Transport Of Metabolites Research Articles

Whole genome-scale assessment of gene fitness of Novosphingobium aromaticavorans during spaceflight

In microgravity, bacteria undergo intriguing physiological adaptations. There have been few attempts to assess global bacterial physiological responses to microgravity, with most studies only focusing on a handful of individual systems. This study assessed the fitness of each gene in the genome of the aromatic compound-degrading Alphaproteobacterium Novosphingobium aromaticavorans during growth in spaceflight. This was accomplished using Comparative TnSeq, which involves culturing the same saturating transposon mutagenized library under two different conditions. To assess gene fitness, a novel comparative TnSeq analytical tool was developed, named TnDivA, that is particularly useful in leveraging biological replicates. In this approach, transposon diversity is represented numerically using a modified Shannon diversity index, which was then converted into effective transposon density. This transformation accounts for variability in read distribution between samples, such as cases where reads were dominated by only a few transposon inserts. Effective density values were analyzed using multiple statistical methods, including log2-fold change, least-squares regression analysis, and Welch’s t-test. The results obtained across applied statistical methods show a difference in the number of significant genes identified. However, the functional categories of genes important to growth in microgravity showed similar patterns. Lipid metabolism and transport, energy production, transcription, translation, and secondary metabolite biosynthesis and transport were shown to have high fitness during spaceflight. This suggests that core metabolic processes, including lipid and secondary metabolism, play an important role adapting to stress and promoting growth in microgravity.

Human mitochondrial uncoupling protein 3 functions as a metabolite transporter.

Since its discovery, a major debate about mitochondrial uncoupling protein 3 (UCP3) has been whether its metabolic actions result primarily from mitochondrial inner membrane proton transport, a process that decreases respiratory efficiency and ATP synthesis. However, UCP3 expression and activity are induced by conditions that would seem at odds with inefficient 'uncoupled' respiration, including fasting and exercise. Here, we demonstrate that the bacterially expressed human UCP3, reconstituted into liposomes, catalyses a strict exchange of aspartate, malate, sulphate and phosphate. The R282Q mutation abolishes the transport activity of the protein. Although the substrate specificity and inhibitor sensitivity of UCP3 display similarity with that of its close homolog UCP2, the two proteins significantly differ in their transport mode and kinetic constants.

Metabolomic Profiling of Obese Patients with Altered Intestinal Permeability Undergoing a Very Low-Calorie Ketogenic Diet.

A healthy intestinal permeability facilitates the selective transport of nutrients, metabolites, water, and bacterial products, involving cellular, neural, hormonal, and immune factors. An altered intestinal permeability indicates pathologic phenotypes and is associated with the exacerbation of obesity and related comorbidities. To investigate the impact of altered permeability in obese patients undergoing a calorie-restrictive dietary regimen (VLCKD), we collected urinary and fecal samples from obese patients with both normal and altered permeability (determined based on the lactulose/mannitol ratio) before and after treatment. The analysis of volatile organic compounds (VOCs) aids in understanding the metabolites produced by the intestinal microbiota in this unique ecological niche. Furthermore, we examined clinical and anthropometric variables from the cohort and compared them to significant VOC panels. Consequently, we identified specific markers in the metabolomics data that differentiated between normal and altered profiles before and after the diet. These markers indicated how the variable contribution specifically accounted for interleukins and lipopolysaccharides (LPS). The targeted metabolomics experiment detected no differences in measured short-chain fatty acids (SCFA). In summary, our study evaluated metabolomic markers capable of distinguishing low-grade inflammation conditions, exacerbated in more advanced stages of obesity with altered intestinal permeability.

Screening of Saccharomyces cerevisiae metabolite transporters by 13C isotope substrate labeling.

