Interconnections of Reactive Oxygen Species Homeostasis and Circadian Rhythm in Neurospora crassa.

Abstract Significance: Both circadian rhythm and the production of reactive oxygen species (ROS) are fundamental features of aerobic eukaryotic cells. The circadian clock enhances the fitness of organisms by enabling them to anticipate cycling changes in the surroundings. ROS generation in the cell is often altered in response to environmental changes, but oscillations in ROS levels may also reflect endogenous metabolic fluctuations governed by the circadian clock. On the other hand, an effective regulation and timing of antioxidant mechanisms may be crucial in the defense of cellular integrity. Thus, an interaction between the circadian timekeeping machinery and ROS homeostasis or signaling in both directions may be of advantage at all phylogenetic levels. Recent Advances: The Frequency-White Collar-1 and White Collar-2 oscillator (FWO) of the filamentous fungus Neurospora crassa is well characterized at the molecular level. Several members of the ROS homeostasis were found to be controlled by the circadian clock, and ROS levels display circadian rhythm in Neurospora. On the other hand, multiple data indicate that ROS affect the molecular oscillator. Critical Issues: Increasing evidence suggests the interplay between ROS homeostasis and oscillators that may be partially or fully independent of the FWO. In addition, ROS may be part of a complex cellular network synchronizing non-transcriptional oscillators with timekeeping machineries based on the classical transcription-translation feedback mechanism. Future Directions: Further investigations are needed to clarify how the different layers of the bidirectional interactions between ROS homeostasis and circadian regulation are interconnected. Antioxid. Redox Signal. 00, 000-000

Redox proteomics of the inflammatory secretome identifies a common set of redoxins and other glutathionylated proteins released in inflammation, influenza virus infection and oxidative stress

Protein cysteines can form transient disulfides with glutathione (GSH), resulting in the production of glutathionylated proteins, and this process is regarded as a mechanism by which the redox state of the cell can regulate protein function. Most studies on redox regulation of immunity have focused on intracellular proteins. In this study we have used redox proteomics to identify those proteins released in glutathionylated form by macrophages stimulated with lipopolysaccharide (LPS) after pre-loading the cells with biotinylated GSH. Of the several proteins identified in the redox secretome, we have selected a number for validation. Proteomic analysis indicated that LPS stimulated the release of peroxiredoxin (PRDX) 1, PRDX2, vimentin (VIM), profilin1 (PFN1) and thioredoxin 1 (TXN1). For PRDX1 and TXN1, we were able to confirm that the released protein is glutathionylated. PRDX1, PRDX2 and TXN1 were also released by the human pulmonary epithelial cell line, A549, infected with influenza virus. The release of the proteins identified was inhibited by the anti-inflammatory glucocorticoid, dexamethasone (DEX), which also inhibited tumor necrosis factor (TNF)-α release, and by thiol antioxidants (N-butanoyl GSH derivative, GSH-C4, and N-acetylcysteine (NAC), which did not affect TNF-α production. The proteins identified could be useful as biomarkers of oxidative stress associated with inflammation, and further studies will be required to investigate if the extracellular forms of these proteins has immunoregulatory functions

Ergothioneine Biosynthesis and Functionality in the Opportunistic Fungal Pathogen, Aspergillus fumigatus.

Ergothioneine (EGT; 2-mercaptohistidine trimethylbetaine) is a trimethylated and sulphurised histidine derivative which exhibits antioxidant properties. Here we report that deletion of Aspergillus fumigatus egtA (AFUA_2G15650), which encodes a trimodular enzyme, abrogated EGT biosynthesis in this opportunistic pathogen. EGT biosynthetic deficiency in A. fumigatus significantly reduced resistance to elevated H2O2 and menadione, respectively, impaired gliotoxin production and resulted in attenuated conidiation. Quantitative proteomic analysis revealed substantial proteomic remodelling in ΔegtA compared to wild-type under both basal and ROS conditions, whereby the abundance of 290 proteins was altered. Specifically, the reciprocal differential abundance of cystathionine γ-synthase and β-lyase, respectively, influenced cystathionine availability to effect EGT biosynthesis. A combined deficiency in EGT biosynthesis and the oxidative stress response regulator Yap1, which led to extreme oxidative stress susceptibility, decreased resistance to heavy metals and production of the extracellular siderophore triacetylfusarinine C and increased accumulation of the intracellular siderophore ferricrocin. EGT dissipated H2O2 in vitro, and elevated intracellular GSH levels accompanied abrogation of EGT biosynthesis. EGT deficiency only decreased resistance to high H2O2 levels which suggests functionality as an auxiliary antioxidant, required for growth at elevated oxidative stress conditions. Combined, these data reveal new interactions between cellular redox homeostasis, secondary metabolism and metal ion homeostasis

