed in association with rosalexin abundance. These collective findings expand our knowledge of how multiple interconnected diterpenoid pathways arose in maize via duplication of hormone-metabolic genes and enable the utilization of a common precursor to form modular chemical defense layers. Significance StatementPlant diterpenoids play critical roles in crop development, stress defense and ecological adaptation. In maize, diterpenoids serve as key components of chemical defenses against pests and diseases with direct impact on crop immunity and vigor. Enzymes of the diterpene synthase and cytochrome P450 families largely drive diterpenoid chemical diversity. This study reports the discovery and characterization of the pathway forming rosalexin diterpenoids in maize. Pathogen-elicited accumulation and in vitro antifungal activity of rosalexin metabolites supports a physiological function in maize chemical defense.

plant biology↗

bioRxiv2026

Heatwave winners and losers: cryptic coral holobionts differ in thermal tolerance

Extreme climatic events are reshaping ecosystems worldwide as individual organisms vary markedly in their ability to withstand these disturbances. Deciphering patterns of persistence on local scales is therefore critical for predicting biodiversity trajectories under intensifying climate extremes. In this study, we examined variation in thermal stress responses among individuals of the coral Stylophora pistillata species complex during a heatwave at Heron Island Reef, Australia. More than half of the focal coral colonies died on the reef, and survival of coral fragments maintained under ex situ common thermal stress conditions was significantly correlated with the survival of their source colony. This demonstrates that survival differences result largely from biological factors rather than differential thermal exposure across reef habitats. Under common garden conditions, we observed striking differences in bleaching severity and survival times among three sympatric cryptic taxa and their highly host-specific symbiont community. Within the most locally common taxon, corals from historically warmer and more seasonally variable reef habitats seem more susceptible to bleaching, contrary to expectations. Together, these results reveal how biological differences among cryptic taxa and among individuals can shape coral responses during a heatwave and advance our understanding of coral vulnerability in a rapidly warming world.

ecology↗

bioRxiv2025

Feeder-free generation of functional dendritic cells from human pluripotent stem cells

The scarcity of primary conventional dendritic cells (cDCs) and the limited effectiveness of monocyte-derived dendritic cells (moDCs) have long hindered progress in human dendritic cell research and immunotherapy. We developed a feeder-free differentiation platform that generates CD1c+CD141+ hPSC-cDCs phenotypically aligned with CD141+ tissue-resident cDC2 subsets found in human tissues. We further optimized the differentiation process using a Design-of-Experiments framework to refine cytokine and serum conditions, enhancing differentiation efficiency while reducing cytokine demand. These hPSC-cDCs exhibit efficient antigen uptake, defined cytokine responses, and robust priming of antigen-specific CD8+ T cell proliferation and effector differentiation, outperforming moDCs in direct comparison. Together, this work establishes a robust, and generalizable platform for mechanistic studies and translational development of dendritic cell-based vaccines and standardized ex vivo T cell expansion.

immunology↗

bioRxiv2025

Structural and functional basis of PU.1-BAF interaction enables targeting of lineage-specific transcription

Chromatin remodeling complexes like BAF finely regulate transcriptional programs by working in concert with transcription factors. However, evidence is lacking as to whether TFs interact directly with BAF and if so, what the mechanistic and structural principles governing these critical interactions are. Here, we establish direct engagement between a crucial and therapeutically relevant full-length human TF, PU.1 (SPI1), and BAF. Within this 1MDa+ complex, we precisely map the binding site of PU.1 to a YEATS-like domain on BAF60A and elucidate the structure of the PU.1-BAF60A complex. This work reveals that upon binding to BAF, a disordered region within the TF adopts a helical conformation, and that disruption of this functionally critical interface via knockdown abrogates the ability of PU.1 to rescue cell viability. To explore the druggability of TF-BAF protein-protein interactions (PPIs), we conducted a high-throughput screen that identified small molecules capable of disrupting the PU.1-BAF60A PPI by binding to BAF60A. Co-crystal structures reveal distinct compound binding modes that converge on a critical PU.1-BAF60A interaction hotspot. These findings define, for the first time, the structural interface between a human TF and a chromatin remodeling complex and establish a platform that enables the targeting of these interactions, a novel mechanism in cancer therapeutics.