The transportome of Saccharomyces cerevisiae comprises approximately 340 membrane-bound proteins, of which very few are well-characterized. Elucidating transporter proteins' function is essential not only for understanding central cellular processes in metabolite exchange with the external milieu but also for optimizing the production of value-added compounds in microbial cell factories. Here, we describe the application of 13C-labeled stable isotopes and detection by targeted LC-MS/MS as a screening tool for identifying Saccharomyces cerevisiae metabolite transporters. We compare the transport assay's sensitivity, reproducibility, and accuracy in yeast transporter mutant cell lines and Xenopus oocytes. As proof of principle, we analyzed the transport profiles of five yeast amino acid transporters. We first cultured yeast transporter deletion or overexpression mutants on uniformly labeled 13C-glucose and then screened their ability to facilitate the uptake or export of an unlabeled pool of amino acids. Individual transporters were further studied by heterologous expression in Xenopus oocytes, followed by an uptake assay with 13C labeled yeast extract. Uptake assays in Xenopus oocytes showed higher reproducibility and accuracy. Although having lower accuracy, the results from S. cerevisiae indicated the system's potential for initial high-throughput screening for native metabolite transporters. We partially confirmed previously reported substrates for all five amino acid transporters. In addition, we propose broader substrate specificity for two of the transporter proteins. The method presented here demonstrates the application of a comprehensive screening platform for the knowledge expansion of the transporter-substrate relationship for native metabolites in S. cerevisiae.

A novel complementary pathway of cordycepin biosynthesis in Cordyceps militaris

We determined whether there exists a complementary pathway of cordycepin biosynthesis in wild-type Cordyceps militaris, high-cordycepin-producing strain C. militaris GYS60, and low-cordycepin-producing strain C. militaris GYS80. Differentially expressed genes were identified from the transcriptomes of the three strains. Compared with C. militaris, in GYS60 and GYS80, we identified 145 and 470 upregulated and 96 and 594 downregulated genes. Compared with GYS80, in GYS60, we identified 306 upregulated and 207 downregulated genes. Gene Ontology analysis revealed that upregulated genes were mostly involved in detoxification, antioxidant, and molecular transducer in GYS60. By Clusters of Orthologous Groups of Proteins and Kyoto Encyclopedia of Genes and Genomes analyses, eight genes were significantly upregulated: five genes related to purine metabolism, one to ATP production, one to secondary metabolite transport, and one to RNA degradation. In GYS60, cordycepin was significantly increased by upregulation of ATP production, which promoted 3′,5′-cyclic AMP production. Cyclic AMP accelerated 3′-AMP accumulation, and cordycepin continued to be synthesized and exported. We verified the novel complementary pathway by adding the precursor adenosine and analyzing the expression of four key genes involved in the main pathway of cordycepin biosynthesis. Adenosine addition increased cordycepin production by 51.2% and 10.1%, respectively, in C. militaris and GYS60. Four genes in the main pathway in GYS60 were not upregulated.

Proteomic investigation reveals the role of bacterial laccase from Bacillus pumilus in oxidative stress defense

The wide distribution of laccases in nature makes them involved in different biological processes. However, little information is known about how laccase participates in the defense machinery of bacteria against oxidative stress. The present study aimed to elucidate the oxidative stress response mechanism of Bacillus pumilus ZB1 and the functional role of bacterial laccase in stress defense. The oxidative stress caused by methyl methanesulfonate (MMS) significantly induced laccase activity and its transcript level. The morphological analysis revealed that the defense of B. pumilus ZB1 against oxidative stress was activated. Based on the proteomic study, 114 differentially expressed proteins (DEPs) were up-regulated and 79 DEPs were down-regulated. In COG analysis, 66.40% DEPs were classified into the category “Metabolism”. We confirmed that laccase was up-regulated in response to MMS stress and its functional annotation was related to “Secondary metabolites biosynthesis, transport and catabolism”. Based on protein-protein interaction prediction, two up-regulated DEPs (YcnJ and GabP) showed interaction with laccase and contributed to the formation of laccase stability and adaptability. The overexpressed laccase might improve the antioxidative property of B. pumilus ZB1. These findings provide an insight and the guidelines for better exploitation of bioremediation using bacterial laccase. SignificanceBacillus pumilus is a gram-positive bacterium that has the potential for many applications, such as bioremediation. The expression of bacterial laccase is significantly influenced by oxidative stress, while the underlying mechanism of laccase overexpression in bacteria has not been fully studied. Elucidation of the biological process may benefit the bioremediation using bacteria in the future. In this study, the differentially expressed proteins were analyzed using a TMT-labeling proteomic approach when B. pumilus was treated with methyl methanesulfonate (MMS). Reactive oxygen species induced by MMS activated the secondary metabolites biosynthesis, transport, and catabolism in B. pumilus, including laccase overexpression. Moreover, the simultaneously up-regulated YcnJ and GabP may benefit the synthesis and the stability of laccase, then improve the antioxidative property of B. pumilus against environmental stress. Our findings advance the understanding of the adaptive mechanism of B. pumilus to environmental conditions.

