Biology subjects

Johnson, R.

Publications and source records attributed to Johnson, R..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Rhizaria in the oligotrophic ocean exhibit clear temporal and vertical variability

Recently studies have shown that Rhizaria, a super-group of marine protists, have a large role in pelagic ecosystems. They are unique in that they construct mineral tests out of silica, calcium carbonate, or strontium sulfate. As a consequence, Rhizaria can have large impacts on the oceans cycling of carbon and other elements. However, less is known about Rhizaria ecology or their role in the pelagic food-web. Some taxa, like certain Radiolaria, are mixotrophic, hosting algal symbionts. While other taxa are flux-feeders or even predatory carnivores. Some prior research has suggested that Rhizaria will partition vertically in the water column, likely due to different trophic strategies. However, very few studies have investigated their populations over extended periods of time. In this study, we present data investigating Rhizaria abundance and vertical distribution from over a year of monthly cruises in the Sargasso Sea. This study represents the first quantification of Rhizaria throughout the mesopelagic zone in an oligotrophic system for an extended period of time. We use this data to investigate the hypothesis that Rhizaria taxonomic groups will partition due to trophic mode. We also investigate how their abundance varies in accordance with environmental parameters. Rhizaria abundance was quantified using an Underwater Vision Profiler (UVP5), an in-situ imaging device. Ultimately, we show that different Rhizaria taxa will have unique vertical distribution patterns. Models relating their abundance to environmental parameters have mixed results, yet particle concentration is a common predictive variable, supporting the importance of heterotrophy amongst many taxa.

ecology↗

bioRxiv2024

The global fish and invertebrate abundance value of mangroves

Mangroves are a critical coastal habitat that provides a suite of ecosystem services and supports livelihoods. We undertake the first global analysis to estimate density and abundance of 37 commercially important fish and invertebrates that are known to extensively use mangroves. Geomorphic mangrove type, sea surface salinity and temperature, and length of mangrove forest edge were important in predicting the density of commercial fish and invertebrates, with deltaic systems supporting the highest densities. The model predicted high densities throughout parts of southeast Asia, the northern coast of South America, the Red Sea, and the Caribbean and Central America. Application of our model onto the global mangrove extent, estimates that mangroves support the annual abundance of nearly 800 billion young-of-year fish and invertebrates contained in our model. Our results confirm the critical role of mangroves globally in supporting fish and fisheries, and further builds the case for their conservation and restoration.

ecology↗

bioRxiv2024

Brain-wide arousal signals are segregated from movement planning in the superior colliculus

The superior colliculus (SC) is traditionally considered a brain region that functions as an interface between processing visual inputs and generating eye movement outputs. Although its role as a primary reflex center is thought to be conserved across vertebrate species, evidence suggests that the SC has evolved to support higher-order cognitive functions including spatial attention. When it comes to oculomotor areas such as the SC, it is critical that high precision fixation and eye movements are maintained even in the presence of signals related to ongoing changes in cognition and brain state, both of which have the potential to interfere with eye position encoding and movement generation. In this study, we recorded spiking responses of neuronal populations in the SC while monkeys performed a memory-guided saccade task and found that the activity of some of the neurons fluctuated over tens of minutes. By leveraging the statistical power afforded by high-dimensional neuronal recordings, we were able to identify a low-dimensional pattern of activity that was correlated with pupil size and simultaneously recorded data in the prefrontal cortex (PFC), consistent with slow changes in the monkeys arousal levels while they were performing the task. Importantly, we found that the spiking responses of deep-layer SC neurons were less correlated with this brain-wide arousal signal, and that neural activity associated with changes in pupil size and saccade tuning did not overlap in population activity space with movement initiation signals. Taken together, these findings provide a framework for understanding how signals related to cognition and arousal can be embedded in the population activity of oculomotor structures without compromising the fidelity of the motor output.

neuroscience↗

bioRxiv2024

Activity of Salmonella SPI-1 inhibits the TLR4-dependent transcriptional but not translational response during macrophage infection