Multi-messenger observations of a binary neutron star merger

On 2017 August 17 a binary neutron star coalescence candidate (later designated GW170817) with merger time 12:41:04 UTC was observed through gravitational waves by the Advanced LIGO and Advanced Virgo detectors. The Fermi Gamma-ray Burst Monitor independently detected a gamma-ray burst (GRB 170817A) with a time delay of ~1.7 s with respect to the merger time. From the gravitational-wave signal, the source was initially localized to a sky region of 31 deg2 at a luminosity distance of 40+8-8 Mpc and with component masses consistent with neutron stars. The component masses were later measured to be in the range 0.86 to 2.26 Mo. An extensive observing campaign was launched across the electromagnetic spectrum leading to the discovery of a bright optical transient (SSS17a, now with the IAU identification of AT 2017gfo) in NGC 4993 (at ~40 Mpc) less than 11 hours after the merger by the One- Meter, Two Hemisphere (1M2H) team using the 1 m Swope Telescope. The optical transient was independently detected by multiple teams within an hour. Subsequent observations targeted the object and its environment. Early ultraviolet observations revealed a blue transient that faded within 48 hours. Optical and infrared observations showed a redward evolution over ~10 days. Following early non-detections, X-ray and radio emission were discovered at the transient’s position ~9 and ~16 days, respectively, after the merger. Both the X-ray and radio emission likely arise from a physical process that is distinct from the one that generates the UV/optical/near-infrared emission. No ultra-high-energy gamma-rays and no neutrino candidates consistent with the source were found in follow-up searches. These observations support the hypothesis that GW170817 was produced by the merger of two neutron stars in NGC4993 followed by a short gamma-ray burst (GRB 170817A) and a kilonova/macronova powered by the radioactive decay of r-process nuclei synthesized in the ejecta

Production of Native Bispecific Antibodies in Rabbits

BACKGROUND: A natural bispecific antibody, which can be produced by exchanging Fab arms of two IgG4 molecules, was first described in allergic patients receiving therapeutic injections with two distinct allergens. However, no information has been published on the production of natural bispecific antibody in animals. Even more important, establishment of an animal model is a useful approach to investigate and characterize the naturally occurring antibody. METHODOLOGY/PRINCIPAL FINDINGS: We demonstrated that a natural bispecific antibody can also be generated in New Zealand white rabbits by immunization with synthesized conjugates. These antibodies showed bispecificity to the components that were simultaneously used to immunize the animals. We observed a trend in our test animals that female rabbits exhibited stronger bispecific antibody responses than males. The bispecific antibody was monomeric and primarily belonged to immunoglobulin (Ig) G. Moreover, bispecific antibodies were demonstrated by mixing 2 purified monospecific antibodies in vivo and in vitro. CONCLUSIONS/SIGNIFICANCE: Our results extend the context of natural bispecific antibodies on the basis of bispecific IgG4, and may provide insights into the exploration of native bispecific antibodies in immunological diseases

Subcellular distribution of glutathione and cysteine in cyanobacteria

Glutathione plays numerous important functions in eukaryotic and prokaryotic cells. Whereas it can be found in virtually all eukaryotic cells, its production in prokaryotes is restricted to cyanobacteria and proteobacteria and a few strains of gram-positive bacteria. In bacteria, it is involved in the protection against reactive oxygen species (ROS), osmotic shock, acidic conditions, toxic chemicals, and heavy metals. Glutathione synthesis in bacteria takes place in two steps out of cysteine, glutamate, and glycine. Cysteine is the limiting factor for glutathione biosynthesis which can be especially crucial for cyanobacteria, which rely on both the sufficient sulfur supply from the growth media and on the protection of glutathione against ROS that are produced during photosynthesis. In this study, we report a method that allows detection and visualization of the subcellular distribution of glutathione in Synechocystis sp. This method is based on immunogold cytochemistry with glutathione and cysteine antisera and computer-supported transmission electron microscopy. Labeling of glutathione and cysteine was restricted to the cytosol and interthylakoidal spaces. Glutathione and cysteine could not be detected in carboxysomes, cyanophycin granules, cell walls, intrathylakoidal spaces, periplasm, and vacuoles. The accuracy of the glutathione and cysteine labeling is supported by two observations. First, preadsorption of the antiglutathione and anticysteine antisera with glutathione and cysteine, respectively, reduced the density of the gold particles to background levels. Second, labeling of glutathione and cysteine was strongly decreased by 98.5% and 100%, respectively, in Synechocystis sp. cells grown on media without sulfur. This study indicates a strong similarity of the subcellular distribution of glutathione and cysteine in cyanobacteria and plastids of plants and provides a deeper insight into glutathione metabolism in bacteria

Adult-Onset Obesity Reveals Prenatal Programming of Glucose-Insulin Sensitivity in Male Sheep Nutrient Restricted during Late Gestation

