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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 523 records · Page 29
All sourcesNASA NTRSDOE OSTI
DOE OSTI2015

The cytoskeletal inhibitors latrunculin A and blebbistatin exert antitumorigenic properties in human hepatocellular carcinoma cells by interfering with intracellular HuR trafficking

The impact of the RNA-binding protein HuR for the post-transcriptional deregulation of tumor-relevant genes is well established. Despite of elevations in HuR expression levels, an increase in cytoplasmic HuR abundance in many cases correlates with a high grade of malignancy. Here, we demonstrated that administration of the actin-depolymerizing macrolide latrunculin A, or blebbistatin, an inhibitor of myosin II ATPase activity, caused a dose- and time-dependent reduction in the high cytoplasmic HuR content of HepG2 and Huh7 hepatocellular carcinoma (HCC) cells. Subcellular fractionation revealed that in addition, both inhibitors strongly attenuated cytoskeletal and membrane-bound HuR abundance and conversely increased the HuR amount in nuclear cell fractions. Concomitant with changes in intracellular HuR localization, both cytoskeletal inhibitors markedly decreased the half-lives of cyclooxygenase-2 (COX-2), cyclin A and cyclin D{sub 1} encoding mRNAs resulting in a significant reduction in their expression levels in HepG2 cells. Importantly, a similar reduction in the expression of these HuR targets was achieved by a RNA interference (RNAi)-mediated knockdown of either HuR or nonmuscle myoin IIA. Using polysomal fractionation, we further demonstrate that the decrease in cytoplasmic HuR by latrunculin A or blebbistatin is accompanied by a marked change in the allocation of HuR and its mRNA cargo from polysomes to ribonucleoprotein (RNP) particles. Functionally, the basal migration and prostaglandin E{sub 2} synthesis are similarly impaired in inhibitor-treated and stable HuR-knockdown HepG2 cells. Our data demonstrate that interfering with the actomyosin-dependent HuR trafficking may comprise a valid therapeutic option for antagonizing pathologic posttranscriptional gene expression by HuR and furthermore emphasize the potential benefit of HuR inhibitory strategies for treatment of HCC. - Highlights: • We tested the effects of latrunculin A and blebbistatin on different HuR activities. • Both cytoskeletal inhibitors disturbed the RNP-polysome HuR allocation in HepG2 cells. • Accordingly, both inhibitors reduced the stability and levels of HuR controlled mRNAs. • Functionally, important HuR-triggered cell functions were impaired by both compounds. • Targeting of HuR trafficking implies a useful approach for novel antitumor therapy.

60 APPLIED LIFE SCIENCES↗
NASA NTRS2001

Extracellular calcium sensing and extracellular calcium signaling

The cloning of a G protein-coupled extracellular Ca(2+) (Ca(o)(2+))-sensing receptor (CaR) has elucidated the molecular basis for many of the previously recognized effects of Ca(o)(2+) on tissues that maintain systemic Ca(o)(2+) homeostasis, especially parathyroid chief cells and several cells in the kidney. The availability of the cloned CaR enabled the development of DNA and antibody probes for identifying the CaR's mRNA and protein, respectively, within these and other tissues. It also permitted the identification of human diseases resulting from inactivating or activating mutations of the CaR gene and the subsequent generation of mice with targeted disruption of the CaR gene. The characteristic alterations in parathyroid and renal function in these patients and in the mice with "knockout" of the CaR gene have provided valuable information on the CaR's physiological roles in these tissues participating in mineral ion homeostasis. Nevertheless, relatively little is known about how the CaR regulates other tissues involved in systemic Ca(o)(2+) homeostasis, particularly bone and intestine. Moreover, there is evidence that additional Ca(o)(2+) sensors may exist in bone cells that mediate some or even all of the known effects of Ca(o)(2+) on these cells. Even more remains to be learned about the CaR's function in the rapidly growing list of cells that express it but are uninvolved in systemic Ca(o)(2+) metabolism. Available data suggest that the receptor serves numerous roles outside of systemic mineral ion homeostasis, ranging from the regulation of hormonal secretion and the activities of various ion channels to the longer term control of gene expression, programmed cell death (apoptosis), and cellular proliferation. In some cases, the CaR on these "nonhomeostatic" cells responds to local changes in Ca(o)(2+) taking place within compartments of the extracellular fluid (ECF) that communicate with the outside environment (e.g., the gastrointestinal tract). In others, localized changes in Ca(o)(2+) within the ECF can originate from several mechanisms, including fluxes of calcium ions into or out of cellular or extracellular stores or across epithelium that absorb or secrete Ca(2+). In any event, the CaR and other receptors/sensors for Ca(o)(2+) and probably for other extracellular ions represent versatile regulators of numerous cellular functions and may serve as important therapeutic targets.

