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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 721 records · Page 40
All sourcesNASA NTRSDOE OSTI
DOE OSTI2019

Integration of transcriptomic and proteomic data identifies biological functions in cell populations from human infant lung

Systems biology uses computational approaches to integrate diverse data types to understand cell and organ behavior. Data derived from complementary technologies, for example transcriptomic and proteomic analyses, are providing new insights into development and disease. We compared mRNA and protein profiles from purified endothelial, epithelial, immune, and mesenchymal cells from normal human lung tissue. Signatures for each cell type were identified and compared at both mRNA and protein levels. Cell specific biological processes and pathways were predicted by analysis of concordant and discordant RNA-protein pairs. Cell clustering and gene set enrichment comparisons identified shared versus unique processes associated with transcriptomic and/or proteomic data. Clear cell-cell correlations between mRNA and protein data were obtained from each cell type. Approximately 40% of RNA-protein pairs were coherently expressed. While the correlation between RNA and their protein products was relatively low (approximately 0.4), cell specific signature genes involved in functional processes characteristic of each cell type, were more highly correlated with their protein products. Consistency of cell specific RNA-protein signatures indicated an essential framework for the function of each cell type. Visualization and reutilization of the protein and RNA profiles are supported by a new web application, "LungProteomics," which is incorporated into the LGEA web portal and freely accessible at (https://research.cchmc.org/pbge/lunggens/lungProteomics/human_sorted_profiles.html).

human lung, sorted cells, integrated omics, Transc↗
DOE OSTI2017

Comparative genomics of geographically distant Fusarium fujikuroi isolates revealed two distinct pathotypes correlating with secondary metabolite profiles

Fusarium fujikuroi causes bakanae (“foolish seedling”) disease of rice which is characterized by hyper-elongation of seedlings resulting from production of gibberellic acids (GAs) by the fungus. This plant pathogen is also known for production of harmful mycotoxins, such as fusarins, fusaric acid, apicidin F and beauvericin. Recently, we generated the first de novo genome sequence of F. fujikuroi strain IMI 58289 combined with extensive transcriptional, epigenetic, proteomic and chemical product analyses. GA production was shown to provide a selective advantage during infection of the preferred host plant rice. Here, we provide genome sequences of eight additional F. fujikuroi isolates from distant geographic regions. The isolates differ in the size of chromosomes, most likely due to variability of subtelomeric regions, the type of asexual spores (microconidia and/or macroconidia), and the number and expression of secondary metabolite gene clusters. Whilst most of the isolates caused the typical bakanae symptoms, one isolate, B14, caused stunting and early withering of infected seedlings. In contrast to the other isolates, B14 produced no GAs but high amounts of fumonisins during infection on rice. Furthermore, it differed from the other isolates by the presence of three additional polyketide synthase (PKS) genes (PKS40, PKS43, PKS51) and the absence of the F. fujikuroi-specific apicidin F (NRPS31) gene cluster. Analysis of additional field isolates confirmed the strong correlation between the pathotype (bakanae or stunting/withering), and the ability to produce either GAs or fumonisins. Deletion of the fumonisin and fusaric acid-specific PKS genes in B14 reduced the stunting/withering symptoms, whereas deletion of the PKS51 gene resulted in elevated symptom development. Phylogenetic analyses revealed two subclades of F. fujikuroi strains according to their pathotype and secondary metabolite profiles.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Low-dose radiation affects cardiac physiology: gene networks and molecular signaling in cardiomyocytes