Changes in gene expression during bacterial infection are the combined result of altered transcription and translation, with the latter comparatively understudied. Gram-negative bacteria rapidly trigger cytokine gene transcription in macrophages through the activation of pathogen associated molecular pattern receptors, for example detection of Salmonella lipopolysaccharide (LPS) from the bacterial cell envelope by Toll-like receptor 4 (TLR4). Here, through time-resolved parallel translatomic and transcriptomic profiling, we now show temporal TLR4-specific translational upregulation of cell signalling proteins in macrophages induced by Salmonella. While transcriptional upregulation of these genes is dampened through the activity of the Salmonella SPI-1 type three secretion system, a robust translational response remains. These data reveal an important host-pathogen translational regulatory network that modifies the innate immune response of macrophages to infection.

molecular biology↗

bioRxiv2023

Syncytia Formation Promotes Virus Resistance to Interferon and Neutralizing Antibodies

SARS-CoV-2, like many viruses, generates syncytia but the role of syncytia formation in viral evolution remains unknown. Using SARS-CoV-2 and SARS-CoV-2 Spike (S) replacement vesicular stomatitis (VSV), we show that S-mediated syncytia impair the antiviral effects of interferons in cultured cells, human lung cell cultures, and hACE2 transgenic mice. Amino acid substitutions that modulate syncytia formation in Delta- and Omicron-encoded S have parallel effects on viral interferon resistance. S-mediated syncytia compromise antibody-mediated virus neutralization in cultured cells. We recapitulate interferon and neutralizing antibody resistance in syncytia generated by the orthoreovirus p14 fusion-associated small transmembrane (FAST) protein in VSV, influenza virus, and seasonal coronavirus OC43 infections. These findings explain selection of SARS-CoV-2 fusogenic variants in humans and, more generally, the evolution of fusogenic viruses driven by adaptive and innate immunity.

microbiology↗

bioRxiv2023

Together, Neuropilin 1 and Neuropilin 2 direct α5 integrin trafficking through GTPase-activating-protein p120RasGAP in endothelial cells to promote fibronectin fibrillogenesis

Integrin trafficking to and from membrane adhesions is a crucial mechanism that dictates many aspects of a cells behaviour, including motility, polarisation, and invasion. In endothelial cells (ECs), the intracellular traffic of 5 integrin is regulated by both neuropilin 1 (NRP1) and neuropilin 2 (NRP2), yet the redundancies in function between these co-receptors remain unclear. Moreover, the endocytic complexes that participate in NRP-directed traffic remain poorly annotated. Using label-free quantitative mass spectrometry of 5 integrin associations in ECs we identify 5 trafficking pathways that depend on NRP1, NRP2, or both NRPs. We identify a trafficking pathway that depends on both NRPs: one that impinges on the GTPase-activating protein p120RasGAP. This pathway promotes the recycling of 5 integrin from early endosomes. Mechanistically, p120RasGAP enables transit of endocytosed 5 integrin-NRP1-NRP2 complexes to Rab11+ recycling endosomes, promoting cell polarisation and fibronectin (FN) fibrillogenesis. Silencing of both NRP receptors, or p120RasGAP, results in the accumulation of 5 integrin in early endosomes, a loss of 5 integrin from surface adhesions, and attenuated EC polarisation. Importantly, endothelial-specific deletion of both NRP1 and NRP2 in the postnatal retina recapitulated our in vitro findings, severely impairing FN fibrillogenesis and polarised sprouting. Our data assign an essential role for p120RasGAP during integrin traffic in ECs and support a hypothesis that NRP receptors can co-traffic internalised cargoes.

cell biology↗

bioRxiv2023

Microeukaryote metabolism across the western North Atlantic Ocean revealed through autonomous underwater profiling