Non-enzymatic glycation reduces glucose transport in the human cartilage endplate independently of matrix porosity or proteoglycan content.

Intervertebral disc degeneration is associated with low back pain, which is a leading cause of disability. While the precise causes of disc degeneration are unknown, inadequate nutrient and metabolite transport through the cartilage endplate (CEP) may be one important factor. Prior work shows that CEP transport properties depend on the porosity of the CEP matrix, but little is known about the role of CEP characteristics that could influence transport properties independently from porosity. Here, we show that CEP transport properties depend on the extent of non-enzymatic glycation of the CEP matrix. Using in vitro ribosylation to induce non-enzymatic glycation and promote the formation of advanced glycation end products, we found that ribosylation reduced glucose partition coefficients in human cadaveric lumbar CEP tissues by 10.7%, on average, compared with donor- and site-matched CEP tissues that did not undergo ribosylation (p = 0.04). These reductions in glucose uptake were observed in the absence of differences in CEP porosity (p = 0.89) or in the amounts of sulfated glycosaminoglycans (sGAGs, p = 0.47) or collagen (p = 0.61). To investigate whether ribosylation altered electrostatic interactions between fixed charges on the sGAG molecules and the mobile free ions, we measured the charge density in the CEP matrix using equilibrium partitioning of a cationic contrast agent using micro-computed tomography. After contrast enhancement, mean X-ray attenuation was 11.9% lower in the CEP tissues that had undergone ribosylation (p = 0.02), implying the CEP matrix was less negatively charged. Taken together, these findings indicate that non-enzymatic glycation negatively impacts glucose transport in the CEP independent of matrix porosity or sGAG content and that the effects may be mediated by alterations to matrix charge density.

The Root-Colonizing Endophyte Piriformospora indica Supports Nitrogen-Starved Arabidopsis thaliana Seedlings with Nitrogen Metabolites.

The root-colonizing endophytic fungus Piriformospora indica promotes the root and shoot growth of its host plants. We show that the growth promotion of Arabidopsis thaliana leaves is abolished when the seedlings are grown on media with nitrogen (N) limitation. The fungus neither stimulated the total N content nor did it promote 15NO3- uptake from agar plates to the leaves of the host under N-sufficient or N-limiting conditions. However, when the roots were co-cultivated with 15N-labelled P. indica, more labels were detected in the leaves of N-starved host plants but not in plants supplied with sufficient N. Amino acid and primary metabolite profiles, as well as the expression analyses of N metabolite transporter genes suggest that the fungus alleviates the adaptation of its host from the N limitation condition. P. indica alters the expression of transporter genes, which participate in the relocation of NO3-, NH4+ and N metabolites from the roots to the leaves under N limitation. We propose that P. indica participates in the plant's metabolomic adaptation against N limitation by delivering reduced N metabolites to the host, thus alleviating metabolic N starvation responses and reprogramming the expression of N metabolism-related genes.

Contribution of radixin and ezrin to the maintenance of hepatocytes' excretory function in health and disease

Background & aimsExcretory liver failure is frequently associated with poor prognosis in critically ill patients. It is characterized by the loss of canalicular membrane export pumps at the hepatocyte membrane. The membrane export pump Multidrug resistant-associated protein (MRP) 2 is pivotal in hepatocytes for brushed membrane morphology and transport of various metabolites. In addition, MRP2 anchoring proteins of the Ezrin/Radixin/Moesin (ERM) family are crucial for the correct MRP2 location, integration, and function in different tissues. In hepatocytes, altered ERM signaling is elementary for developing excretory liver failure. MethodsPolarized human HepaRG cells, primary human hepatocytes, and hepatocyte-specific Ezrin knockout mice are employed to investigate ERM expression and function in health and the bile duct ligation model of obstructive cholestasis. ResultsERM-scaffolding protein Ezrin has no relevant function in maintaining the canalicular structure in hepatocytes during health and disease. ConclusionsHomeostasis of the canalicular pole in hepatocytes is maintained exclusively by Radixin but not Ezrin, and Radixin dysfunction promotes cholestasis.