There are 160,000 cancer patients worldwide treated with particle radiotherapy (RT). With the advent of proton, and high (H) charge (Z) and energy (E) HZE ionizing particle RT, the cardiovascular diseases risk estimates are uncertain. In addition, future deep space exploratory-type missions will expose humans to unknown but low doses of particle irradiation (IR). Here, we examined molecular responses using transcriptome profiling in left ventricular murine cardiomyocytes isolated from mice that were exposed to 90 cGy, 1 GeV proton ( 1 H) and 15 cGy, 1 GeV/nucleon iron ( 56 Fe) over 28 days after exposure. Unsupervised clustering analysis of gene expression segregated samples according to the IR response and time after exposure, with 56 Fe-IR showing the greatest level of gene modulation. 1 H-IR showed little differential transcript modulation. Network analysis categorized the major differentially expressed genes into cell cycle, oxidative responses, and transcriptional regulation functional groups. Transcriptional networks identified key nodes regulating expression. Validation of the signal transduction network by protein analysis and gel shift assay showed that particle IR clearly regulates a long-lived signaling mechanism for ERK1/2, p38 MAPK signaling and identified NFATc4, GATA4, STAT3, and NF-κB as regulators of the response at specific time points. These data suggest that the molecular responses and gene expression to 56 Fe-IR in cardiomyocytes are unique and long-lasting. Our study may have significant implications for the efforts of National Aeronautics and Space Administration to develop heart disease risk estimates for astronauts and for patients receiving conventional and particle RT via identification of specific HZE-IR molecular markers.

62 RADIOLOGY AND NUCLEAR MEDICINE↗
DOE OSTI2023

Comparative transcriptome analysis of rainbow trout gonadal cells (RTG-2) infected with U and J genogroup infectious hematopoietic necrosis virus

Infectious hematopoietic necrosis virus (IHNV) is the causative pathogen of infectious hematopoietic necrosis, outbreaks of which are responsible for significant losses in rainbow trout aquaculture. Strains of IHNV isolated worldwide have been classified into five major genogroups, J, E, L, M, and U. To date, comparative transcriptomic analysis has only been conducted individually for the J and M genogroups. In this study, we compared the transcriptome profiles in U genogroup and J genogroup IHNV-infected RTG-2 cells with mock-infected RTG-2 cells. The RNA-seq results revealed 17,064 new genes, of which 7,390 genes were functionally annotated. Differentially expressed gene (DEG) analysis between U and J IHNV-infected cells revealed 2,238 DEGs, including 1,011 downregulated genes and 1,227 upregulated genes. Among the 2,238 DEGs, 345 new genes were discovered. The DEGs related to immune responses, cellular signal transduction, and viral diseases were further analyzed. RT-qPCR validation confirmed that the changes in expression of the immune response-related genes trpm2 , sting , itgb7 , ripk2 , and irf1 , cellular signal transduction-related genes irl , cacnb2 , bmp2l , gadd45α , and plk2 , and viral disease-related genes mlf1, mtor, armc5 , pik3r1 , and c-myc were consistent with the results of transcriptome analysis. Taken together, our findings provide a comprehensive transcriptional analysis of the differential virulence of the U and J genogroups of IHNV, and shed new light on the pathogenic mechanisms of IHNV strains.

Zhao, Jing-Zhuang↗
DOE OSTI2026

Compilation and utilization of a sorghum transcriptome compendium for gene regulatory network analysis and crop trait engineering

Sorghum bicolor (Sorghum) is a drought and heat tolerant C4 grass crop used to produce grain, forage, biofuels, and other bioproducts. Genetic improvement of sorghum hybrid crops is aided by a large and diverse germplasm, sorghum's diploid inbreeding genetics, and a relatively small genome that has facilitated genomic research. Over the past 20 years, the sorghum research community characterized the cytogenetic and recombinant landscapes of sorghum's 10 chromosomes, sequenced and annotated the sorghum genome, and used that information to identify genes/alleles that modulate flowering time, plant height, seed shattering, and other important traits. More recently, >1000 RNA-seq transcriptome profiles were collected from 15 sorghum genotypes to help understand the genetic basis of variation in growth and development of sorghum stems, tillers, roots, and leaves, and the regulation of biosynthetic pathways that produce epicuticular wax, dhurrin, and RFOs, compounds that contribute to sorghum's resilience. Transcriptome studies were designed to identify differentially expressed genes that are co-expressed during development or in response to a treatment to enable construction of gene regulatory networks. Co-expression and network analysis identified transcription factors and their cognate binding sites in target gene promoters and signaling pathways that modulate gene regulatory networks providing gene editing targets for further trait optimization. RNA-seq data from >20 experiments targeting sorghum organs, tissues, cell types, developmental stages, and responses to environmental conditions (i.e., diel, day-length, shading, water-deficit, temperature) has been compiled in a sorghum transcriptome compendium. The goal of this resource paper is to describe compendium content, accessibility, and a compendium data analysis pipeline and to illustrate the types of information that can be derived from the compendium with a focus on the elucidation of gene regulatory networks useful for guiding the improvement of sorghum traits through gene editing.

