Biology subjects

Graham, E.

Publications and source records attributed to Graham, E..

8 recordsLinked to original sources

bioRxiv2025

Wee1 opposes APC/C(Cdh1) activity to promote S-phase entry

Wee1 phosphorylates and inhibits CDK activity to inhibit mitotic entry and establish a G2 DNA damage checkpoint. Consequently, Wee1 inhibitors are in clinical trials, developed to be synthetically lethal in TP53 mutant tumours that become reliant on a Wee1-mediated DNA damage checkpoint. However, Wee1 inhibitors have efficacy in TP53 wild-type tumours and many trials have been terminated due to high levels of toxic side-effects, suggesting that Wee1 has unknown functions. Here, we show that Wee1 promotes cell cycle re-entry from quiescence (G0) by opposing the activity of the E3 ubiquitin ligase, APC/CCdh1. Wee1 phosphorylates Cdh1 (FZR1) at key residues that mediate the interaction between Cdh1 and APC/C. Cells with loss-of-function of Wee1 during G0/G1 have delayed S-phase entry, an impaired G1/S transition, abnormal S-phase accumulation of the CDK inhibitor p21 and enter a p21-dependent G2 arrest. Reduced expression of APC/CCdh1 or p21 renders cells more sensitive to acute Wee1 inhibition and both pathways are downregulated in acquired Wee1 inhibitor resistance. Our study reveals a new cell cycle control mechanism that has implications for how Wee1 inhibitors should be used in the clinic.

cell biology↗

bioRxiv2025

CD8+ T cells mediate vaccination-induced lymphatic containment of latent Mycobacterium tuberculosis infection following immunosuppression, while B cells are dispensable

It is estimated that two billion people are latently infected with Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB). Latent Mtb infection (LTBI) can occur in multiple organs, including the lymphatics. The risk of LTBI reactivation increases in immunocompromised conditions, such as coinfection with human immunodeficiency virus (HIV), and during treatment of autoimmune diseases and organ transplantation. The immunological correlates of protection against TB, including against reactivation of LTBI, remain largely elusive. Here, we used a mouse model of latent lymphatic Mtb infection to dissect the immunological mechanisms underlying LTBI containment versus reactivation. We show that immunosuppression-mediated reactivation of lymphatic LTBI and the subsequent spread to non-lymphatic organs can be prevented by vaccination with multiple recombinant BCG (rBCG) strains despite the deficiency of CD4+ T cells. Using spatial transcriptomics, multi-parameter imaging, network analysis and bioinformatic integration of histopathological images, we reveal that immunosuppression is associated with a distinct repositioning of non-CD4 immune cells at the edge of TB lesions within the infection-draining cervical lymph nodes. While B cells increased in numbers, they are dispensable for the containment of LTBI. Lymphatic Mtb infection in different immune cell-deficient mouse strains, antibody-mediated cell depletion and adoptive transfer experiments into highly susceptible mice unequivocally show that vaccination-mediated prevention of LTBI reactivation is critically dependent on CD8+ T cells. These findings have profound implications for our understanding of immunity to TB and the management of LTBI. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=92 SRC="FIGDIR/small/634479v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@151fdd6org.highwire.dtl.DTLVardef@1efed47org.highwire.dtl.DTLVardef@c2e7e0org.highwire.dtl.DTLVardef@555c7_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2023

Collagen Tubular Airway-on-Chip for Extended Epithelial Culture andInvestigation of Ventilation Dynamics

The lower respiratory tract is a hierarchical network of compliant tubular structures that are made from extracellular matrix proteins with a wall lined by an epithelium. While microfluidic airway-on-a-chip models incorporate the effects of shear and stretch on the epithelium, week-long air-liquid-interface (ALI) culture remains limited to static conditions. The circular cross-section and substrate compliance associated with intact airways have yet to be recapitulated to allow studies of epithelial injuries under physiological and ventilation conditions. To overcome these limitations, we present a collagen tube-based airway model. Sustaining a functional human bronchial epithelium during two-week perfusion is accomplished by continuously supplying warm, humid air at the apical side and culture medium at the basal side. The model faithfully recapitulates human airways in size, composition, and mechanical microenvironment, allowing for the first time dynamic studies of elastocapillary phenomena associated with regular breathing as well as mechanical ventilation, along with the impact on epithelial cells. Findings reveal the epithelium to become increasingly damaged when subjected to repetitive collapse and reopening as opposed to overdistension and suggest expiratory flow resistance to reduce atelectasis. We expect the model to find broad potential applications in organ-on-a-chip applications for various tubular tissues.