Review↗
DOE OSTI2017

Maternal exposure to selenium and cadmium, fetal growth, and placental expression of steroidogenic and apoptotic genes

Background: Cadmium (Cd) and selenium (Se) antagonistically influence redox balance and apoptotic signaling, with Cd potentially promoting and Se inhibiting oxidative stress and apoptosis. Alterations to placental redox and apoptotic functions by maternal exposure to Cd and Se during pregnancy may explain some of the Cd and Se associations with fetal development. Objectives: Investigate associations between Cd and Se concentrations in maternal toenails with placental expression patterns of tumor necrosis factor (TNF) and steroidogenic genes involved in redox reactions and test associations with fetal growth. Methods: In a sub-sample from the Rhode Island Child Health Study (n = 173), we investigated the relationships between: (1) maternal toenail Cd and Se concentrations and fetal growth using logistic regression, (2) Cd and Se interactions with factor scores from placental TNF and steroidogenic expression patterns (RNAseq) using linear models, and (3) TNF and steroidogenic expression factors with fetal growth via analysis of covariance. Results: Se was associated with decreased odds of intrauterine growth restriction (IUGR) (OR = 0.27, p-value = 0.045). Cd was associated with increased odds of IUGR (OR = 1.95, p-value = 0.13) and small for gestational age (SGA) births (OR = 1.46, p-value = 0.11), though not statistically significant. Cd and Se concentrations were antagonistically associated with placental TNF and steroidogenic expression patterns, which also differed by birth size. Conclusions: Se may act as an antagonist to Cd and as a modifiable protective factor in fetal growth restriction, and these data suggest these effects may be due to associated variations in the regulation of genes involved in placental redox balance and/or apoptotic signaling. - Highlights: • Tested cadmium and selenium associations with fetal growth and gene expression. • Maternal selenium associated with decreased odds of intrauterine growth restriction. • Cadmium may increase oxidative and apoptotic placental expression patterns. • Selenium may decrease oxidative and apoptotic placental expression patterns.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2015

Genome-Wide Identification of Evolutionarily Conserved Alternative Splicing Events in Flowering Plants

Alternative splicing (AS) plays important roles in many plant functions, but its conservation across the plant kingdom is not known. We describe a methodology to identify AS events and identify conserved AS events across large phylogenetic distances using RNASeq datasets. We applied this methodology to transcriptome data from nine angiosperms including Amborella, the single sister species to all other extant flowering plants. AS events within 40–70% of the expressed multi-exonic genes per species were found, 27,120 of which are conserved among two or more of the taxa studied. While many events are species specific, many others are shared across long evolutionary distances suggesting they have functional significance. Conservation of AS event data provides an estimate of the number of ancestral AS events present at each node of the tree representing the nine species studied. Furthermore, the presence or absence of AS isoforms between species with different whole genome duplication (WGD) histories provides the opportunity to examine the impact of WDG on AS potential. Examining AS in gene families identifies those with high rates of AS, and conservation can distinguish ancient events vs. recent or species specific adaptations. The MADS-box and SR protein families are found to represent families with low and high occurrences of AS, respectively, yet their AS events were likely present in the MRCA of angiosperms.