RNA-seq↗
DOE OSTI2022

Nonclassical autophagy activation pathways are essential for production of infectious Influenza A virus in vitro

Autophagy is a critical mechanism deployed by eukaryotic cells in response to stress, including viral infection, to boost the innate antimicrobial responses. However, an increasing number of pathogens hijack the autophagic machinery to facilitate their own replication. Influenza A virus (IAV), responsible for several global pandemics, has an intricate dependence on autophagy for successful replication in mammalian cells. To elucidate key chokepoints in the host stress responses facilitating IAV replication, we constructed a meta-transcriptome of IAV and host gene expression dynamics during early (1–3 hpi), mid (4–6 hpi), and late (8–12 hpi) stages of the viral replication cycle at two multiplicities of infection (MOI): 1 and 5. We supplemented the global transcriptome study with phosphoproteomic analysis of stress-activated protein kinase (SAPK/JNK) signaling in lung carcinoma (predominantly used as an in vitro model of IAV replication) and normal human bronchial epithelial cells. We report significant differences in the activation profiles of autophagy regulating genes upon IAV infection at the two MOI as well as divergent dependence on ULK1 signaling within the normal and cancer cells. Regardless of the cell model, JNK-Thr187 signaling was crucial for the production of infectious viral particles.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Ectomycorrhizal fungi decompose soil organic matter using oxidative mechanisms adapted from saprotrophic ancestors

Ectomycorrhizal fungi are thought to have a key role in mobilizing organic nitrogen that is trapped in soil organic matter (SOM). However, the extent to which ectomycorrhizal fungi decompose SOM and the mechanism by which they do so remain unclear, considering that they have lost many genes encoding lignocellulose-degrading enzymes that are present in their saprotrophic ancestors. Spectroscopic analyses and transcriptome profiling were used to examine the mechanisms by which five species of ectomycorrhizal fungi, representing at least four origins of symbiosis, decompose SOM extracted from forest soils. In the presence of glucose and when acquiring nitrogen, all species converted the organic matter in the SOM extract using oxidative mechanisms. The transcriptome expressed during oxidative decomposition has diverged over evolutionary time. Each species expressed a different set of transcripts encoding proteins associated with oxidation of lignocellulose by saprotrophic fungi. The decomposition ‘toolbox’ has diverged through differences in the regulation of orthologous genes, the formation of new genes by gene duplications, and the recruitment of genes from diverse but functionally similar enzyme families. The capacity to oxidize SOM appears to be common among ectomycorrhizal fungi. We propose that the ancestral decay mechanisms used primarily to obtain carbon have been adapted in symbiosis to scavenge nutrients instead.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2014

Short-term calorie restriction feminizes the mRNA profiles of drug metabolizing enzymes and transporters in livers of mice

Calorie restriction (CR) is one of the most effective anti-aging interventions in mammals. A modern theory suggests that aging results from a decline in detoxification capabilities and thus accumulation of damaged macromolecules. The present study aimed to determine how short-term CR alters mRNA profiles of genes that encode metabolism and detoxification machinery in the liver. Male C57BL/6 mice were fed CR (0, 15, 30, or 40%) diets for one month, followed by mRNA quantification of 98 xenobiotic processing genes (XPGs) in the liver, including 7 uptake transporters, 39 phase-I enzymes, 37 phase-II enzymes, 10 efflux transporters, and 5 transcription factors. In general, 15% CR did not alter mRNAs of most XPGs, whereas 30 and 40% CR altered over half of the XPGs (32 increased and 29 decreased). CR up-regulated some phase-I enzymes (fold increase), such as Cyp4a14 (12), Por (2.3), Nqo1 (1.4), Fmo2 (5.4), and Fmo3 (346), and numerous number of phase-II enzymes, such as Sult1a1 (1.2), Sult1d1 (2.0), Sult1e1 (33), Sult3a1 (2.2), Gsta4 (1.3), Gstm2 (1.3), Gstm3 (1.7), and Mgst3 (2.2). CR feminized the mRNA profiles of 32 XPGs in livers of male mice. For instance, CR decreased the male-predominantly expressed Oatp1a1 (97%) and increased the female-predominantly expressed Oatp1a4 (11). In conclusion, short-term CR alters the mRNA levels of over half of the 98 XPGs quantified in livers of male mice, and over half of these alterations appear to be due to feminization of the liver. - Highlights: • Utilized a graded CR model in male mice • The mRNA profiles of xenobiotic processing genes (XPGs) in liver were investigated. • CR up-regulates many phase-II enzymes. • CR tends to feminize the mRNA profiles of XPGs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