Non-alcoholic fatty liver disease and gut microbial dysbiosis- underlying mechanisms and gut microbiota mediated treatment strategies.

Non-alcoholic fatty liver disease (NAFLD) is by far the most prevalent form of liver disease worldwide. It's also the leading cause of liver-related hospitalizations and deaths. Furthermore, there is a link between obesity and NAFLD risk. A projected 25% of the world's population grieves from NAFLD, making it the most common chronic liver disorder. Several factors, such as obesity, oxidative stress, and insulin resistance, typically accompany NAFLD. Weight loss, lipid-lowering agents, thiazolidinediones, and metformin help prominently control NAFLD. Interestingly, pre-clinical studies demonstrate gut microbiota's potential causal role in NAFLD. Increased intestinal permeability and unhindered transport of microbial metabolites into the liver are the major disruptions due to gut microbiome dysbiosis, contributing to the development of NAFLD by dysregulating the gut-liver axis. Hence, altering the pathogenic bacterial population using probiotics, prebiotics, synbiotics, and fecal microbiota transplantation (FMT) could benefit patients with NAFLD. Therefore, it is crucial to acknowledge the importance of microbiota-mediated therapeutic approaches for NAFLD and comprehend the underlying mechanisms that establish a connection between NAFLD and gut microbiota. This review provides a comprehensive overview of the affiliation between dysbiosis of gut microbiota and the progress of NAFLD, as well as the potential benefits of prebiotic, probiotic, synbiotic supplementation, and FMT in obese individuals with NAFLD.

Large-scale genomic analyses with machine learning uncover predictive patterns associated with fungal phytopathogenic lifestyles and traits

Invasive plant pathogenic fungi have a global impact, with devastating economic and environmental effects on crops and forests. Biosurveillance, a critical component of threat mitigation, requires risk prediction based on fungal lifestyles and traits. Recent studies have revealed distinct genomic patterns associated with specific groups of plant pathogenic fungi. We sought to establish whether these phytopathogenic genomic patterns hold across diverse taxonomic and ecological groups from the Ascomycota and Basidiomycota, and furthermore, if those patterns can be used in a predictive capacity for biosurveillance. Using a supervised machine learning approach that integrates phylogenetic and genomic data, we analyzed 387 fungal genomes to test a proof-of-concept for the use of genomic signatures in predicting fungal phytopathogenic lifestyles and traits during biosurveillance activities. Our machine learning feature sets were derived from genome annotation data of carbohydrate-active enzymes (CAZymes), peptidases, secondary metabolite clusters (SMCs), transporters, and transcription factors. We found that machine learning could successfully predict fungal lifestyles and traits across taxonomic groups, with the best predictive performance coming from feature sets comprising CAZyme, peptidase, and SMC data. While phylogeny was an important component in most predictions, the inclusion of genomic data improved prediction performance for every lifestyle and trait tested. Plant pathogenicity was one of the best-predicted traits, showing the promise of predictive genomics for biosurveillance applications. Furthermore, our machine learning approach revealed expansions in the number of genes from specific CAZyme and peptidase families in the genomes of plant pathogens compared to non-phytopathogenic genomes (saprotrophs, endo- and ectomycorrhizal fungi). Such genomic feature profiles give insight into the evolution of fungal phytopathogenicity and could be useful to predict the risks of unknown fungi in future biosurveillance activities.

C4 maize and sorghum are more sensitive to rapid dehydration than C3 wheat and sunflower.

The high productive potential, heat resilience, and greater water use efficiency of C4 over C3 plants attract considerable interest in the face of global warming and increasing population, but C4 plants are often sensitive to dehydration, questioning the feasibility of their wider adoption. To resolve the primary effect of dehydration from slower from secondary leaf responses originating within leaves to combat stress, we conducted an innovative dehydration experiment. Four crops grown in hydroponics were forced to a rapid yet controlled decrease in leaf water potential by progressively raising roots of out of the solution while measuring leaf gas exchange. We show that, under rapid dehydration, assimilation decreased more steeply in C4 maize and sorghum than in C3 wheat and sunflower. This reduction was due to a rise of nonstomatal limitation at triple the rate in maize and sorghum than in wheat and sunflower. Rapid reductions in assimilation were previously measured in numerous C4 species across both laboratory and natural conditions. Hence, we deduce that high sensitivity to rapid dehydration might stem from the disturbance of an intrinsic aspect of C4 bicellular photosynthesis. We posit that an obstruction to metabolite transport between mesophyll and bundle sheath cells could be the cause.