54 ENVIRONMENTAL SCIENCES↗
NASA NTRS2023

Uncovering Unique Molecular Adaptations in the Arabidopsis Thaliana Cvi-0 Ecotype

This research proposal aims to investigate the unique molecular adaptations exhibited by Arabidopsis Thaliana, specifically focusing on the Cape Verde Islands (Cvi-0) ecotype, in response to microgravity conditions. The study examines data from NASA’s Open Science Data Repository and applies a multifaceted RNAseq analysis pipeline using tools in the UseGalaxy.org open platform. Through transcriptomic analysis, differential gene expression patterns were identified in Cvi-0, revealing an absence of heat shock protein (HSP) upregulation and an upregulation of Rubisco Activase (RCA) and chloroplast-related pathways. To test the hypothesis that these adaptations may contribute to Cvi-0’s increased adaptability in microgravity, a three-fold experimental design is proposed. Four experimental groups will be cultivated under simulated microgravity and ground control conditions, including Cvi-0, Col-0, and genetically modified Col-0 with silenced HSP genes, and genetically modified Col-0 with upregulated RCA gene. Growth parameters will be measured to assess plant resilience, and RNA sequencing will provide transcriptomic data for pathway analysis. Anticipated outcomes include improved markers of plant health (mass, growth, etc.) of Cvi-0 in simulated microgravity and enhanced resilience in genetically altered Col-0 variants, providing insights into potential mechanisms of adaptation. This research would bear significance for space agriculture, nutrition for extended space missions, and sustainable terrestrial crop enhancement. Moreover, the insights gained could reshape crop engineering on Earth, enhancing robustness to climate induced stresses and bolstering global food security. The proposal’s trajectory blends scientific curiosity with practical applicability, forging a path towards sustainable food production and improving human exploration beyond our planet.

GL4HS↗
DOE OSTI2016

The role of signal transducer and activator of transcription 3 in Rift Valley fever virus infection

Rift Valley fever (RVF) is a zoonotic disease that can cause severe illness in humans and livestock, triggering spontaneous abortion in almost 100% of pregnant ruminants. In this study, we demonstrate that signal transducer and activator of transcription 3 (STAT3) is phosphorylated on its conserved tyrosine residue (Y705) following RVFV infection. This phosphorylation was dependent on a major virulence factor, the viral nonstructural protein NSs. Loss of STAT3 had little effect on viral replication, but rather resulted in cells being more susceptible to RVFV-induced cell death. Phosphorylated STAT3 translocated to the nucleus, coinciding with inhibition of fos, jun, and nr4a2 gene expression, and the presence of STAT3 and NSs at the nr4a2 promoter. NSs was found predominantly in the cytoplasm of STAT3 null cells, indicating that STAT3 influences NSs nuclear localization. Collectively, these data demonstrate that STAT3 functions in a pro-survival capacity through modulation of NSs localization. - Highlights: • STAT3 is phosphorylated on tyrosine residue 705 following RVFV infection. • Phosphorylation of STAT3 was dependent on the viral protein NSs. • STAT3 -/- MEFs were more susceptible to RVFV-induced cell death. • Loss of STAT3 led to an increase in pro-apoptotic gene expression. • STAT3 functions in a pro-survival capacity by modulation of NSs localization.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2020

Perimatrix of middle ear cholesteatoma: A granulation tissue with a specific transcriptomic signature

Objectives/Hypothesis: To establish comprehensive transcriptomic profiles of cholesteatoma perimatrix tissue and granulation tissue from chronic otitis media (COM) that did not develop cholesteatoma, which can indicate molecular pathways involved in the cholesteatoma perimatrix pathology and invasiveness. Study Design: Retrospective Case Series. Methods: Transcriptome data were obtained from cholesteatoma perimatrix tissue and COM granulation tissue by an Illumina iScan microarray. Differentially expressed genes (DEGs) were subsequently analyzed using both bioinformatical functional annotation and network analysis. Expression of candidate genes (MMP9 and LCN2) was validated by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) on a larger group of samples. Results: Analysis of the transcriptome led to the identification of 169 differentially expressed genes between investigated tissues. Bioinformatic analysis suggested that most significant biological processes involving DEGs were previously described in cholesteatoma pathology. Network analysis identified ERBB2, TFAP2A, and TP63 as major hubs of the DEGs molecular network. Furthermore, it was observed that the cellular component most significantly enriched in DEGs was extracellular space containing 47 DEGs. Using qRT-PCR, it was confirmed that mRNA levels of the major extracellular hub (MMP9) are increased, whereas its interacting molecule (LCN2) mRNA levels were decreased in cholesteatoma perimatrix tissue compared to COM granulation tissue. Conclusions: The current study approach offers an overall look at molecular mechanisms that describe the cholesteatoma entity by focusing exclusively on the perimatrix processes in comparison to COM granulation tissue. The observed differences in gene expression between cholesteatoma perimatrix and COM granulation tissue could suggest novel markers potentially influenced by the perimatrix–matrix molecular interplay, which is not present in COM without cholesteatoma. Level of Evidence: NA. Laryngoscope, 130:E220–E227, 2020. © 2019 The American Laryngological, Rhinological and Otological Society, Inc.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2015