Entire expressed peripheral blood transcriptome in pediatric severe malarial anemia

Abstract This study on severe malarial anemia (SMA: Hb < 6.0 g/dL), a leading global cause of childhood morbidity and mortality, compares the entire expressed whole blood host transcriptome between Kenyan children (3-48 mos.) with non-SMA (Hb ≥ 6.0 g/dL, n = 39) and SMA ( n = 18). Differential expression analyses reveal 1403 up-regulated and 279 down-regulated transcripts in SMA, signifying impairments in host inflammasome activation, cell death, and innate immune and cellular stress responses. Immune cell profiling shows decreased memory responses, antigen presentation, and immediate pathogen clearance, suggesting an immature/improperly regulated immune response in SMA. Module repertoire analysis of blood-specific gene signatures identifies up-regulation of erythroid genes, enhanced neutrophil activation, and impaired inflammatory responses in SMA. Enrichment analyses converge on disruptions in cellular homeostasis and regulatory pathways for the ubiquitin-proteasome system, autophagy, and heme metabolism. Pathway analyses highlight activation in response to hypoxic conditions [Hypoxia Inducible Factor (HIF)−1 target and Reactive Oxygen Species (ROS) signaling] as a central theme in SMA. These signaling pathways are also top-ranking in protein abundance measures and a Ugandan SMA cohort with available transcriptomic data. Targeted RNA-Seq validation shows strong concordance with our entire expressed transcriptome data. These findings identify key molecular themes in SMA pathogenesis, offering potential targets for new malaria therapies.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2019

Function-driven single-cell genomics uncovers cellulose-degrading bacteria from the rare biosphere

Assigning a functional role to a microorganism has historically relied on cultivation of isolates or detection of environmental genome-based biomarkers using a posteriori knowledge of function. However, the emerging field of function-driven single-cell genomics aims to expand this paradigm by identifying and capturing individual microbes based on their in situ functions or traits. To identify and characterize yet uncultivated microbial taxa involved in cellulose degradation, we developed and benchmarked a function-driven single-cell screen, which we applied to a microbial community inhabiting the Great Boiling Spring (GBS) Geothermal Field, northwest Nevada. Our approach involved recruiting microbes to fluorescently labeled cellulose particles, and then isolating single microbe-bound particles via fluorescence-activated cell sorting. The microbial community profiles prior to sorting were determined via bulk sample 16S rRNA gene amplicon sequencing. The flow-sorted cellulose-bound microbes were subjected to whole genome amplification and shotgun sequencing, followed by phylogenetic placement. Next, putative cellulase genes were identified, expressed and tested for activity against derivatives of cellulose and xylose. Alongside typical cellulose degraders, including members of the Actinobacteria, Bacteroidetes, and Chloroflexi, we found divergent cellulases encoded in the genome of a recently described candidate phylum from the rare biosphere, Goldbacteria, and validated their cellulase activity. As this genome represents a species-level organism with novel and phylogenetically distinct cellulolytic activity, we propose the name Candidatus ‘Cellulosimonas argentiregionis’. We expect that this function-driven single-cell approach can be extended to a broad range of substrates, linking microbial taxonomy directly to in situ function.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2017