BACKGROUND: Obesity invokes a range of metabolic disturbances, but the transition from a poor to excessive nutritional environment may exacerbate adult metabolic dysfunction. The current study investigated global maternal nutrient restriction during early or late gestation on glucose tolerance and insulin sensitivity in the adult offspring when lean and obese. METHODS/PRINCIPAL FINDINGS: Pregnant sheep received adequate (1.0M; CE, n = 6) or energy restricted (0.7M) diet during early (1-65 days; LEE, n = 6) or late (65-128 days; LEL, n = 7) gestation (term approximately 147 days). Subsequent offspring remained on pasture until 1.5 years when all received glucose and insulin tolerance tests (GTT & ITT) and body composition determination by dual energy x-ray absorptiometry (DXA). All animals were then exposed to an obesogenic environment for 6-7 months and all protocols repeated. Prenatal dietary treatment had no effect on birth weight or on metabolic endpoints when animals were 'lean' (1.5 years). Obesity revealed generalised metabolic 'inflexibility' and insulin resistance; characterised by blunted excursions of plasma NEFA and increased insulin(AUC) (from 133 to 341 [s.e.d. 26] ng.ml(-1).120 mins) during a GTT, respectively. For LEL vs. CE, the peak in plasma insulin when obese was greater (7.8 vs. 4.7 [s.e.d. 1.1] ng.ml(-1)) and was exacerbated by offspring sex (i.e. 9.8 vs. 4.4 [s.e.d. 1.16] ng.ml(-1); LEL male vs. CE male, respectively). Acquisition of obesity also significantly influenced the plasma lipid and protein profile to suggest, overall, greater net lipogenesis and reduced protein metabolism. CONCLUSIONS: This study indicates generalised metabolic dysfunction with adult-onset obesity which also exacerbates and 'reveals' programming of glucose-insulin sensitivity in male offspring prenatally exposed to maternal undernutrition during late gestation. Taken together, the data suggest that metabolic function appears little compromised in young prenatally 'programmed' animals so long as weight is adequately controlled. Nutritional excess in adulthood exacerbates any programmed phenotype, indicating greater vigilance over weight control is required for those individuals exposed to nutritional thrift during gestation

Population-Related Variation in Plant Defense more Strongly Affects Survival of an Herbivore than Its Solitary Parasitoid Wasp

The performance of natural enemies, such as parasitoid wasps, is affected by differences in the quality of the host’s diet, frequently mediated by species or population-related differences in plant allelochemistry. Here, we compared survival, development time, and body mass in a generalist herbivore, the cabbage moth, Mamestra brassicae, and its solitary endoparasitoid, Microplitis mediator, when reared on two cultivated (CYR and STH) and three wild (KIM, OH, and WIN) populations of cabbage, Brassica oleracea. Plants either were undamaged or induced by feeding of larvae of the cabbage butterfly, Pieris rapae. Development and biomass of M. brassicae and Mi. mediator were similar on both cultivated and one wild cabbage population (KIM), intermediate on the OH population, and significantly lower on the WIN population. Moreover, development was prolonged and biomass was reduced on herbivore-induced plants. However, only the survival of parasitized hosts (and not that of healthy larvae) was affected by induction. Analysis of glucosinolates in leaves of the cabbages revealed higher levels in the wild populations than cultivars, with the highest concentrations in WIN plants. Multivariate statistics revealed a negative correlation between insect performance and total levels of glucosinolates (GS) and levels of 3-butenyl GS. However, GS chemistry could not explain the reduced performance on induced plants since only indole GS concentrations increased in response to herbivory, which did not affect insect performance based on multivariate statistics. This result suggests that, in addition to aliphatic GS, other non-GS chemicals are responsible for the decline in insect performance, and that these chemicals affect the parasitoid more strongly than the host. Remarkably, when developing on WIN plants, the survival of Mi. mediator to adult eclosion was much higher than in its host, M. brassicae. This may be due to the fact that hosts parasitized by Mi. mediator pass through fewer instars, and host growth is arrested when they are only a fraction of the size of healthy caterpillars. Certain aspects of the biology and life-history of the host and parasitoid may determine their response to chemical challenges imposed by the food plant

Tracing animal genomic evolution with the chromosomal-level assembly of the freshwater sponge Ephydatia muelleri

Abstract The genomes of non-bilaterian metazoans are key to understanding the molecular basis of early animal evolution. However, a full comprehension of how animal-specific traits such as nervous systems arose is hindered by the scarcity and fragmented nature of genomes from key taxa, such as Porifera. Ephydatia muelleri is a freshwater sponge found across the northern hemisphere. Here we present its 326 Mb genome, assembled to high contiguity (N50: 9.88 Mb) with 23 chromosomes on 24 scaffolds. Our analyses reveal a metazoan-typical genome architecture, with highly shared synteny across Metazoa, and suggest that adaptation to the extreme temperatures and conditions found in freshwater often involves gene duplication. The pancontinental distribution and ready laboratory culture of E. muelleri make this a highly practical model system, which with RNAseq, DNA methylation and bacterial amplicon data spanning its development and range allows exploration of genomic changes both within sponges and in early animal evolution