Systematic drug safety evaluation based on public genomic expression (Connectivity Map) data: Myocardial and infectious adverse reactions as application cases

Adverse drug reaction (ADR) is of great importance to both regulatory agencies and the pharmaceutical industry. Various techniques, such as quantitative structure–activity relationship (QSAR) and animal toxicology, are widely used to identify potential risks during the preclinical stage of drug development. Despite these efforts, drugs with safety liabilities can still pass through safety checkpoints and enter the market. This situation raises the concern that conventional chemical structure analysis and phenotypic screening are not sufficient to avoid all clinical adverse events. Genomic expression data following in vitro drug treatments characterize drug actions and thus have become widely used in drug repositioning. In the present study, we explored prediction of ADRs based on the drug-induced gene-expression profiles from cultured human cells in the Connectivity Map (CMap) database. The results showed that drugs inducing comparable ADRs generally lead to similar CMap expression profiles. Based on such ADR-gene expression association, we established prediction models for various ADRs, including severe myocardial and infectious events. Drugs with FDA boxed warnings of safety liability were effectively identified. We therefore suggest that drug-induced gene expression change, in combination with effective computational methods, may provide a new dimension of information to facilitate systematic drug safety evaluation. - Highlights: • Drugs causing common toxicity lead to similar in vitro gene expression changes. • We built a model to predict drug toxicity with drug-specific expression profiles. • Drugs with FDA black box warnings were effectively identified by our model. • In vitro assay can detect severe toxicity in the early stage of drug development.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

NPRL2 reduces the niraparib sensitivity of castration-resistant prostate cancer via interacting with UBE2M and enhancing neddylation

In this study, we explored the regulatory effects of nitrogen permease regulator 2-like (NPRL2) on niraparib sensitivity, a PARP inhibitor (PARPi) in castrate-resistant prostate cancer (CRPC). Data from The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression (GTEx) program were retrospectively examined. Gene-set enrichment analysis (GSEA) was conducted between high and low NRPL2 expression prostate adenocarcinoma (PRAD) cases in TCGA. CCK-8 assay, Western blot analysis of apoptotic proteins, and flow cytometric analysis of apoptosis were applied to test niraparib sensitivity. Immunofluorescent (IF) staining and co-immunoprecipitation (co-IP) were conducted to explore the proteins interacting with NPRL2. Results showed that the upregulation of a canonical protein-coding transcript of NPRL2 (ENST00000232501.7) is associated with an unfavorable prognosis. Bioinformatic analysis predicts a physical interaction between NPRL2 and UBE2M, which is validated by a following Co-IP assay. This interaction increases NPRL2 stability by reducing polyubiquitination and proteasomal degradation. Depletion of NPRL2 or UBE2M significantly increases the niraparib sensitivity of CRPC cells and enhances niraparib-induced tumor growth inhibition in vivo. NPRL2 cooperatively enhances UBE2M-mediated neddylation and facilitates the degradation of multiple substrates of Cullin-RING E3 ubiquitin ligases (CRLs). In conclusion, this study identified a novel NPRL2-UBE2M complex in modulating neddylation and niraparib sensitivity of CRPC cells. Therefore, targeting NPRL2 might be considered as an adjuvant strategy for PARPi therapy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

Coordinated resource allocation to plant growth–defense tradeoffs

Summary Plant resource allocation patterns often reveal tradeoffs that favor growth (G) over defense (D), or vice versa. Ecologists most often explain G–D tradeoffs through principles of economic optimality, in which negative trait correlations are attributed to the reconciliation of fitness costs. Recently, researchers in molecular biology have developed ‘big data’ resources including multi‐omic (e.g. transcriptomic, proteomic and metabolomic) studies that describe the cellular processes controlling gene expression in model species. In this synthesis, we bridge ecological theory with discoveries in multi‐omics biology to better understand how selection has shaped the mechanisms of G–D tradeoffs. Multi‐omic studies reveal strategically coordinated patterns in resource allocation that are enabled by phytohormone crosstalk and transcriptional signal cascades. Coordinated resource allocation justifies the framework of optimality theory, while providing mechanistic insight into the feedbacks and control hubs that calibrate G–D tradeoff commitments. We use the existing literature to describe the coordinated resource allocation hypothesis (CoRAH) that accounts for balanced cellular controls during the expression of G–D tradeoffs, while sustaining stored resource pools to buffer the impacts of future stresses. The integrative mechanisms of the CoRAH unify the supply‐ and demand‐side perspectives of previous G–D tradeoff theories.