Early Transcriptomic Changes in the Ileal Pouch Provide Insight into the Molecular Pathogenesis of Pouchitis and Ulcerative Colitis

Background: Ulcerative colitis (UC) only involves the colonic mucosa. Yet, nearly 50% of patients with UC who undergo total proctocolectomy with ileal pouch anal anastomosis develop UC-like inflammation of the ileal pouch (pouchitis). By contrast, patients with familial adenomatous polyposis (FAP) with ileal pouch anal anastomosis develop pouchitis far less frequently. We hypothesized that pathogenic events associated with the development of UC are recapitulated by colonic-metaplastic transcriptomic reprogramming of the UC pouch. Methods: We prospectively sampled pouch and prepouch ileum mucosal biopsies in patients with UC with ileal pouch anal anastomosis 4, 8, and 12 months after their pouch was in continuity. Mucosal samples were also obtained from patients with FAP. Transcriptional profiles of the UC and FAP pouch and prepouch ileum were investigated via RNA sequencing and compared with data from a previously published microarray study. Results: Unlike patients with FAP, subjects with UC exhibited a large set of differentially expressed genes between the pouch and prepouch ileum as early as 4 months after pouch functionalization. Functional pathway analysis of differentially expressed genes in the UC pouch revealed an enhanced state of immune/inflammatory response and extracellular matrix remodeling. Moreover, >70% of differentially expressed genes mapped to published inflammatory bowel diseases microarray data sets displayed directional changes consistent with active UC but not with Crohn's disease. Conclusions: The UC pouch, well before histologic inflammation, already displays a systems-level gain of colon-associated genes and loss of ileum-associated genes. Patients with UC exhibit a unique transcriptomic response to ileal pouch creation that can be observed well before disease and may in part explain their susceptibility to the development of pouchitis.

familial adenomatous polyposis↗
DOE OSTI2026

Effects_of_heterologous_expression_of_glycolysis_enzymes_on_product_formation_in_Clostridium_thermocellum

Glycolysis is central toClostridium thermocellummetabolism; however, engineered strains for high ethanol titer exhibit a decrease in yield, suggesting the presence of glycolytic bottlenecks. We expressed heterologoustpi,fba, andgapDHgenes fromThermoanaerobacterium saccharolyticumandZymomonas mobilis, along with 26 non-phosphorylatinggapNvariants, to identify limiting reactions. We demonstrated functional expression and increased activity for several Fba and Tpi enzymes in the engineered strain LL1711. DespiteC. thermocellum'slow native FBA activity compared to other industrial strains, increasing Fba or Tpi activity via heterologous expression had no significant effect on cellobiose uptake or ethanol titers in high-substrate fermentations. Furthermore, 25 of 26 tested gapN genes proved highly toxic toC. thermocellumupon transformation. In conclusion, none of the tested glycolytic enzyme modifications improved fermentation profiles. These results suggest that the primary metabolic limitation is not at the FBA or TPI reactions, supporting a shift in future engineering efforts toward downstream fermentation pathways.

6-bisphosphate aldolase; glyceraldehyde-3-phosphat↗
NASA NTRSDate not supplied

Spaceflight-induced Changes in Microbial Virulence and the Impact to the Host Immune Response