biochemistry↗

bioRxiv2025

In Vivo Multiplexed Modeling Reveals Diverse Roles of the TBX2 Subfamily and Egr1 in Ras-Driven Lung Adenocarcinoma

The TBX2 subfamily of T-box transcription factors (including Tbx2, Tbx3, Tbx4, Tbx5) plays an essential role in lung development. Downregulation of these genes in human Lung adenocarcinoma (LUAD) suggests that these genes may be tumor suppressive, however because downregulation appears to occur primarily via epigenetic change, it remains unclear if these changes causally drive tumor progression or are merely the consequence of upstream events. Herein, we developed the first multiplexed mouse model to study the impact of TBX2 subfamily loss, alongside associated signaling genes Egr1, Chd2, Tnfaip3a, and Atf3, in Ras-driven lung cancer. Using TuBa-seq, a high-throughput tumor-barcoding system, we quantified the growth effects of these knockouts during early and late tumorigenesis. Chd2 loss consistently suppressed tumor progression, while Tbx2 loss exhibited stage-dependent effects. Notably, Egr1 emerged as a potent tumor suppressor, with its knockout increasing tumor size ([~]5x) at 20 weeks, surpassing Rb1 loss. Transcriptomic analyses of Egr1-deficient tumors suggested immune dysregulation, including heightened inflammation and potential markers of T cell exhaustion in the tumor microenvironment. These findings indicate that Egr1 may play a role in suppressing tumor growth through modulating immune dynamics, offering new insights into the interplay between tumor progression and immune regulation in LUAD.

cancer biology↗

bioRxiv2025

A novel human blood-brain barrier model reveals pericytes as critical regulators of viral neuroinvasion

The blood-brain barrier (BBB) plays a vital role in regulating the passage of biomolecules between the bloodstream and the central nervous system (CNS) while also protecting the CNS from pathogens. Pericytes reside at the interface between the endothelial cells that form the vessel walls and the brain parenchyma. These cells are critical for maintaining BBB integrity and play key roles in regulating vessel permeability, blood flow, and immune cell migration. In this study, we developed a novel serum-free protocol to generate neural crest cell-derived pericytes (NCC-PCs) from human pluripotent stem cells (hPSCs). These NCC-PCs enhance BMEC barrier function and can be co-cultured with hPSC-derived brain microvascular endothelial cells (BMECs) in a contact co-culture BBB model that recapitulates the in vivo cellular interactions at the BBB. We used this model to evaluate the pathological consequences of BBB cell infection by highly neuroinvasive flaviviruses. Our results identify a previously undescribed role for NCC-PCs in maintaining BMEC barrier integrity during infection and reducing the spread of viral infection to the CNS.

microbiology↗

bioRxiv2024

The safety and efficacy of Ultrasound Histotripsy and human pluripotent stem cell-derived hepatic spheroid implantation as a potential therapy for treatment of congenital metabolic liver disease: assessment in an immunocompetent rodent model.

BackgroundLiver disease secondary to an inborn or genetic error of metabolism is a rare group of conditions often associated with chronic ill health and reduced survival. Curative treatment is mainly limited to liver transplantation with major long-term risks. Cell therapy is a promising alternative, but current approaches are ineffective. AimTo develop histotripsy, a non-invasive high intensity ultrasound procedure for liver tissue mechanical ablation, combined with hepatocyte stem cell implantation as a novel method of reversing liver failure from genetic disease. This study assessed the safety and feasibility of this approach in healthy rodents. MethodsUnder general anaesthesia, adult rats (n=12) underwent laparotomy and ultrasound histotripsy to the exposed liver. Around 1 million cells were injected into a single histotripsy cavity in each animal under direct vision (n=10) with two receiving only histotripsy without cell injection. On completion of cell implant haemostasis was secured, laparotomy incision closed, and the animals recovered. Groups of animals were terminated immediately and after 4 hrs, 8 hrs, 24hrs, 4 days and 7 days. Liver and vital organs were assessed for procedure related injuries and evidence of viable implanted cells by histology and immunohistochemistry. ResultsAll animals successfully recovered, and no complication was observed throughout the study. Created cavities were successfully identified in histological analysis of rat. The presence of human cells was verified using anti-human nuclei antibody confirming successful implantation of liver organoids into decellularized cavities. ConclusionsIn this feasibility study, we demonstrated suitability of histotripsy to create decellularized cavities in liver parenchyma. In addition, feasibility of direct transplantation of undissociated liver organoids into the created cavities was demonstrated as a potential approach to treat inborn liver disease by creating nodules of healthy cells capable of performing loss metabolic function. Therapeutic efficacy of this approach will be evaluated in an upcoming study.