Cortistatin deficiency reveals a dysfunctional brain endothelium with impaired gene pathways, exacerbated immune activation, and disrupted barrier integrity

BackgroundBrain activity governing cognition and behaviour depends on the fine-tuned microenvironment provided by a tightly controlled blood–brain barrier (BBB). Brain endothelium dysfunction is a hallmark of BBB breakdown in most neurodegenerative/neuroinflammatory disorders. Therefore, the identification of new endogenous molecules involved in endothelial cell disruption is essential to better understand BBB dynamics. Cortistatin is a neuroimmune mediator with anti-inflammatory and neuroprotective properties that exerts beneficial effects on the peripheral endothelium. However, its role in the healthy and injured brain endothelium remains to be evaluated. Herein, this study aimed to investigate the potential function of endogenous and therapeutic cortistatin in regulating brain endothelium dysfunction in a neuroinflammatory/neurodegenerative environment.MethodsWild-type and cortistatin-deficient murine brain endothelium and human cells were used for an in vitro barrier model, where a simulated ischemia-like environment was mimicked. Endothelial permeability, junction integrity, and immune response in the presence and absence of cortistatin were evaluated using different size tracers, immunofluorescence labelling, qPCR, and ELISA. Cortistatin molecular mechanisms underlying brain endothelium dynamics were assessed by RNA-sequencing analysis. Cortistatin role in BBB leakage was evaluated in adult mice injected with LPS.ResultsThe endogenous lack of cortistatin predisposes endothelium weakening with increased permeability, tight-junctions breakdown, and dysregulated immune activity. We demonstrated that both damaged and uninjured brain endothelial cells isolated from cortistatin-deficient mice, present a dysregulated and/or deactivated genetic programming. These pathways, related to basic physiology but also crucial for the repair after damage (e.g., extracellular matrix remodelling, angiogenesis, response to oxygen, signalling, and metabolites transport), are dysfunctional and make brain endothelial barrier lacking cortistatin non-responsive to any further injury. Treatment with cortistatin reversed in vitro hyperpermeability, tight-junctions disruption, inflammatory response, and reduced in vivo BBB leakage.ConclusionsThe neuropeptide cortistatin has a key role in the physiology of the cerebral microvasculature and its presence is crucial to develop a canonical balanced response to damage. The reparative effects of cortistatin in the brain endothelium were accompanied by the modulation of the immune function and the rescue of barrier integrity. Cortistatin-based therapies could emerge as a novel pleiotropic strategy to ameliorate neuroinflammatory/neurodegenerative disorders with disrupted BBB.

A Simulation of the Effects of Diffusion on Hyperpolarized [1-13C]-Pyruvate Signal Evolution.

Hyperpolarized [1-13C]-pyruvate magnetic resonance imaging is an emerging metabolic imaging method that offers unprecedented spatiotemporal resolution for monitoring tumor metabolism in vivo. To establish robust imaging biomarkers of metabolism, we must characterize phenomena that may modulate the apparent pyruvate-to-lactate conversion rate (kPL). Here, we investigate the potential effect of diffusion on pyruvate-to-lactate conversion, as failure to account for diffusion in pharmacokinetic analysis may obscure true intracellular chemical conversion rates. Changes in hyperpolarized pyruvate and lactate signal were calculated using a finite-difference time domain simulation of a two-dimensional tissue model. Signal evolution curves with intracellular kPL values from 0.02 to 1.00 s-1 were analyzed using spatially invariant one-compartment and two-compartment pharmacokinetic models. A second spatially variant simulation incorporating compartmental instantaneous mixing was fit with the same one-compartment model. When fitting with the one-compartment model, apparent kPL underestimated intracellular kPL by approximately 50% at an intracellular kPL of 0.02 s-1. This underestimation increased for larger kPL values. However, fitting the instantaneous mixing curves showed that diffusion accounted for only a small part of this underestimation. Fitting with the two-compartment model yielded more accurate intracellular kPL values. This work suggests diffusion is not a significant rate-limiting factor in pyruvate-to-lactate conversion given that our model assumptions hold true. In higher order models, diffusion effects may be accounted for by a term characterizing metabolite transport. Pharmacokinetic models used to analyze hyperpolarized pyruvate signal evolution should focus on carefully selecting the analytical model for fitting rather than accounting for diffusion effects.