INTRODUCTION Over the past 50 years, many microorganisms have displayed unexpected responses relevant to infectious disease when grown in microgravity and microgravity analogue environments, including changes in stress resistance, biofilm production, antibiotic sensitivity, final cell concentration, gene expression, enhanced host-pathogen interaction, and virulence. In parallel, astronaut studies have characterized a persistent spaceflight-induced dysregulation of the human immune system; consisting of altered leukocyte distribution, reductions in T and NK cell function, altered cytokine profiles, and reactivation of latent herpesviruses. Further, astronauts have some degree of clinical incidence, primarily infectious disease episodes and atopic dermatitis. The impact of the microgravity environment on host-pathogen interactions and potential for clinical disease remains understudied and poorly characterized. SPECIFIC AIMS In this study, the following Specific Aims are being investigated, using the microbial pathogens, Salmonella enterica Enteritidis, Pseudomonas aeruginosa, Burkholderia cepacia, Streptococcus pneumoniae, and enterohemorrhagic Escherichia coli (EHEC). Aim 1: Characterize the effect of spaceflight analogue culture on microbial pathogenesis-related stress responses and in vitro host-pathogen interactions. Analyses include microbial stress responses as well as colonization and viability following pathogen challenge of three-dimensional (3-D) tissue co-culture models containing immune cells. We specifically selected obligate and opportunistic pathogens that are medically important and have been or are likely to be found aboard spacecraft. Aim 2: Characterize the effect of spaceflight analogue culture on the virulence potential of pathogenic microorganisms. Virulence will be assessed using a mouse model of infection. SIGNIFICANCE The goal of this study is to use spaceflight analogue conditions to gain insight into the breadth of medically-significant International Space Station microorganisms that have altered virulence and the impact of those changes on the immune response of the host. This information will provide critical understanding into the impact of microgravity on potential alterations in microbial virulence and associated infectious disease risk to crew health during spaceflight missions.

C M Ott↗
DOE OSTI2012

Structural evolution and tissue-specific expression of tetrapod-specific second isoform of secretory pathway Ca{sup 2+}-ATPase

Highlights: Black-Right-Pointing-Pointer Full-length secretory pathway Ca-ATPase (SPCA2) cloned from rat duodenum. Black-Right-Pointing-Pointer ATP2C2 gene (encoding SPCA2) exists only in genomes of Tetrapoda. Black-Right-Pointing-Pointer Rat and pig SPCA2 are expressed in intestines, lung and some secretory glands. Black-Right-Pointing-Pointer Subcellular localization of SPCA2 may depend on tissue type. Black-Right-Pointing-Pointer In rat duodenum, SPCA2 is localized in plasma membrane-associated compartments. -- Abstract: Secretory pathway Ca-ATPases are less characterized mammalian calcium pumps than plasma membrane Ca-ATPases and sarco-endoplasmic reticulum Ca-ATPases. Here we report analysis of molecular evolution, alternative splicing, tissue-specific expression and subcellular localization of the second isoform of the secretory pathway Ca-ATPase (SPCA2), the product of the ATP2C2 gene. The primary structure of SPCA2 from rat duodenum deduced from full-length transcript contains 944 amino acid residues, and exhibits 65% sequence identity with known SPCA1. The rat SPCA2 sequence is also highly homologous to putative human protein KIAA0703, however, the latter seems to have an aberrant N-terminus originating from intron 2. The tissue-specificity of SPCA2 expression is different from ubiquitous SPCA1. Rat SPCA2 transcripts were detected predominantly in gastrointestinal tract, lung, trachea, lactating mammary gland, skin and preputial gland. In the newborn pig, the expression profile is very similar with one remarkable exception: porcine bulbourethral gland gave the strongest signal. Upon overexpression in cultured cells, SPCA2 shows an intracellular distribution with remarkable enrichment in Golgi. However, in vivo SPCA2 may be localized in compartments that differ among various tissues: it is intracellular in epidermis, but enriched in plasma membranes of the intestinal epithelium. Analysis of SPCA2 sequences from various vertebrate species argue that ATP2C2 gene radiated from ATP2C1 (encoding SPCA1) during adaptation of tetrapod ancestors to terrestrial habitats.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2019

Multiomics resolution of molecular events during a day in the life of Chlamydomonas

The unicellular green alga Chlamydomonas reinhardtii displays metabolic flexibility in response to a changing environment. We analyzed expression patterns of its three genomes in cells grown under light–dark cycles. Nearly 85% of transcribed genes show differential expression, with different sets of transcripts being up-regulated over the course of the day to coordinate cellular growth before undergoing cell division. Parallel measurements of select metabolites and pigments, physiological parameters, and a subset of proteins allow us to infer metabolic events and to evaluate the impact of the transcriptome on the proteome. Among the findings are the observations that Chlamydomonas exhibits lower respiratory activity at night compared with the day; multiple fermentation pathways, some oxygen-sensitive, are expressed at night in aerated cultures; we propose that the ferredoxin, FDX9, is potentially the electron donor to hydrogenases. The light stress-responsive genes PSBS , LHCSR1 , and LHCSR3 show an acute response to lights-on at dawn under abrupt dark-to-light transitions, while LHCSR3 genes also exhibit a later, second burst in expression in the middle of the day dependent on light intensity. Each response to light (acute and sustained) can be selectively activated under specific conditions. Our expression dataset, complemented with coexpression networks and metabolite profiling, should constitute an excellent resource for the algal and plant communities.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

