Biology subjects

Boyle, N.

Publications and source records attributed to Boyle, N..

9 recordsLinked to original sources

bioRxiv2026

Dual-targeted NADK2 Links Mitochondrial Redox Homeostasis to Carbon Partitioning and Heterotrophic Growth in Chlamydomonas reinhardtii

NAD kinases (NADKs) can modulate NADP(H) levels across shifting environmental conditions. In Chlamydomonas reinhardtii, we localized CreNADK1 to the chloroplast, CreNADK3 to the cytoplasm, and CreNADK2 to both the chloroplast and mitochondria. Importantly, during mixotrophic (light + acetate) and photoautotrophic (light) growth, most CreNADK2 is chloroplast localized whereas an increased proportion resides in mitochondria when the cells are grown heterotrophically (dark + acetate). In Crenadk2 mutants, a diminished ability to synthesize mitochondrial NADP(H) inhibits the TCA cycle and markedly retards heterotrophic growth. In the light, photosynthesis becomes the primary energy source, and the presence of chloroplastic CreNADK1 enables near-normal growth in mutant cells, with inhibition of TCA cycle activity partially ameliorated via rerouting of acetate metabolism through the glyoxylate shunt, bypassing the TCA cycle and diminishing mitochondrial ROS production. These findings demonstrate spatial plasticity of NAD(H)Ks and the implementation of bypass metabolism to enable growth of Crenadk2 mutants.

cell biology↗

bioRxiv2026

BeeMonitor: Automated IoT video surveillance and an AI-powered video processing system for monitoring the foraging and nesting behavior of cavity-nesting solitary bees

O_LISolitary bee species that use artificial trap nests are important for agricultural crop production and as indicators of habitat quality. Quantifying cavity-nesting solitary bee foraging and nesting behavior is essential for real-time analysis of population numbers and pollination activity, as well as understanding how environmental conditions shape reproductive success and population dynamics. However, manual observation is labor-intensive, prone to observer bias, and unable to deliver continuous data. Existing automated systems either require individual bee marking or detect presence without resolving nest-tube-level entry and exit events. C_LIO_LIWe developed BeeMonitor, an integrated hardware and computer-vision pipeline that detects nest entry and exit events in cavity-nesting solitary bees from continuous video, using Osmia cornifrons (the horn-faced mason bee) as a model system. A low-cost Raspberry Pi handles solar-powered field recording, while the software combines object detection (YOLOv26), a custom multiple-object tracker (BeeTrack), and a Random Forest classifier trained on trajectory-derived features to distinguish genuine events from incidental detections. C_LIO_LIOver a 29-day deployment, hardware reliability averaged 97.5% recording coverage. The pipeline achieved 91.3% precision and 87.3% recall (F1 = 0.893), generalizing robustly under leave-one-video-out cross-validation (mean F1 = 0.904). Detected foraging trips correlated strongly with brood cell counts (R2 = 0.849, p < 0.001, n = 19), and a Random Forest model (AUC = 0.820) identified solar radiation as the dominant driver of foraging activity, followed by temperature. C_LIO_LIBeeMonitor demonstrates that automated computer vision can reliably extract ecologically relevant behavioral data from continuous video, enabling real-time analysis of pollinator behavior and abundance at a temporal and spatial resolution unattainable through manual observation. Its modular design supports adaptation to other species and monitoring contexts. C_LI

animal behavior and cognition↗

bioRxiv2025

Investigating Overflow Metabolism in Heterotrophic Cultures of the Green Alga Chromochloris zofingiensis

Chromochloris zofingiensis is of interest for its ability to perform a reversible trophic switch in the presence of glucose that is characterized by a shutdown of photosynthesis and an accumulation of energy storage metabolites. Previous work has shown that this trophic switch is accompanied by overflow metabolism and the production of lactate in aerobic conditions. This trophic switch is not observed in nutrient replete media. We utilized isotopically assisted metabolic flux analysis to characterize intracellular flux distributions that are associated with different metabolic phenotypes observed in this organism in different media formulations in light and dark conditions. The results of this analysis showed differences in flux through carbon fixation reactions, the TCA cycle and through the reaction catalyzed by pyruvate kinase. This analysis was complemented with transcriptomics data collected for C. zofingiensis grown in iron limited conditions to provide further evidence towards the negative impact of iron limitation on both photosynthetic and respiratory activity. Overflow metabolism allows this alga to compensate for the lower energy production that results from iron limitation. This work highlights how nutrient availability can lead to drastic changes in the metabolism of C. zofingiensis.