D-2-hydroxyglutarate regulates human brain vascular endothelial cell proliferation and barrier function.

Gain-of-function mutations in isocitrate dehydrogenase (IDH) genes result in excessive production of (D)-2-hydroxyglutarate (D-2HG) which intrinsically modifies tumor cell epigenetics and impacts surrounding noncancerous cells through nonepigenetic pathways. However, whether D-2HG has a paracrine effect on endothelial cells in the tumor microenvironment needs further clarification. We quantified microvessel density by immunohistochemistry using tissue sections from 60 high-grade astrocytic gliomas with or without IDH mutation. Microvessel density was found to be reduced in tumors carrying an IDH mutation. Ex vivo experiments showed that D-2HG inhibited endothelial cell migration, wound healing, and tube formation by suppressing cell proliferation but not viability, possibly through reduced activation of the mTOR/STAT3 pathway. Further, D-2HG reduced fluorescent dextran permeability and decreased paracellular T-cell transendothelial migration by augmenting expression of junctional proteins thereby collectively increasing endothelial barrier function. These results indicate that D-2HG may influence the tumor vascular microenvironment by reducing the intratumoral vasculature density and by inhibiting the transport of metabolites and extravasation of circulating cells into the astrocytoma microenvironment. These observations provide a rationale for combining IDH inhibition with antitumor immunological/angiogenic approaches and suggest a molecular basis for resistance to antiangiogenic drugs in patients whose tumors express a mutant IDH allele.

γ-Aminobutyric Acid Priming Alleviates Acid-Aluminum Toxicity to Creeping Bentgrass by Regulating Metabolic Homeostasis.

Aluminum (Al) toxicity is a major limiting factor for plant growth and crop production in acidic soils. This study aims to investigate the effects of γ-aminobutyric acid (GABA) priming on mitigating acid-Al toxicity to creeping bentgrass (Agrostis stolonifera) associated with changes in plant growth, photosynthetic parameters, antioxidant defense, key metabolites, and genes related to organic acids metabolism. Thirty-seven-old plants were primed with or without 0.5 mM GABA for three days and then subjected to acid-Al stress (5 mmol/L AlCl3·6H2O, pH 4.35) for fifteen days. The results showed that acid-Al stress significantly increased the accumulation of Al and also restricted aboveground and underground growths, photosynthesis, photochemical efficiency, and osmotic balance, which could be effectively alleviated by GABA priming. The application of GABA significantly activated antioxidant enzymes, including superoxide dismutase, peroxidase, catalase, and ascorbate peroxidase, to reduce oxidative damage to cells under acid-Al stress. Metabolomics analysis demonstrated that the GABA pretreatment significantly induced the accumulation of many metabolites such as quinic acid, pyruvic acid, shikimic acid, glycine, threonine, erythrose, glucose-6-phosphate, galactose, kestose, threitol, ribitol, glycerol, putrescine, galactinol, and myo-inositol associated with osmotic, antioxidant, and metabolic homeostases under acid-Al stress. In addition, the GABA priming significantly up-regulated genes related to the transportation of malic acid and citric acid in leaves in response to acid-Al stress. Current findings indicated GABA-induced tolerance to acid-Al stress in relation to scavenging of reactive oxygen species, osmotic adjustment, and accumulation and transport of organic metabolites in leaves. Exogenous GABA priming could improve the phytoremediation potential of perennial creeping bentgrass for the restoration of Al-contaminated soils.

Role of ABC Proteins in Secondary Metabolism and Immune (=Defensive) Response in Seaweeds.

Laurencia seaweed species synthesize a broad range of secondary metabolites, mainly terpenes (e.g., elatol), exhibiting diverse ecological roles, such as defense against fouling and herbivores. Recently, an intricate cellular machinery was described concerning terpenes biosynthetic pathways, storage inside corps en cerise (CC), and regulated exocytosis in these species. But for seaweeds in general, the proteins involved in transmembrane transport of secondary metabolites remain unknown. Assays with Rhodamine-123 and cyclosporine A (CSA) revealed the presence of ABC transporters in CC membrane of Laurencia dendroidea. In vivo incubation assays with CSA resulted in CC morphological changes, reduced intracellular elatol concentrations, and increased biofouling cover on the seaweed surface. Cultivation assays in the presence of a marine pathogenic bacteria induced the expression of ABC proteins belonging to the subfamilies ABCB, ABCD, ABCF, and ABCG. The latter subfamily is known to be associated with the transport of plant terpenes. Our results shed new light on the role of ABC proteins in key mechanisms of the defensive system in seaweeds against fouling and herbivory.