bioengineering↗

bioRxiv2024

Method for determining of cytotoxicity based on the release of fluorescent proteins

This paper describes a method for determining the cytotoxicity of chemical compounds based on the detection of fluorescent proteins - in this case, green fluorescent protein (GFP) and red fluorescent protein (RFP), which are released into the medium from dead cells. This method is similar in principle to the lactate dehydrogenase test (LDH test), but it does not require a reaction with a chromogenic substrate. This method also makes it possible to independently determine the viability of different lines when used in cocultures. Experiments were preformed on a classical monolayer, spheroids and 3D cultures in alginate hydrogel. Capecitabine was used as a model cytotoxic agent. We included liver cells (Huh7) in a coculture model and determined changes in the cytotoxicity levels of capecitabine against NCI-H1299 cells. The experimental part also found that there were differences in sensitivity to capecitabine depending on the type of 3D cultures used.

pharmacology and toxicology↗

bioRxiv2024

The DiffInvex evolutionary model for conditional somatic selection identifies chemotherapy resistance genes in 10,000 cancer genomes

Tumors often show an initial response to chemotherapy, but then develop resistance, leading to relapse and poor prognosis. We hypothesized that a genomic comparison of mutations in pre-treated versus treatment-naive tumors would serve to identify genes that confer resistance. A challenge in such an analysis is that therapy alters mutation burdens and signatures, confounding association studies and complicating identifying causal, selected mutations. We developed DiffInvex, a framework for identifying changes in selection acting on individual genes in somatic genomes. Crucially, DiffInvex draws on a mutation rate baseline that accounts for these shifts in neutral mutagenesis during cancer evolution. We applied DiffInvex to 9,953 cancer whole-genomes from 29 cancer types from 8 studies, containing both WGS of treatment-naive tumors and tumors pre-treated by various drugs, identifying genes where point mutations are under conditional positive or negative selection for a certain chemotherapeutic, suggesting resistance mechanisms occurring via point mutation. DiffInvex confirmed well-known chemoresistance-driver mutations in EGFR, ESR1, KIT and AR genes as being under conditional positive selection, with additional cancer types identified for EGFR and KIT. Additionally, DiffInvex identified 11 genes with treatment-associated selection for different classes of therapeutics. In most cases, these genes were common cancer genes including PIK3CA, APC, MAP2K4 and MAP3K1. This suggests that tumor resistance to therapy via mutation often occurs via selective advantages conferred by known driver genes, rather than via mutations in specialized resistance genes. Various gene-chemotherapy associations were further supported in tests for functional impact of mutations, again implemented in a conditional selection setting, as well as replicating in independent panel or exome sequencing data. In addition to nominating drug resistance genes that could be targeted by future therapeutics, DiffInvex can also be applied to diverse analysis in cancer evolution, such as comparing normal and tumoral tissues, or analyzing subclonal evolution, identifying changes in selection over time.

genomics↗

bioRxiv2024

Sex-based difference in immune responses and efficacy of the pneumococcal conjugate vaccine