Protists (microeukaryotes) are key contributors to marine carbon cycling, influencing the transfer of energy to higher trophic levels and the vertical movement of carbon to the ocean interior. Their physiology, ecology, and interactions with the chemical environment are still poorly understood in offshore ecosystems, and especially in the deep ocean. Using the Autonomous Underwater Vehicle (AUV) Clio, the microbial community along a 1,050 km transect in the western North Atlantic Ocean was surveyed at 10-200 m vertical depth increments to capture metabolic microeukaryote signatures spanning a gradient of oligotrophic, continental margin, and productive coastal ecosystems. Plankton biomass was collected along the surface of this transect and across depth features, and taxonomy and metabolic function were examined using a paired metatranscriptomic and metaproteomic approach. A shift in the microeukaryote community composition was observed from the euphotic zone through the mesopelagic and into the bathypelagic ocean. A diverse surface assemblage consisting of haptophytes, stramenopiles, dinoflagellates and ciliates was represented in both the transcript and protein fractions, with foraminifera, radiolaria, picozoa, and discoba proteins enriched at >200 m depth, and fungal proteins emerging in waters >3,000 m depth. In the broad microeukaryote community, nitrogen stress biomarkers were found in productive coastal sites, with phosphorus stress biomarkers in offshore waters where Saharan dust input is thought to supply iron and nitrogen. This multi-omics dataset broadens our understanding of how microeukaryotic taxa and their functional processes are structured along environmental gradients of temperature, light, macronutrients, and trace metals.

ecology↗

bioRxiv2023

Connecting Thiamine Availability to the Microbial Community Composition in Chinook Salmon Spawning Habitats of the Sacramento River Basin

Thiamine Deficiency Complex (TDC) is a major emerging threat to global populations of culturally and economically important populations of salmonids. Salmonid eggs and embryos can assimilate exogenous thiamine, and evidence suggests that microbial communities in benthic environments can produce substantial amounts of thiamine. We therefore hypothesize that microbially produced thiamine in both riverine surface water and hyporheic zones could serve to rescue early life stages of salmonids suffering from TDC. The distributions of thiamine and its metabolically related compounds (dTRCs) have never been determined in freshwater systems. Similarly, the microbial cycling of these compounds has never been investigated. Here we determine that all dTRCs are present in femto-picomolar concentrations across diverse salmon spawning habitats in Californias Sacramento River system. We observed that thiamine concentrations in the Sacramento River are orders of magnitude lower than marine environments, indicating substantial differences in thiamine cycling between these two environments. Our data suggest that the hyporheic zone is likely the source of thiamine to the overlying surface water. Temporal variations in dTRC concentration were observed where highest concentrations were seen when Chinook salmon were actively spawning. Significant correlations were identified between the richness of differentially abundant ASVs and dTRC concentrations. The influence of these ASVs on dTRC concentrations provide evidence of dTRC cycling by microbes in the hyporheic zone, which would influence the conditions where embryonic salmon incubate. Together, these results indicate a connection between microbial communities in freshwater habitats and the availability of thiamine to spawning TDC-impacted California Central Valley Chinook salmon.

microbiology↗

bioRxiv2023

Salmonella impairs macrophage immunity through effector-independent rapid translational induction in response to membrane puncture by the SPI-1 injectisome

During bacterial infection both the host cell and its invader must divert intracellular resources to synthesise specific proteins in a timely manner. For the host, these factors may be needed for innate immune responses, including programmed cell death, and in the bacteria newly synthesized proteins may be survival factors needed to counteract host responses. Salmonella is an important food-borne bacterial pathogen that invades and multiplies within host cells. It is well established that invasion of epithelial cells is dependent upon the SPI-1 Type III injectisome, a biological needle that penetrates and secretes effectors into host cells to promote internalization. However, the importance of the SPI-1 injectisome in infection of professional phagocytes such as macrophages, which are the predominant host cell type during systemic infection, is less clear. Through time resolved parallel transcriptomic and translatomic studies of macrophage infection, we revealed that SPI-1 injectisome-dependent infection of macrophages triggers rapid translation of transcription factor mRNAs, including Early Growth Response 1 (Egr1). Despite the short half-life of EGR1 protein, its swift synthesis within the initial hour of infection is sufficient to inhibit transcription of pro-inflammatory genes and thereby restrain inflammatory responses and programmed cell death within the first hour of during early infection. This transient period of inflammatory suppression in macrophages is exploited by Salmonella to establish infection and sheds new insight on the importance of translational activation in host-pathogen dynamics during Salmonella infection.

