Transcription Factor Engineering in Aspergillus nidulans Leads to the Discovery of an Orsellinaldehyde Derivative Produced via an Unlinked Polyketide Synthase Gene.

Secondary metabolites are generally produced by enzymes encoded by genes within a biosynthetic gene cluster. Transcription factor genes are frequently located within these gene clusters. These transcription factors often drive expression of the other genes of the biosynthetic gene cluster, and overexpression of the transcription factor provides a facile approach to express all genes within a gene cluster, resulting in production of downstream metabolite(s). Unfortunately this approach is not always successful, leading us to engineer more effective hybrid transcription factors. Herein, we attempted to activate a putative cryptic biosynthetic gene cluster in Aspergillus nidulans using a combination of transcription factor engineering and overexpression approaches. This resulted in the discovery of a novel secondary metabolite we term triorsellinaldehyde. Surprisingly, deletion of the polyketide synthase gene within the gene cluster did not prevent triorsellinaldehyde production. However, targeted deletion of a polyketide synthase gene elsewhere in the genome revealed its role in triorsellinaldehyde biosynthesis.

A Dynamic Foundation: Aberrations of Sleep Architecture and Its Association With Clinical and Sub-clinical Psychopathology.

This investigation centered on the hypnagogic and hypnopompic wake-sleep/sleep-wake transition states and the associated exploration of hypnagogic and hypnopompic experiences (HHEs), and sleep paralysis (SP) on psychiatric exacerbation and paradoxical masking. The study aims to discern causality by examining how these sleep-related experiences may contribute to the emergence or exacerbation of psychiatric and neurodegenerative conditions, particularly, pertaining to the clinical or sub-clinical demographic of Schizotypal Personality Disorder (STPD), Mood Disorders, Schizophrenia, Post-Traumatic Stress Disorder (PTSD), Generalized Anxiety Disorder (GAD), Narcolepsy, Panic Disorder, specific phobias, or heightened psychotic sensitivity. Methodologically, this study employed a comprehensive literature review, drawing from a range of studies across sleep medicine, psychiatry, and psychology, utilizing PubMed-indexed peer-reviewed scientific literature, sourcing from academic institutions, Google Scholar, and open-access publications. This interdisciplinary approach allowed for a nuanced and systematic understanding of the potential links between specific sleep-wake/wake-sleep aberrations and their masking or exacerbation of clinical/sub-clinical psychiatric symptomatology in this particular demographic. Insights gained from the outcome of this study hold promise for advancing understanding of the interrelationship between sleep neurobiology and psychiatric disorders. Additionally, the findings may inform targeted therapeutic interventions tailored to mitigate the impact of sleep-wake disruptions on vulnerable populations. The overarching objective is to bridge current gaps in knowledge, cultivating a more profound understanding with direct implications for both clinical practice and ongoing research endeavors. The study outcomes provide an intriguing understanding of the complex relationship between sleep neurobiology and psychiatric disorders, paving the way for targeted therapeutic interventions to alleviate the effects of sleep-wake disruptions, and addressing critical gaps in knowledge with direct implications for clinical practice and ongoing research.

A heterologous expression platform in Aspergillus nidulans for the elucidation of cryptic secondary metabolism biosynthetic gene clusters: discovery of the Aspergillus fumigatus sartorypyrone biosynthetic pathway.

Aspergillus fumigatus is a serious human pathogen causing life-threatening Aspergillosis in immunocompromised patients. Secondary metabolites (SMs) play an important role in pathogenesis, but the products of many SM biosynthetic gene clusters (BGCs) remain unknown. In this study, we have developed a heterologous expression platform in Aspergillus nidulans, using a newly created genetic dereplication strain, to express a previously unknown BGC from A. fumigatus and determine its products. The BGC produces sartorypyrones, and we have named it the spy BGC. Analysis of targeted gene deletions by HRESIMS, NMR, and microcrystal electron diffraction (MicroED) enabled us to identify 12 products from the spy BGC. Seven of the compounds have not been isolated previously. We also individually expressed the polyketide synthase (PKS) gene spyA and demonstrated that it produces the polyketide triacetic acid lactone (TAL), a potentially important biorenewable platform chemical. Our data have allowed us to propose a biosynthetic pathway for sartorypyrones and related natural products. This work highlights the potential of using the A. nidulans heterologous expression platform to uncover cryptic BGCs from A. fumigatus and other species, despite the complexity of their secondary metabolomes.