Vaccine-mediated protection and susceptibility to Streptococcus pneumoniae (pneumococcus) infections are influenced by biological sex. The incidence of invasive pneumococcal disease remains higher in males compared to females even after the introduction of the pneumococcal conjugate vaccine (PCV). However, sex-based differences in the immune response to this conjugate vaccine remain unexplored. To investigate those differences, we vaccinated adult male and female mice with PCV and assessed cellular and humoral immune responses. Compared to females, male mice displayed lower levels of T follicular helper cells, germinal center B cells and plasmablasts, which are all required for antibody production following vaccination. This was linked to lower IgG and IgM levels against pneumococci and lower isotype switching to IgG3 in vaccinated males. Additionally, sera of vaccinated male mice had lower efficacy in several anti-pneumococcal functions including neutralization of bacterial binding to pulmonary epithelial cells as well as direct cytotoxicity against S. pneumoniae. Importantly, while the vaccine was highly protective in females, vaccinated males succumbed to infection more readily and were more susceptible to both lung-localized infection and systemic spread following S. pneumoniae challenge. These findings identify sex-based differences in immune responses to PCV that can inform future vaccine strategies.

immunology↗

bioRxiv2023

SARS-CoV-2 infection of human pluripotent stem cell-derived vascular cells reveals smooth muscle cells as key mediators of vascular pathology during infection

Although respiratory symptoms are the most prevalent disease manifestation of infection by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), nearly 20% of hospitalized patients are at risk for thromboembolic events. This prothrombotic state is considered a key factor in the increased risk of stroke, which is observed clinically during both acute infection and long after symptoms clear. Here we develop a model of SARS-CoV-2 infection using human-induced pluripotent stem cell-derived endothelial cells (ECs), pericytes (PCs), and smooth muscle cells (SMCs) to recapitulate the vascular pathology associated with SARS-CoV-2 exposure. Our results demonstrate that perivascular cells, particularly SMCs, are a susceptible vascular target for SARS-CoV-2 infection. Utilizing RNA sequencing, we characterize the transcriptomic changes accompanying SARS-CoV-2 infection of SMCs, PCs, and ECs. We observe that infected SMCs shift to a pro-inflammatory state and increase the expression of key mediators of the coagulation cascade. Further, we show human ECs exposed to the secretome of infected SMCs produce hemostatic factors that contribute to vascular dysfunction, despite not being susceptible to direct infection. The findings here recapitulate observations from patient sera in human COVID-19 patients and provide mechanistic insight into the unique vascular implications of SARS-CoV-2 infection at a cellular level.

microbiology↗

bioRxiv2023

The G-alpha Gpa1 directs septin localization in the mating projection of Saccharomyces cerevisiae through its Ubiquitination Domain and Endocytic Machinery

The septin cytoskeleton plays a key role in the morphogenesis of the yeast mating projection, forming structures at the base of the projection. The yeast mating response uses the G-protein coupled receptor (GPCR), Ste2, to detect mating pheromone and initiate mating projection morphogenesis. Desensitization of the G, Gpa1, by the Regulator of G-protein Signaling (RGS), Sst2, is required for proper septin organization and morphogenesis. We hypothesized that Gpa1 would utilize known septin regulators to control septin organization. We found that single deletions of the septin chaperone Gic1, the Cdc42 GAP Bem3, and the endocytic adaptor proteins Ent1 and Ent2 rescued the polar cap accumulation of septins in the hyperactive G. We hypothesized that hyperactive G might increase the rate of endocytosis of a pheromone-responsive cargo, thereby altering where septins are localized. Mathematical modeling predicted that changes in endocytosis could explain the septin organizations we find in WT and mutant cells. Our results show that Gpa1-induced disorganization of septins requires clathrin-mediated endocytosis. Both the GPCR and the G are known to be internalized by clathrin-mediated endocytosis during the pheromone response. Deletion of the GPCR C-terminus to block internalization partially rescued septin organization. However, deleting the Gpa1 ubiquitination domain required for its endocytosis completely abrogated septin accumulation at the polarity site. Our data support a model where the location of endocytosis serves as a spatial mark for septin structure assembly and that desensitization of the G delays its endocytosis sufficiently that septins are placed peripheral to the site of Cdc42 polarity.