systems biology↗

bioRxiv2025

High-Throughput Antibody Neutralization Screening in Massively Parallel Droplet Arrays

Neutralizing antibodies provide rapid immune defense against infectious diseases, but are difficult to discover at scale because neutralization assays require live reporter cells and soluble monoclonal antibodies. Here we report Droplet Reporter Cell Testing for Neutralization (DrReCT-Neutralization) to screen antibody gene libraries for their ability to neutralize viral infections. We established the necessary engineered cell lines and validated the DrReCT screening platform using synthetic oligoclonal libraries, followed by an example discovery campaign that demonstrated scalable functional antibody data collection against viral diseases.

immunology↗

bioRxiv2022

Investigating the unique ability of Trichodesmium to fix carbon and nitrogen simultaneously using MiMoSA

The open ocean is an extremely competitive environment, partially due to the dearth of nutrients. Trichodesmium erythraeum, a marine diazotrophic cyanobacterium, is a keystone species in the ocean due to its ability to fix nitrogen and leak 30-50% into the surrounding environment, providing a valuable source of a necessary macronutrient to other species. While there are other diazotrophic cyanobacteria that play an important role in the marine nitrogen cycle, Trichodesmium is unique in its ability to fix both carbon and nitrogen simultaneously during the day without the use of specialized cells called heterocysts to protect nitrogenase from oxygen. Here, we use the advanced modeling framework called Multiscale Multiobjective Systems Analysis (MiMoSA) to investigate how Trichodesmium erythraeum can reduce dimolecular nitrogen to ammonium in the presence of oxygen. Our simulations indicate that nitrogenase inhibition is best modeled as Michealis Menten competitive inhibition and that cells along the filament maintain microaerobia using high flux through Mehlers reactions in order to protect nitrogenase from oxygen. We also examined the effect of location on metabolic flux and found that cells at the end of filaments operate in distinctly different metabolic modes than internal cells despite both operating in a photoautotrophic mode. These results give us important insight into how this species is able to operate photosynthesis and nitrogen fixation simultaneously, giving it a distinct advantage over other diazotrophic cyanobacteria because they can harvest light directly to fuel the energy demand of nitrogen fixation. IMPORTANCETrichodesmium erythraeum is a marine cyanobacterium responsible for approximately half of all biologically fixed nitrogen, making it an integral part of the global nitrogen cycle. Interestingly, unlike other nitrogen fixing cyanobacteria, Trichodesmium does not use temporal or spatial separation to protect nitrogenase from oxygen poisoning; instead, it operates photosynthesis and nitrogen fixation reactions simultaneously during the day. Unfortunately, the exact mechanism the cells utilize to operate carbon and nitrogen fixation simultaneously is unknown. Here, we use an advanced metabolic modeling framework to investigate and identify the most likely mechanisms Trichodesmium uses to protect nitrogenase from oxygen. The model predicts that cells operate in a microaerobic mode, using both respiratory and Mehler reactions to dramatically reduce intracellular oxygen concentrations.

systems biology↗

bioRxiv2022

Chlamydomonas reinhardtii triose-phosphate/phosphate translocator3 (TPT3): A major chloroplast shunt for the export of fixed carbon and reductant

Modulation of export of photoassimilates from the chloroplast is essential for controlling the distribution of fixed carbon in the cell and maintaining optimum photosynthetic rates. In this study we identified chloroplast triose phosphate/phosphate translocators 2 and 3 (CreTPT2 and CreTPT3) in the green alga Chlamydomonas reinhardtii that exhibited similar substrate specificities but were differentially expressed over the diel cycle. We focused mostly on analyzing CreTPT3 because of its high level of expression and the severe phenotype exhibited by tpt3 relative to the tpt2 mutants. Null mutants for CreTPT3 had a pleiotropic phenotype that impacted growth, photosynthetic activities, metabolite profiles, carbon partitioning, and organelle-specific accumulation of H2O2. These analyses demonstrated that CreTPT3 is a dominant conduit on the chloroplast envelope for the transport of photoassimilate. In addition, CreTPT3 can serve as a safety valve that moves excess reductant out of the chloroplast and appears to be essential for preventing the cells from experiencing oxidative stress and accumulating of reactive oxygen species, even under low/moderate light intensities. Finally, our studies indicate subfunctionalization of the CreTPT transporters and suggest that there are differences in managing the export of photoassimilates from the chloroplasts of Chlamydomonas and vascular plants.