A chloroplast diacylglycerol lipase modulates glycerolipid pathway balance in Arabidopsis

SUMMARY Two parallel pathways compartmentalized in the chloroplast and the endoplasmic reticulum contribute to thylakoid lipid synthesis in plants, but how these two pathways are coordinated during thylakoid biogenesis and remodeling remains unknown. We report here the molecular characterization of a homologous ADIPOSE TRIGLYCERIDE LIPASE‐LIKE gene, previously referred to as ATGLL . The ATGLL gene is ubiquitously expressed throughout development and rapidly upregulated in response to a wide range of environmental cues. We show that ATGLL is a chloroplast non‐regioselective lipase with a hydrolytic activity preferentially towards 16:0 of diacylglycerol (DAG). Comprehensive lipid profiling and radiotracer labeling studies revealed a negative correlation of ATGLL expression and the relative contribution of the chloroplast lipid pathway to thylakoid lipid biosynthesis. Additionally, we show that genetic manipulation of ATGLL expression resulted in changes in triacylglycerol levels in leaves. We propose that ATGLL, through affecting the level of prokaryotic DAG in the chloroplast, plays important roles in balancing the two glycerolipid pathways and in maintaining lipid homeostasis in plants.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

Indirubin-3′-(2,3 dihydroxypropyl)-oximether (E804) is a potent modulator of LPS-stimulated macrophage functions

Indirubin is a deep-red bis-indole isomer of indigo blue, both of which are biologically active ingredients in Danggui Longhui Wan, an ancient Chinese herbal tea mixture used to treat neoplasia and chronic inflammation and to enhance detoxification of xenobiotics. Multiple indirubin derivatives have been synthesized and shown to inhibit cyclin-dependent kinases (CDKs) and glycogen-synthase kinase (GSK-3β) with varying degrees of potency. Several indirubins are also aryl hydrocarbon receptor (AhR) agonists, with AhR-associated activities covering a wide range of potencies, depending on molecular structure. This study examined the effects of indirubin-3′-(2,3 dihydroxypropyl)-oximether (E804), a novel indirubin with potent STAT3 inhibitory properties, on basal and LPS-inducible activities in murine RAW264.7 macrophages. Using a focused commercial qRT-PCR array platform (SuperArray®), the effects of E804 on expression of a suite of genes associated with stress and toxicity were determined. Most genes up-regulated by LPS treatment were suppressed by E804; including LPS-induced expression of pro-inflammatory cytokines and receptors, apoptosis control genes, and oxidative stress response genes. Using qRT-PCR as a follow up to the commercial arrays, E804 treatment suppressed LPS-induced COX-2, iNOS, IL-6 and IL-10 gene expression, though the effects on iNOS and COX-2 protein expression were less dramatic. E804 also inhibited LPS-induced secretion of IL-6 and IL-10. Functional endpoints, including iNOS and lysozyme enzymatic activity, phagocytosis of fluorescent latex beads, and intracellular killing of bacteria, were also examined, and in each experimental condition E804 suppressed activities. Collectively, these results indicate that E804 is a potent modulator of pro-inflammatory profiles in LPS-treated macrophages. -- Highlights: ► RAW 264.7 macrophages were treated with 1 μM Indirubin E804, 1 μg/ml LPS, or both. ► E804 suppresses LPS-induced expression of iNOS, IL-6, COX-2, and IL-10. ► E804 suppresses LPS-induced iNOS and lysozyme activity, and IL-6 and IL-10 secretion. ► E804 suppressed phagocytosis and intracellular killing by macrophages.

60 APPLIED LIFE SCIENCES↗