cell biology↗

bioRxiv2023

Tracking the progression of Alzheimer's disease with peripheral blood monocytes

BackgroundAlzheimers disease (AD) is the most common form of dementia with the symptoms gradually worsening over the years. However, the driving pathological processes occur well before the appearance of symptoms. AD patients display signs of systemic inflammation, suggesting that it could precede the well-established AD hallmarks. We recently showed that the innate immune response in the form of monocyte activation is detectable at the pre-clinical stage. ObjectivesOur goal here is to characterize changes of gene expression in peripheral blood monocytes from patients at different stages of AD progression and validate potential biomarkers for a better prognosis and diagnosis of AD clinical spectrum. ResultsWe performed a whole transcriptome analysis on monocytes purified from healthy subjects, Mild Cognitive Impairment (MCI) and AD patients, and established the list of genes differentially expressed in monocytes during the disease evolution. We observed that, in the top 500 genes differentially expressed, a majority of these genes were upregulated (65%) during AD progression. These genes are mainly involved in chemokine/cytokine-mediated signaling pathways. We further confirmed several biomarkers by quantitative PCR and immunoblotting and showed that they are often deregulated at pre-clinical stages of the disease (MCI stage), supporting the hyperactivation of monocytes in MCI patients. PerspectivesOur findings provide evidence that the pre-clinical stage of AD can be detected in monocytes using a specific set of biomarkers, highlighting the importance to study the early innate immune response in AD. Our results open the possibility to use these biomarkers with different diagnostic methodologies to better predict and efficiently treat AD.

neuroscience↗

bioRxiv2022

A dolabralexin-deficient mutant provides insight into specialized diterpenoid metabolism in maize (Zea mays).

Two major groups of maize (Zea mays) specialized metabolites, termed kauralexins and dolabralexins, serve as known or predicted diterpenoid defenses against pathogens, herbivores, and other environmental stressors. To consider physiological roles of the recently discovered dolabralexin pathway, we examined dolabralexin structural diversity, tissue-specificity, and stress-elicited production in a defined biosynthetic pathway mutant. Metabolomics analyses support a larger number of dolabralexin pathway products than previously known. We identified dolabradienol as a previously undetected pathway metabolite and characterized its enzymatic production. Transcript and metabolite profiling showed that dolabralexin biosynthesis and accumulation predominantly occurs in primary roots and shows quantitative variation across genetically diverse inbred lines. Generation and analysis of CRISPR-Cas9-derived loss-of- function Kaurene Synthase-Like 4 (Zmksl4) mutants demonstrated dolabralexin production deficiency, thus supporting ZmKSL4 as the diterpene synthase responsible for the conversion of geranylgeranyl pyrophosphate precursors into dolabradiene and downstream pathway products. Zmksl4 mutants further display altered root-to-shoot ratios and root architecture in response to water deficit, consistent with an interactive role of maize dolabralexins in plant vigor during abiotic stress.

plant biology↗

bioRxiv2022

Single extracellular vesicle analysis in human amniotic fluid shows evidence of phenotype alterations in preeclampsia

Amniotic fluid surrounding the developing fetus is a complex biological fluid rich in metabolically active bio-factors. The presence of extracellular vesicles (EVs) in amniotic fluid has been mainly related to fetal urine. We here characterized EVs from term amniotic fluid in terms of surface marker expression using different orthogonal techniques. EVs appeared to be a heterogeneous population expressing markers of renal, placental, epithelial and stem cells. Moreover, we compared amniotic fluid EVs from normal pregnancies with those of preeclampsia, a hypertensive disorder affecting up to 8% of pregnancies worldwide. An increase of CD105 (endoglin) expressing EVs was observed in preeclamptic amniotic fluid by bead-based cytofluorimetric analysis, and further confirmed using a chip-based analysis. HLA-G, a typical placental marker, was not co-expressed by the majority of CD105+ EVs, suggesting their origin from amniotic fluid cells. At a functional level, preeclampsia-derived EVs, but not normal pregnancy EVs, showed an antiangiogenic effect, possibly due to the decoy effect of endoglin. In addition, several miRNAs were differentially expressed in preeclampsia-derived EVs and directly related to the modulation of angiogenesis and trophoblast function. Our results provide a characterization of term amniotic fluid-EVs, supporting their origin from fetal and placental cells. In preeclampsia, the observed antiangiogenic characteristics of amniotic fluid-EVs may reflect the hypoxic and antiangiogenic microenvironment and could possibly impact on the developing fetus or on the surrounding fetal membranes.