bioengineering↗

bioRxiv2023

A latent cardiomyocyte regeneration potential in the human heart

Cardiomyocytes in the adult human heart show a regenerative capacity, with an annual renewal rate around 0.5%. Whether this regenerative capacity of human cardiomyocytes is employed in heart failure has been controversial. Using retrospective 14C birth dating we analyzed cardiomyocyte renewal in patients with end-stage heart failure. We show that cardiomyocyte generation is minimal in end-stage heart failure patients at rates 18-50 times lower compared to the healthy heart. However, patients receiving left ventricle support device therapy, who showed significant functional and structural cardiac improvement, had a >6-fold increase in cardiomyocyte renewal relative to the healthy heart. Our findings reveal a substantial cardiomyocyte regeneration potential in human heart disease, which could be exploited therapeutically.

cell biology↗

bioRxiv2022

Universal Amplicon Sequencing of North Imperial Valley Wetlands Microbiomes

DNA sequencing of complex microbial communities allows for the classification and quantification of thousands of distinct organisms in diverse environmental niches. We present a three domain "Universal Amplicon" (UA) method to simultaneously amplify DNA from the ribosomal small subunit locus from bacteria, archaea, and eukaryotes (and their organelles) using a single pair of amplification primers. We demonstrate the amenability of the UA to multiplexed Illumina library preparation and MiSeq-based sequencing. We validate the UA by sequencing a commercially available microbial community of known quantitative composition and through direct comparison to a shotgun metagenomics dataset. Following validation, we apply the UA to a time-course study of the wetlands of the Northern Imperial Valley in California and show substantial and variable microbial life in the Salton Sea and nearby waters. We demonstrate that the microbial ecology of the Salton Sea varies on at least a monthly basis and is distinct from the surrounding area. Finally, we contribute an open-source Shiny app for real-time analysis of complex metagenomic communities, with application to this study and far beyond.

ecology↗

bioRxiv2022

Alterations of pulmonary surfactant function by e-cigarette vapour

E-cigarette (EC) and vaping use continue to remain popular amongst teenage and young adult populations, despite several reports of vaping associated lung injury. This popularity is due in part to the vast variety of appealing flavours and nicotine concentrations easily accessible on the market. One of the first compounds that EC aerosols comes into contact within the lungs during a deep inhalation is pulmonary surfactant. This lipid protein mixture lines the alveoli, reducing surface tension and preventing alveolar collapse. Impairment of surfactants critical surface tension reducing activity can contribute to lung dysfunction. Currently, information on how EC aerosols impacts pulmonary surfactant remains limited. We hypothesized that exposure to EC aerosol impairs the surface tension reducing ability of surfactant. Bovine Lipid Extract Surfactant (BLES) was used as a model surfactant in a direct exposure syringe system. BLES (2ml) was placed in a syringe (30ml) attached to an EC. The generated aerosol was drawn into the syringe and then expelled, repeated 30 times. Biophysical analysis after exposure was completed using a constrained drop surfactometer (CDS). Minimum surface tensions increased after exposure to the EC aerosol. Variation in device used, addition of nicotine, or temperature of the aerosol had no additional effect. Two e-liquid flavours, menthol and red wedding, had further detrimental effects, resulting in higher surface tension than the vehicle exposed BLES. Alteration of surfactant properties through interaction with the produced aerosol was observed with a basic e-liquid vehicle, however additional compounds produced by added flavourings appeared to be able to increase inhibition. In conclusion, EC aerosols alter surfactant function through increases in minimum surface tension. This impairment may contribute to lung dysfunction and susceptibility to further injury.