Branched-chain amino acid biosynthesis in fungi.

Branched-chain amino acids (BCAAs)-isoleucine, leucine, and valine-are synthesized by fungi. These amino acids are important components of proteins and secondary metabolites. The biochemical pathway for BCAA biosynthesis is well-characterized in the yeast Saccharomyces cerevisiae. The biosynthesis of these three amino acids is interconnected. Different precursors are metabolized in multiple steps through shared enzymes to produce isoleucine and valine, and the valine biosynthesis pathway branches before the penultimate step to a series of leucine biosynthesis-specific steps to produce leucine. Recent efforts have made advances toward characterization of the BCAA biosynthesis pathway in several fungi, revealing diversity in gene duplication and functional divergence in the genes for these enzymatic steps in different fungi. The BCAA biosynthesis pathway is regulated by the transcription factor LEU3 in S. cerevisiae, and LeuB in Aspergillus nidulans and Aspergillus fumigatus, and the activity of these transcription factors is modulated by the leucine biosynthesis pathway intermediate α-isopropylmalate. Herein, we discuss recent advances in our understanding of the BCAA pathway and its regulation, focusing on filamentous ascomycete fungi and comparison with the well-established process in yeast.

Dendrite initiation and propagation in lithium metal solid-state batteries.

All-solid-state batteries with a Li anode and ceramic electrolyte have the potential to deliver a step change in performance compared with today's Li-ion batteries1,2. However, Li dendrites (filaments) form on charging at practical rates and penetrate the ceramic electrolyte, leading to short circuit and cell failure3,4. Previous models of dendrite penetration have generally focused on a single process for dendrite initiation and propagation, with Li driving the crack at its tip5-9. Here we show that initiation and propagation are separate processes. Initiation arises from Li deposition into subsurface pores, by means of microcracks that connect the pores to the surface. Once filled, further charging builds pressure in the pores owing to the slow extrusion of Li (viscoplastic flow) back to the surface, leading to cracking. By contrast, dendrite propagation occurs by wedge opening, with Li driving the dry crack from the rear, not the tip. Whereas initiation is determined by the local (microscopic) fracture strength at the grain boundaries, the pore size, pore population density and current density, propagation depends on the (macroscopic) fracture toughness of the ceramic, the length of the Li dendrite (filament) that partially occupies the dry crack, current density, stack pressure and the charge capacity accessed during each cycle. Lower stack pressures suppress propagation, markedly extending the number of cycles before short circuit in cells in which dendrites have initiated.

Extra-Uterine Leiomyoma Presenting After Hysterectomy.

Leiomyomas are a common gynaecological finding affecting 20-30% of women over the age of 35, with prevalence decreasing following menopause. Around 25% of women present clinically with a leiomyoma, which are most commonly found within the uterus. Rarer extra-uterine locations include the broad ligament, cervix, and vagina. We present a rare case of an extra-uterine leiomyoma located in the perineum of a 59-year-old female. Our case highlights the importance for extra-uterine leiomyoma to be considered as a differential diagnosis in patients presenting with a pelvic mass following hysterectomy.

Co-option of an extracellular protease for transcriptional control of nutrient degradation in the fungus Aspergillus nidulans.

Nutrient acquisition is essential for all organisms. Fungi regulate their metabolism according to environmental nutrient availability through elaborate transcription regulatory programs. In filamentous fungi, a highly conserved GATA transcription factor AreA and its co-repressor NmrA govern expression of genes involved in extracellular breakdown, uptake, and metabolism of nitrogen nutrients. Here, we show that the Aspergillus nidulans PnmB protease is a moonlighting protein with extracellular and intracellular functions for nitrogen acquisition and metabolism. PnmB serves not only as a secreted protease to degrade extracellular nutrients, but also as an intracellular protease to control the turnover of the co-repressor NmrA, accelerating AreA transcriptional activation upon nitrogen starvation. PnmB expression is controlled by AreA, which activates a positive feedback regulatory loop. Hence, we uncover a regulatory mechanism in the well-established controls determining the response to nitrogen starvation, revealing functional evolution of a protease gene for transcriptional regulation and extracellular nutrient breakdown.