molecular biology↗

bioRxiv2023

Atypical Chemokine Receptor 1 (Ackr1)-deficient Mice Resist Lethal SARS-CoV-2 Challenge

High pro-inflammatory chemokine levels have been reported in blood and lung in patients with COVID-19. To investigate specific roles in pathogenesis, we studied the regulation of chemokine ligands and receptors in the lungs of 4-6-month-old wild type C57BL/6 mice infected with the MA10 mouse-adapted strain of SARS-CoV-2. We found that atypical chemokine receptor 1 (Ackr1, also known as Duffy antigen receptor for chemokines/DARC) was the most highly upregulated chemokine receptor in infected lung, where it localized to endothelial cells of veins and arterioles. In a screen of 7 leukocyte chemoattractant or chemoattractant receptor knockout mouse lines, Ackr1-/- mice were unique in having lower mortality after SARS-CoV-2 infection, particularly in males. ACKR1 is a non-signaling chemokine receptor that in addition to endothelium is also expressed on erythrocytes and Purkinje cells of the cerebellum. It binds promiscuously to both inflammatory CC and CXC chemokines and has been reported to control chemokine availability which may influence the shape of chemotactic gradients and the ability of leukocytes to extravasate and produce immunopathology. Of note, erythrocyte ACKR1 deficiency is fixed in sub-Saharan African populations where COVID-19 has been reported to result in low mortality compared to worldwide data. Our data suggest the possibility of a causal contribution of ACKR1 deficiency to low sub-Saharan COVID-19 mortality and identify ACKR1 as a possible drug target in the disease.

immunology↗

bioRxiv2023

MBNL1 regulates programmed postnatal switching between regenerative and differentiated cardiac states.

Discovering determinants of cardiomyocyte maturity and the maintenance of differentiated states is critical to both understanding development and potentially reawakening endogenous regenerative programs in adult mammalian hearts as a therapeutic strategy. Here, the RNA binding protein Muscleblind-like 1 (MBNL1) was identified as a critical regulator of cardiomyocyte differentiated states and their regenerative potential through transcriptome-wide control of RNA stability. Targeted MBNL1 overexpression early in development prematurely transitioned cardiomyocytes to hypertrophic growth, hypoplasia, and dysfunction, whereas loss of MBNL1 function increased cardiomyocyte cell cycle entry and proliferation through altered cell cycle inhibitor transcript stability. Moreover, MBNL1-dependent stabilization of the estrogen-related receptor signaling axis was essential for maintaining cardiomyocyte maturity. In accordance with these data, modulating MBNL1 dose tuned the temporal window of cardiac regeneration, where enhanced MBNL1 activity arrested myocyte proliferation, and MBNL1 deletion promoted regenerative states with prolonged myocyte proliferation. Collectively these data suggest MBNL1 acts as a transcriptome-wide switch between regenerative and mature myocyte states postnatally and throughout adulthood.

cell biology↗

bioRxiv2023

Neural and behavioral correlates of edible cannabis-induced poisoning: characterizing a novel preclinical model