biophysics↗

bioRxiv2022

Importance of Mobile Genetic Element Immunity in Numerically Abundant Trichodesmium Clades

The colony-forming cyanobacteria Trichodesmium spp. are considered one of the most important nitrogen-fixing genera in the warm, low nutrient, open ocean. Despite this central biogeochemical role, many questions about their evolution, physiology, and trophic interactions remain unanswered. To address these questions, we describe the genetic potential of the genus via significantly improved genomic assemblies of strains Trichodesmium thiebautii H94, Trichodesmium erythraeum 2175, and 17 new Trichodesmium metagenome-assembled genomes (MAGs, >50% complete) from hand-picked, Trichodesmium colonies spanning the Atlantic Ocean. Phylogenomics identified [~]four N2 fixing clades of Trichodesmium across the transect, with T. thiebautii dominating the colony-specific reads. Pangenomic analyses showed that all T. thiebautii MAGs are enriched in defense mechanisms and encode a vertically inherited Type III-B Clustered Regularly Interspaced Short Palindromic Repeats and associated protein-based immunity system (CRISPR-Cas hereafter). Surprisingly, this CRISPR-Cas system was absent in all T. erythraeum genomes and MAGs, vertically inherited by T. thiebautii, and correlated with increased signatures of horizontal gene transfer. Multiple lines of evidence indicate that the CRISPR-Cas system is functional in situ: 1. Trichodesmium CRISPR spacer sequences with 100% identical hits to field-assembled, putative phage genome fragments were identified, 2. High Trichodesmium spacer sequence variability indicating rapid adaptation, and 3. metaproteomic and transcriptomic expression analyses detecting the CRISPR-Cas system components in Trichodesmium colonies from the Atlantic and Pacific Oceans. These data suggest that phage or mobile genetic element immunity in T. thiebautii could contribute to their success, gene diversity, and numerical dominance over T. erythraeum in the oceans, thus warranting further Trichodesmium virome investigations. Significance statementOur work identifies CRISPR-Cas immunity as a phylogenetically distinct, environmentally expressed factor in the speciation of closely related N2-fixing Trichodesmium clades. These findings suggest that differential phage predation and resistance could be a previously overlooked selective pressure in the genus, potentially leading to the current numerical dominance of T. thiebautii over T. erythraeum in the oceans. Furthermore, while the currently CO2-limited T. erythraeum is expected to be a winner of anthropogenic climate change, their predicted higher phage sensitivity than T. thiebautii could challenge this outcome.

microbiology↗

bioRxiv2017

Undocumented Potential For Primary Productivity In A Globally-Distributed Bacterial Photoautotroph

Aerobic anoxygenic phototrophs (AAnPs) are common in the global oceans and are associated with photoheterotrophic activity. To date, AAnPs have not been identified in the surface ocean that possess the potential for carbon fixation. Using the Tara Oceans metagenomic dataset, we have reconstructed draft genomes of four bacteria that possess the genomic potential for anoxygenic phototrophy, carbon fixation via the Calvin-Benson-Bassham cycle, and the oxidation of sulfite and thiosulfate. Forming a monophyletic clade within the Alphaproteobacteria and lacking cultured representatives, the organisms compose minor constituents of local microbial communities (0.1-1.0%), but are globally distributed, present in multiple samples from the North Pacific, Mediterranean Sea, the East Africa Coastal Province, and the South Atlantic. These organisms represent a shift in our understanding of microbially-mediated photoautotrophy in the global oceans and provide a previously undiscovered route of primary productivity.\n\nSignificance StatementIn examining the genomic content of organisms collected during the Tara Oceans expedition, we have identified a novel clade within the Alphaproteobacteria that has the potential for photoautotrophy. Based on genome observations, these organisms have the potential to couple inorganic sulfur compounds as electron donors to fix carbon into biomass. They are globally distributed, present in samples from the North Pacific, Mediterranean Sea, East Africa Coastal Current, and the South Atlantic. This discovery may require re-examination of the microbial communities in the global ocean to understand and constrain the impacts of this group of organisms on the global carbon cycle.

microbiology↗