Effect of Ion Irradiation on Nanoindentation Fracture and Deformation in Silicon Carbide.

Silicon carbide is desirable for many nuclear applications, making it necessary to understand how it deforms after irradiation. Ion implantation combined with nanoindentation is commonly used to measure radiation-induced changes to mechanical properties; hardness and modulus can be calculated from load-displacement curves, and fracture toughness can be estimated from surface crack lengths. Further insight into indentation deformation and fracture is required to understand the observed changes to mechanical properties caused by irradiation. This paper investigates indentation deformation using high-resolution electron backscatter diffraction (HR-EBSD) and Raman spectroscopy. Significant differences exist after irradiation: fracture is suppressed by swelling-induced compressive residual stresses, and the plastically deformed region extends further from the indentation. During focused ion beam cross-sectioning, indentation cracks grow, and residual stresses are modified. The results clarify the mechanisms responsible for the modification of apparent hardness and apparent indentation toughness values caused by the compressive residual stresses in ion-implanted specimens. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s11837-021-04636-8.

Duplication and Functional Divergence of Branched-Chain Amino Acid Biosynthesis Genes in Aspergillus nidulans.

Fungi, bacteria, and plants, but not animals, synthesize the branched-chain amino acids: leucine, isoleucine, and valine. While branched-chain amino acid (BCAA) biosynthesis has been well characterized in the yeast Saccharomyces cerevisiae, it is incompletely understood in filamentous fungi. The three BCAAs share several early biosynthesis steps before divergence into specific pathways. In Aspergillus nidulans, the genes for the first two dedicated steps in leucine biosynthesis have been characterized, but the final two have not. We used sequence searches of the A. nidulans genome to identify two genes encoding ß-isopropylmalate dehydrogenase, which catalyzes the penultimate step of leucine biosynthesis, and six genes encoding BCAA aminotransferase, which catalyzes the final step in biosynthesis of all three BCAA. We have used combinations of gene knockouts to determine the relative contribution of each of these genes to BCAA biosynthesis. While both ß-isopropylmalate dehydrogenase genes act in leucine biosynthesis, the two most highly expressed BCAA aminotransferases are responsible for BCAA biosynthesis. We have also characterized the expression of leucine biosynthesis genes using reverse transcriptase-quantitative PCR and found regulation in response to leucine availability is mediated through the Zn(II)2Cys6 transcription factor LeuB. IMPORTANCE Branched-chain amino acid (BCAA) biosynthesis is important for pathogenic fungi to successfully cause disease in human and plant hosts. The enzymes for their production are absent from humans and, therefore, provide potential antifungal targets. While BCAA biosynthesis is well characterized in yeasts, it is poorly understood in filamentous fungal pathogens. Developing a thorough understanding of both the genes encoding the metabolic enzymes for BCAA biosynthesis and how their expression is regulated will inform target selection for antifungal drug development.

Dietary mitigation of enteric methane emissions from ruminants: A review of plant tannin mitigation options.

Methane gas from livestock production activities is a significant source of greenhouse gas (GHG) emissions which have been shown to influence climate change. New technologies offer a potential to manipulate the rumen biome through genetic selection reducing CH4 production. Methane production may also be mitigated to varying degrees by various dietary intervention strategies. Strategies to reduce GHG emissions need to be developed which increase ruminant production efficiency whereas reducing production of CH4 from cattle, sheep, and goats. Methane emissions may be efficiently mitigated by manipulation of natural ruminal microbiota with various dietary interventions and animal production efficiency improved. Although some CH4 abatement strategies have shown efficacy in vivo, more research is required to make any of these approaches pertinent to modern animal production systems. The objective of this review is to explain how anti-methanogenic compounds (e.g., plant tannins) affect ruminal microbiota, reduce CH4 emission, and the effects on host responses. Thus, this review provides information relevant to understanding the impact of tannins on methanogenesis, which may provide a cost-effective means to reduce enteric CH4 production and the influence of ruminant animals on global GHG emissions.