Chitin Nanofibrils Enabled Core-Shell Microcapsules of Alginate Hydrogel.

An engineered 3D architectural network of the biopolymeric hydrogel can mimic the native cell environment that promotes cell infiltration and growth. Among several bio-fabricated hydrogel structures, core-shell microcapsules inherit the potential of cell encapsulation to ensure the growth and transport of cells and cell metabolites. Herein, a co-axial electrostatic encapsulation strategy is used to create and encapsulate the cells into chitin nanofibrils integrated alginate hydrogel microcapsules. Three parameters that are critical in the electrostatic encapsulation process, hydrogel composition, flow rate, and voltage were optimized. The physicochemical characterization including structure, size, and stability of the core-shell microcapsules was analyzed by scanning electron microscope (SEM), FTIR, and mechanical tests. The cellular responses of the core-shell microcapsules were evaluated through in vitro cell studies by encapsulating NIH/3T3 fibroblast cells. Notably, the bioactive microcapsule showed that the cell viability was found excellent for more than 2 weeks. Thus, the results of this core-shell microcapsule showed a promising approach to creating 3D hydrogel networks suitable for different biomedical applications such as in vitro tissue models for toxicity studies, wound healing, and tissue repair.

Phylogenetic Analysis Guides Transporter Protein Deorphanization: A Case Study of the SLC25 Family of Mitochondrial Metabolite Transporters.

Homology search and phylogenetic analysis have commonly been used to annotate gene function, although they are prone to error. We hypothesize that the power of homology search in functional annotation depends on the coupling of sequence variation to functional diversification, and we herein focus on the SoLute Carrier (SLC25) family of mitochondrial metabolite transporters to survey this coupling in a family-wide manner. The SLC25 family is the largest family of mitochondrial metabolite transporters in eukaryotes that translocate ligands of different chemical properties, ranging from nucleotides, amino acids, carboxylic acids and cofactors, presenting adequate experimentally validated functional diversification in ligand transport. Here, we combine phylogenetic analysis to profile SLC25 transporters across common eukaryotic model organisms, from Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, to Homo sapiens, and assess their sequence adaptations to the transported ligands within individual subfamilies. Using several recently studied and poorly characterized SLC25 transporters, we discuss the potentials and limitations of phylogenetic analysis in guiding functional characterization.

Comprehensive Transcriptome and Metabolome Analyses Reveal Primary Molecular Regulation Pathways Involved in Peanut under Water and Nitrogen Co-Limitation.

The yield and quality of peanut (Arachis hypogaea L.), an oil crop planted worldwide, are often limited by drought stress (DS) and nitrogen (N) deficiency. To investigate the molecular mechanism by which peanut counteracts DS and N deficiency, we conducted comprehensive transcriptomic and metabolomic analyses of peanut leaves. Herein, 829 known differentially accumulated metabolites, 324 differentially expressed transcription factors, and 5294 differentially expressed genes (DEGs) were identified under different water and N conditions. The transcriptome analysis demonstrated that drought-related DEGs were predominantly expressed in "glycolysis/gluconeogenesis" and "glycerolipid metabolism", while N-deficiency-related DEGs were mainly expressed in starch and sucrose metabolism, as well as in the biosynthesis of amino acid pathways. The biosynthesis, transport, and catabolism of secondary metabolites accounted for a large proportion of the 1317 DEGs present in water and N co-limitation. Metabolomic analysis showed that the metabolic accumulation of these pathways was significantly dependent on the stress conditions. Additionally, the roles of metabolites and genes in these pathways, such as the biosynthesis of amino acids and phenylpropanoid biosynthesis under different stress conditions, were discussed. The results demonstrated that different genes, metabolic pathways, and metabolites were related to DS and N deficiency. Thus, this study elucidates the metabolic pathways and functional genes that can be used for the improvement of peanut resistance to abiotic stress.