Accidental exposure to {Delta}9-tetrahydrocannabinol (THC)-containing edible cannabis, leading to cannabis poisoning, is common in children and pets; however, the neural mechanisms underlying these poisonings remain unknown. Therefore, we examined the effects of acute edible cannabis-induced poisoning on neural activity and behavior. Adult Sprague-Dawley rats (6 males, 7 females) were implanted with electrodes in the prefrontal cortex (PFC), dorsal hippocampus (dHipp), cingulate cortex (Cg), and nucleus accumbens (NAc). Cannabis poisoning was then induced by exposure to a mixture of Nutella (6 g/kg) and THC-containing cannabis oil (20 mg/kg). Subsequently, cannabis tetrad and neural oscillations were examined 2, 4, 8, and 24 h after THC exposure. In another cohort (16 males, 15 females), we examined the effects of cannabis poisoning on learning and prepulse inhibition, and the serum and brain THC and 11-hydroxy-THC concentrations. Cannabis poisoning resulted in sex differences in brain and serum THC and 11-hydroxy-THC levels over a 24-h period. It also caused gamma power suppression in the Cg, dHipp, and NAc in a sex- and time-dependent manner. Cannabis poisoning also resulted in hypolocomotion, hypothermia, and anti-nociception in a time-dependent manner and impairments in learning and prepulse inhibition. Our results suggest that the impairments in learning and information processing may be due to the decreased gamma power in the dHipp and PFC. Additionally, most of the changes in neural activity and behavior appear 2 hours after ingestion, suggesting that interventions at or before this time might be effective in reversing or reducing the effects of cannabis poisoning.

neuroscience↗

bioRxiv2022

Live-attenuated pediatric parainfluenza vaccine expressing 6P-stabilized SARS-CoV-2 spike protein is protective against SARS-CoV-2 variants in hamsters

The pediatric live-attenuated bovine/human parainfluenza virus type 3 (B/HPIV3)-vectored vaccine expressing the prefusion-stabilized SARS-CoV-2 spike (S) protein (B/HPIV3/S-2P) was previously evaluated in vitro and in hamsters. To improve its immunogenicity, we generated B/HPIV3/S-6P, expressing S further stabilized with 6 proline mutations (S-6P). Intranasal immunization of hamsters with B/HPIV3/S-6P reproducibly elicited significantly higher serum anti-S IgA/IgG titers than B/HPIV3/S-2P; hamster sera efficiently neutralized variants of concern (VoCs), including Omicron variants. B/HPIV3/S-2P and B/HPIV3/S-6P immunization protected hamsters against weight loss and lung inflammation following SARS-CoV-2 challenge with the vaccine-matched strain WA1/2020 or VoCs B.1.1.7/Alpha or B.1.351/Beta and induced near-sterilizing immunity. Three weeks post-challenge, B/HPIV3/S-2P- and B/HPIV3/S-6P-immunized hamsters exhibited a robust anamnestic serum antibody response with increased neutralizing potency to VoCs, including Omicron sublineages. B/HPIV3/S-6P primed for stronger anamnestic antibody responses after challenge with WA1/2020 than B/HPIV3/S-2P. B/HPIV3/S-6P will be evaluated as an intranasal vaccine to protect infants against both HPIV3 and SARS-CoV-2. AUTHOR SUMMARYSARS-CoV-2 infects and causes disease in all age groups. While injectable SARS-CoV-2 vaccines are effective against severe COVID-19, they do not fully prevent SARS-CoV-2 replication and transmission. This study describes the preclinical comparison in hamsters of B/HPIV3/S-2P and B/HPIV3/S-6P, live-attenuated pediatric vector vaccine candidates expressing the "2P" prefusion stabilized version of the SARS-CoV-2 spike protein, or the further-stabilized "6P" version. B/HPIV3/S-6P induced significantly stronger anti-S serum IgA and IgG responses than B/HPIV3/S-2P. A single intranasal immunization with B/HPIV3/S-6P elicited broad systemic antibody responses in hamsters that efficiently neutralized the vaccine-matched isolate as well as variants of concern, including Omicron. B/HPIV3/S-6P immunization induced near-complete airway protection against the vaccine-matched SARS-CoV-2 isolate as well as two variants. Furthermore, following SARS-CoV-2 challenge, immunized hamsters exhibited strong anamnestic serum antibody responses. Based on these data, B/HPIV3/S-6P will be further evaluated in a phase I study.