Reactive Extrusion and Magnesium (II) N-Heterocyclic Carbene Catalyst in Continuous PLA Production.

Reactive extrusion and magnesium (II) N-heterocyclic carbene catalyst are successfully employed in continuous polylactide synthesis. The possibility of using six-membered N-heterocyclic carbene adducts to act as efficient catalysts towards the sustainable synthesis of poly(l-lactide) through ring-opening polymerization of l-lactide (LA) is first investigated in bulk batch reactions. Under optimized solvent-free conditions, polylactide (PLA) of moderate to high molecular weights and excellent optical activities are successfully achieved. These promising results are further applied in the continuous production of PLA in an extruder.

Biodegradable Drug-Delivery Peptide Nanocapsules.

Branched amphiphilic peptide capsules (BAPCs) are an efficient transport system that can deliver nucleic acids, small proteins, and solutes. The ability of BAPCs to break down is essential to their adoption as a delivery vehicle for human and agricultural applications. Until now, however, BAPCs were shown to be inert to mammalian degradation systems. Here, we demonstrate, using BAPCs encapsulating the toxic urea analogue thiourea, that the common soil fungus Aspergillus nidulans can degrade BAPCs. We provide evidence that this degradation is extracellular through the action of secreted proteases. Our data indicate that BAPCs are likely biodegradable in the environment.

Hybrid Transcription Factor Engineering Activates the Silent Secondary Metabolite Gene Cluster for (+)-Asperlin in Aspergillus nidulans.

Fungi are a major source of valuable bioactive secondary metabolites (SMs). These compounds are synthesized by enzymes encoded by genes that are clustered in the genome. The vast majority of SM biosynthetic gene clusters are not expressed under normal growth conditions, and their products are unknown. Developing methods for activation of these silent gene clusters offers the potential for discovering many valuable new fungal SMs. While a number of useful approaches have been developed, they each have limitations, and additional tools are needed. One approach, upregulation of SM gene cluster-specific transcription factors that are associated with many SM gene clusters, has worked extremely well in some cases, but it has failed more often than it has succeeded. Taking advantage of transcription factor domain modularity, we developed a new approach. We fused the DNA-binding domain of a transcription factor associated with a silent SM gene cluster with the activation domain of a robust SM transcription factor, AfoA. Expression of this hybrid transcription factor activated transcription of the genes in the target cluster and production of the antibiotic (+)-asperlin. Deletion of cluster genes confirmed that the cluster is responsible for (+)-asperlin production, and we designate it the aln cluster. Separately, coinduction of expression of two aln cluster genes revealed the pathway intermediate (2 Z,4 Z,6 E)-octa-2,4,6-trienoic acid, a compound with photoprotectant properties. Our findings demonstrate the potential of our novel synthetic hybrid transcription factor strategy to discover the products of other silent fungal SM gene clusters.

Use of new technologies to evaluate the environmental footprint of feedlot systems.

With increased concern over the effects of livestock production on the environment, a number of new technologies have evolved to help scientists evaluate the environmental footprint of beef cattle. The objective of this review was to provide an overview of some of those techniques. These techniques include methods to measure individual feed intake, enteric methane emissions, ground-level greenhouse gas and ammonia emissions, feedlot and pasture emissions, and identify potential pathogens. The appropriate method to use for measuring emissions will vary depending upon the type of emission, the emission source, and the goals of the research. These methods should also be validated to assure they produce accurate results and achieve the goals of the research project. In addition, we must not forget to properly use existing technologies and methods such as proper feed mixing, feeding management, feed/ingredient sampling, and nutrient analysis.

Unlocking the Potential of Poly(Ortho Ester)s: A General Catalytic Approach to the Synthesis of Surface-Erodible Materials.

Poly(ortho ester)s (POEs) are well-known for their surface-eroding properties and hence present unique opportunities for controlled-release and tissue-engineering applications. Their development and wide-spread investigation has, however, been severely limited by challenging synthetic requirements that incorporate unstable intermediates and are therefore highly irreproducible. Herein, the first catalytic method for the synthesis of POEs using air- and moisture-stable vinyl acetal precursors is presented. The synthesis of a range of POE structures is demonstrated, including those that are extremely difficult to achieve by other synthetic methods. Furthermore, application of this chemistry permits efficient installation of functional groups through ortho ester linkages on an aliphatic polycarbonate.