plant biology↗

bioRxiv2022

Rhythm of The Night (and Day): Predictive metabolic modeling of circadian growth in Chlamydomonas

Algal cells experience strong circadian rhythms under diurnal light, with regular changes in both biomass composition and transcriptomic environment. However, most metabolic models - critical tools for bioengineering organisms - assume a steady state. The conflict between these assumptions and the reality of the cellular environment make such models inappropriate for algal cells, creating a significant obstacle in engineering cells that are viable under natural light. By transforming a set of discreet transcriptomic measurements from synchronized Chlamydomonas cells grown in a 12/12 diel light regime (1) into continuous curves, we produced a complete representation of the cells transcriptome that can be interrogated at any arbitrary timepoint. We clustered these curves, in order to find genes that were expressed in similar patterns, and then also used it to build a metabolic model that can accumulate and catabolize different biomass components over the course of a day. This model predicts qualitative phenotypical outcomes for the sta6 mutant, including excess lipid accumulation (2) and a failure to thrive when grown diurnally in minimal media (3), representing a qualitative prediction of phenotype from genotype even under dynamic conditions. We also extended this approach to simulate all single-knockout mutants with genes represented in the model and identified potential targets for rational engineering efforts. SIGNIFICANCE STATEMENTWe have developed the first transient metabolic model for diurnal growth of algae based on experimental data and capable of predicting phenotype from genotype. This model enables us to evaluate the impact of genetic and environmental changes on the growth, biomass composition and intracellular fluxes of the model green alga, Chlamydomonas reinhardtii. The availability of this model will enable faster and more efficient design of cells for production of fuels, chemicals and pharmaceuticals.

systems biology↗

bioRxiv2021

Immortalization and Functional Screening of Natively Paired Human T Cell Receptor Repertoires

Functional analyses of the T cell receptor (TCR) landscape can reveal critical information about protection from disease and molecular responses to vaccines. However, it has proven difficult to combine advanced next-generation sequencing technologies with methods to decode the peptide-major histocompatibility complex (pMHC) specificity of individual TCRs. Here we developed a new high-throughput approach to enable repertoire-scale functional evaluations of natively paired TCRs. In particular, we leveraged the immortalized nature of physically linked TCR:{beta} amplicon libraries to analyze binding against multiple recombinant pMHCs on a repertoire scale. To exemplify the utility of this approach, we also performed affinity-based functional mapping in conjunction with quantitative next-generation sequencing to track antigen-specific TCRs. These data successfully validated a new immortalization and screening platform to facilitate detailed molecular analyses of human TCRs against diverse antigen targets associated with health, vaccination, or disease.

immunology↗

bioRxiv2021

Quantifying Central Metabolic Fluxes in Human Platelets Using Metabolic Flux Analysis

Platelet metabolism is linked to platelet hyper- and hypoactivity in numerous human diseases. Developing a detailed understanding of the link between metabolic shifts and platelet activation state is integral to improving human health. Here, we show the first application of isotopically nonstationary 13C metabolic flux analysis to quantitatively measure carbon fluxes in both resting and thrombin activated platelets. Resting platelets primarily metabolize glucose to lactate via glycolysis, while acetate is oxidized to fuel the tricarboxylic acid cycle. Upon activation with thrombin, a potent platelet agonist, platelets increase their uptake of glucose 3-fold. This results in an absolute increase in flux throughout central metabolism, but when compared to resting platelets they redistribute carbon dramatically. Activated platelets decrease relative flux to the oxidative pentose phosphate pathway and TCA cycle from glucose and increase relative flux to lactate. These results provide the first report of reaction-level carbon fluxes in platelets and allow us to distinguish metabolic fluxes with much higher resolution than previous studies.

systems biology↗