microbiology↗

bioRxiv2022

Zinc and laboratory emerge date impact honey bee gut microbiota and survival

Honey bees (Apis mellifera) may be exposed to a wide variety of chemicals in the environment, including pesticides, antibiotics, and metals. Zinc, for example, is commonly included in fertilizers, pesticides, and feed additives, and is found in agricultural runoff. Honey bees can be exposed to zinc directly or indirectly by consuming zinc-contaminated nectar and pollen. However, there is a paucity of studies addressing the putative effects of zinc on honey bees health. In this study, we tested the effects of zinc on honey bee survivorship and gut microbiota. To evaluate survivorship, we exposed bees to six concentrations of zinc (0, 50, 100, 250, 500, or 1000 mg/L) and assessed survival daily for 10 days. To evaluate effects of zinc on gut microbiota, we exposed bees to 5 or 100 mg/L zinc. Bees were sampled before (day 0) and after zinc exposure (days 3, 6, and 9). Abdominal contents underwent DNA extraction and 16S rRNA sequencing (V3-V4) on an Illumina MiSeq. Sequences were filtered and processed through QIIME2 and DADA2. Zinc treatment had minimal effects on bacterial DNA concentrations and absolute cell counts while emerge date (the date a bee emerged from the brood comb) had a significant effect with decreased bacterial concentrations and cell counts observed at later emerge dates. Survival was only minimally impacted (>89% survival) at zinc concentrations up to 100 mg/L. Zinc had limited effects on overall gut microbial composition, diversity, and taxonomic abundances, with the greatest differences noted in the bee group exposed to the higher concentration of zinc (100 mg/L). In this group, several beneficial taxa (Lactobacillus, Rhizobiaceae, Gilliamella) were found at reduced abundances, while Paenibacillus, a potentially pathogenic taxa, was found at increased abundances. This suggests that zinc exposure, even at relatively low levels, may negatively impact honey bee health, even if survivorship is not dramatically impacted. Notably, emerge date effects were also observed in microbial composition. These results demonstrate the need to include assessments of honey bee gut microbiota in addition to other metrics of honey bee health and survivorship when evaluating the potential effects of agrochemicals on honey bees

ecology↗

bioRxiv2022

Hepatocyte Growth Factor and β1-integrin signalling axis drives tunneling nanotube formation in A549 lung adenocarcinoma cells

Tunneling nanotubes (TNTs) are thin cytoplasmic protrusions involved in long-distance cellular communication. The presence of TNTs has been found in vivo and in vitro studies in non-small cell lung cancer (NSCLC). Cancer cells transport a range of organelles and signalling molecules along TNTs, to confer a survival phenotype for the recipient cell, contributing toward chemoresistance and malignancy. Despite its important role in cancer progression, the molecular mechanisms underlying TNT formation is not well defined. Within the tumour microenvironment (TME) of NSCLC, hepatocyte growth factor (HGF) and its receptor, c-Met, are mutationally upregulated causing growth, and invasion. In this study, we report a novel crosstalk between HGF/c-Met and {beta}1-integrin involved in the formation of functional TNTs in A549 cells. Through pharmacological inhibitor studies, we discovered Arp2/3 complex, MAPK and PI3K pathways were activated downstream of this crosstalk signalling axis. Furthermore, paxillin was recruited during this key process, localising at the protrusion site of HGF-induced TNTs, and therefore serving as the central link between the upstream and downstream regulators involved. Overall, these results demonstrate a novel strategy to inhibit TNT formation in NSCLC through targeting the HGF/c-Met and {beta}1-integrin signalling axis, thus highlighting the importance of personalised multi-drug targeting in NSCLC.

cancer biology↗

bioRxiv2022

Oxygen levels at the time of activation determine T cell persistence and immunotherapeutic efficacy