Temporal Nitrous Oxide Emissions from Beef Cattle Feedlot Manure after a Simulated Rainfall Event.

Nitrous oxide (NO) is a greenhouse gas (GHG) emitted from agricultural operations. The objective of this research was to quantify NO-N emissions from simulated open-lot beef cattle feedlot pens after rainfall. A recirculating-flow-through, non-steady state chamber system consisting of five 1-m steel pans was designed for quantifying emissions. A lid was placed sequentially on each pan, and headspace air was recirculated between the pan and a real-time NO analyzer, measuring concentrations every 1 s. Air-dried manure (89.2% dry matter) from a commercial feedlot in the Texas Panhandle was placed in the pans and then 0, 6.3, 12.7, 25.4, or 50.8 mm of water was applied to simulate a one-time rainfall event. Emissions of NO-N were monitored for 45 d, where two distinct episodes of NO-N production were observed over time. The first NO-N episode had a duration of 10 h and peaked 2 h after rainfall at a flux of 1.0 to 200 mg m h. The second episode had a duration of 40 d and peaked 15 d after rainfall at a flux of 0.06 to 35 mg m h. The second episode accounted for 69 to 91% of the cumulative NO-N emitted over the 45-d period. Each millimeter of rainfall increased cumulative NO-N emitted by 167.9 mg m ( = 0.99, < 0.001). This rainfall vs. cumulative emissions relationship will be useful for modeling annual NO-N emissions from open-lot beef cattle feedlots, and for assessing the effectiveness of best management practices for reducing feedlot GHG emissions.

Nitrous Oxide Emissions from Open-Lot Cattle Feedyards: A Review.

Nitrous oxide (NO) emissions from concentrated animal feeding operations, including cattle feedyards, have become an important research topic. However, there are limitations to current measurement techniques, uncertainty in the magnitude of feedyard NO fluxes, and a lack of effective mitigation methods. The objective of this review was to assess NO emission from cattle feedyards, including comparison of measured and modeled emission rates, discussion of measurement methods, and evaluation of mitigation options. Published annual per capita flux rates for beef cattle feedyards and open-lot dairies were highly variable and ranged from 0.002 to 4.3 kg NO animal yr. On an area basis, published emission rates ranged from 0 to 41 mg NO m h. From these studies and Intergovernmental Panel on Climate Change emission factors, calculated daily per capita NO fluxes averaged 18 ± 10 g NO animal d (range, 0.04-67 g NO animal d). This variation was due to inconsistency in measurement techniques as well as irregularity in NO production and emission attributable to management, animal diet, and environmental conditions. Based on this review, it is clear that the magnitude and dynamics of NO emissions from open-lot cattle systems are not well understood. Further research is required to quantify feedyard NO fluxes and develop cost-effective mitigation methods.

Tasking on Natural Statistics of Infrared Images.

Natural scene statistics (NSSs) provide powerful, perceptually relevant tools that have been successfully used for image quality analysis of visible light images. Since NSS capture statistical regularities that arise from the physical world, they are relevant to long wave infrared (LWIR) images, which differ from visible light images mainly by the wavelengths captured at the imaging sensors. We show that NSS models of bandpass LWIR images are similar to those of visible light images, but with different parameterizations. Using this difference, we exploit the power of NSS to successfully distinguish between LWIR images and visible light images. In addition, we study distortions unique to LWIR and find directional models useful for detecting the halo effect, simple bandpass models useful for detecting hotspots, and combinations of these models useful for measuring the degree of non-uniformity present in many LWIR images. For local distortion identification and measurement, we also describe a method for generating distortion maps using NSS features. To facilitate our evaluation, we analyze the NSS of LWIR images under pristine and distorted conditions, using four databases, each captured with a different IR camera. Predicting human performance for assessing distortion and quality in LWIR images is critical for task efficacy. We find that NSS features improve human targeting task performance prediction. Furthermore, we conducted a human study on the perceptual quality of noise-and blur-distorted LWIR images and create a new blind image quality predictor for IR images.