molecular biology↗

bioRxiv2020

SARS-CoV-2 RNA reverse-transcribed and integrated into the human genome

Prolonged SARS-CoV-2 RNA shedding and recurrence of PCR-positive tests have been widely reported in patients after recovery, yet these patients most commonly are non-infectious1-14. Here we investigated the possibility that SARS-CoV-2 RNAs can be reverse-transcribed and integrated into the human genome and that transcription of the integrated sequences might account for PCR-positive tests. In support of this hypothesis, we found chimeric transcripts consisting of viral fused to cellular sequences in published data sets of SARS-CoV-2 infected cultured cells and primary cells of patients, consistent with the transcription of viral sequences integrated into the genome. To experimentally corroborate the possibility of viral retro-integration, we describe evidence that SARS-CoV-2 RNAs can be reverse transcribed in human cells by reverse transcriptase (RT) from LINE-1 elements or by HIV-1 RT, and that these DNA sequences can be integrated into the cell genome and subsequently be transcribed. Human endogenous LINE-1 expression was induced upon SARS-CoV-2 infection or by cytokine exposure in cultured cells, suggesting a molecular mechanism for SARS-CoV-2 retro-integration in patients. This novel feature of SARS-CoV-2 infection may explain why patients can continue to produce viral RNA after recovery and suggests a new aspect of RNA virus replication.

genomics↗

bioRxiv2020

RIT1: A Novel Biomarker in Glioblastoma

Glioblastoma is the most common type of malignant brain tumors and the most feared cancer among adults. Its high invasiveness and the lack of successful therapies are responsible for the poor prognosis among these patients. A comprehensive understanding for the early molecular mechanisms in glioblastoma would definitely enhance its diagnosis and treatment strategies. Based on in vitro and in vivo models, it was recently postulated that the deregulated expression of key genes that are known to be involved in early neurogenesis could be the instigator of brain tumorigenesis. RIT1 (Ras Like Without CAAX 1) which encodes an unusual "orphan" GTPase protein belongs to this category of critical genes involved in controlling sequential proliferation and differentiation of adult hippocampal neural progenitor cells. As such, we surveyed RIT1 expression and its potential role in glioblastoma by in-silico means. Our results revealed a significant and progressive upregulation of RIT1 expression in various publicly available datasets. RIT1 expression ranked among the top upregulated genes in glioblastoma cohorts and correlated with poor overall survival. Genetic and epigenetic analysis of RIT1 didnt reveal significant aberrations that could underlie its deregulated expression. Moreover, our results pointed on RIT1 transcriptional activity that could be significantly controlled by STAT3, one of the main players in the onset of glioblastoma. In conclusion, our results are the first proof for the role of RIT1 as an oncogene in glioblastoma paving the way for its potential use as a biomarker and therapeutic target as is the case of the other members in the RAS family.

cancer biology↗

bioRxiv2020

Genetically Encoded, Multivalent Liquid Glycan Array (LiGA)

The Central Dogma of Biology does not allow for the study of glycans using DNA sequencing. We report a "Liquid Glycan Array" (LiGA) platform comprising a library of DNA barcoded M13 virions that display 30-1500 copies of glycans per phage. A LiGA is synthesized by acylation of phage pVIII protein with a dibenzocyclooctyne, followed by ligation of azido-modified glycans. Pulldown of the LiGA with lectins followed by deep sequencing of the barcodes in the bound phage decodes the optimal structure and density of the recognized glycans. The LiGA is target agnostic and can measure the glycan-binding profile of lectins such as CD22 on cells in vitro and immune cells in a live mouse. From a mixture of multivalent glycan probes, LiGAs identifies the glycoconjugates with optimal avidity necessary for binding to lectins on living cells in vitro and in vivo; measurements that cannot be performed with canonical glass slide-based glycan arrays. DedicationThe paper is dedicated to Laura L. Kiessling on the occasion of her 60th birthday.

biochemistry↗