SummaryOxygenation levels are a determinative factor in T cell function. Here we describe that the oxygen tensions sensed by T cells at the moment of activation act to persistently modulate both differentiation and function. We found that in a protocol of CAR-T cell generation, 24 hours of low oxygen levels during initial CD8+ T cell priming is sufficient to enhance antitumour cytotoxicity in a preclinical model. This is the case even when CAR-T cells are subsequently cultured under high oxygen tensions prior to adoptive transfer. Increased hypoxia inducible transcription factor (HIF) expression was able to alter T cell fate in a similar manner to exposure to low oxygen tensions; however, only a controlled or temporary increase in HIF signalling was able to consistently improve cytotoxic function of T cells. These data show that oxygenation levels during and immediately after T cell activation play an essential role in regulating T cell function.

immunology↗

bioRxiv2022

Diet Induced Obesity and Diabetes Enhance Mortality and Reduces Vaccine Efficacy for SARS-CoV-2

Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), the causative agent of Coronavirus disease 2019 (COVID-19), emerged in Wuhan, China, in December 2019. As of October 2022, there have been over 625 million confirmed cases of COVID-19, including over 6.5 million deaths. Epidemiological studies have indicated that comorbidities of obesity and diabetes mellitus are associated with increased morbidity and mortality following SARS-CoV-2 infection. We determined how the comorbidities of obesity and diabetes affect morbidity and mortality following SARS-CoV-2 infection in unvaccinated and adjuvanted spike nanoparticle (NVX-CoV2373) vaccinated mice. We find that obese/diabetic mice infected with SARS-CoV-2 have increased morbidity and mortality compared to age matched normal mice. Mice fed a high-fat diet (HFD) then vaccinated with NVX-CoV2373 produce equivalent neutralizing antibody titers to those fed a normal diet (ND). However, the HFD mice have reduced viral clearance early in infection. Analysis of the inflammatory immune response in HFD mice demonstrates a recruitment of neutrophils that was correlated with increased mortality and reduced clearance of the virus. Depletion of neutrophils in diabetic/obese vaccinated mice reduced disease severity and protected mice from lethality. This model recapitulates the increased disease severity associated with obesity and diabetes in humans with COVID-19 and is an important comorbidity to study with increasing obesity and diabetes across the world. ImportanceSARS-CoV-2 has caused a wide spectrum of disease in the human population, from asymptomatic infections to death. It is important to study the host differences that may alter the pathogenesis of this virus. One clinical finding in COVID19 patients, is that people with obesity or diabetes are at increased risk of severe illness from SARS-CoV-2 infection. We used a high fat diet model in mice to study the effects of obesity and Type 2 diabetes on SARS-CoV-2 infection as well as how these comorbidities alter the response to vaccination. We find that diabetic/obese mice have increased disease after SARS-CoV-2 infection and they have slower clearance of virus. We find that the lungs of these mice have increased neutrophils and that removing these neutrophils protect diabetic/obese mice from disease. This demonstrates why these diseases have increased risk of severe disease and suggests specific interventions upon infection.

microbiology↗

bioRxiv2022

N-linked glycosylation increases horse radish peroxidase rigidity leading to enhanced activity and stability.

Glycosylation is the most prevalent protein post-translational modification, with a quarter of glycosylated proteins having enzymatic properties. Yet the full impact of glycosylation on the protein structure-function relationship, especially in enzymes, is still limited. Here we show glycosylation rigidifies the important commercial enzyme horseradish peroxidase (HRP), which in turn increases its activity and stability. Circular dichroism spectroscopy revealed that glycosylation increased holo-HRPs thermal stability and promoted significant helical structure in the absence of haem (apo-HRP). Glycosylation also resulted in a 10-fold increase in enzymatic turnover towards o-phenylenediamine dihydrochloride when compared to its non-glycosylated form. Utilising a naturally occurring site-specific probe of active site flexibility (Trp117) in combination with red-edge excitation shift fluorescence spectroscopy, we found that glycosylation significantly rigidified the enzyme. In silico simulations confirmed that glycosylation largely decreased protein backbone flexibility, especially in regions close to the active site and the substrate access channel. Thus, our data shows that glycosylation does not just have a passive effect on HRP stability but can exert long range effects that mediate the native enzymes activity and stability through changes in inherent dynamics.

biochemistry↗