Monson, Russell K.↗
DOE OSTI2021

Asc-Seurat: analytical single-cell Seurat-based web application

Abstract Background Single-cell RNA sequencing (scRNA-seq) has revolutionized the study of transcriptomes, arising as a powerful tool for discovering and characterizing cell types and their developmental trajectories. However, scRNA-seq analysis is complex, requiring a continuous, iterative process to refine the data and uncover relevant biological information. A diversity of tools has been developed to address the multiple aspects of scRNA-seq data analysis. However, an easy-to-use web application capable of conducting all critical steps of scRNA-seq data analysis is still lacking. Summary We present Asc-Seurat, a feature-rich workbench, providing an user-friendly and easy-to-install web application encapsulating tools for an all-encompassing and fluid scRNA-seq data analysis. Asc-Seurat implements functions from the Seurat package for quality control, clustering, and genes differential expression. In addition, Asc-Seurat provides a pseudotime module containing dozens of models for the trajectory inference and a functional annotation module that allows recovering gene annotation and detecting gene ontology enriched terms. We showcase Asc-Seurat’s capabilities by analyzing a peripheral blood mononuclear cell dataset. Conclusions Asc-Seurat is a comprehensive workbench providing an accessible graphical interface for scRNA-seq analysis by biologists. Asc-Seurat significantly reduces the time and effort required to analyze and interpret the information in scRNA-seq datasets.

60 APPLIED LIFE SCIENCES↗
NASA NTRSDate not supplied

Bionutrients-1: Utilizing Genomics and Transcriptomics to Assess the Reliability of Microorganisms for In Situ Nutrient Production on Long Duration Missions

The resupply of current long-duration crewed missions to the ISS relies on ground-launched supplies. As NASA looks toward Mars, ground-based resupply will no longer be an option. Critical nutrients, including vitamin C, vitamin K, folate, and thiamin, degrade during long-term storage, and regular consumption of these nutrients is essential for astronaut health. Another challenge of current food systems is the difficulty of consuming sufficient calories when subsisting on the limited flavors of freeze-dried food, which can lead to weight loss. The inclusion of microorganism-based food systems could alleviate both concerns. For example, the fermentation of rehydrated milk into yogurt with microorganisms genetically incorporating genes to produce critical vitamins would allow for both in situ production of nutrients and a fresh food product with additional flavor profiles. In comparison to plant food production, microorganisms require less flight infrastructure. The BioNutrients-1 mission is demonstrating viability of microbial fermentation food production in microgravity and testing the reliability of this approach for long-duration missions lacking resupply. While the BioNutrients-1 mission includes the collection of multiple phenotypic measurements, this status update will focus on the processing of samples for genomics and transcriptomics analyses as well as the planned analysis pipelines. First, the BioNutrients-1 mission seeks to identify microorganisms capable of surviving long-duration storage at ambient temperatures while maintaining genetic fidelity. To achieve this, nine commonly employed microbial species were stored at ambient temperatures in Stasis Packs on the ISS for five years. The viability and mutation rates will be measured at multiple time points for both flown and ground control samples. From an omics perspective, the changes in the bulk rates of point mutations and genetic rearrangements across the Stasis Pack species during the five years of storage will be determined, providing valuable insights into the potential of these microorganisms for long-duration space missions. Second, the BioNutrients-1 mission is characterizing the impact of microgravity on fermentation. Two strains of the yeast Saccharomyces cerevisiae, each encoding antioxidants (β-carotene or zeaxanthin) were flown to ISS for storage and fermentation within simplified bioreactors (Production Packs). The impact of microgravity on the expression of the antioxidant production genes and general metabolic genes will be determined using RNA sequencing. Ultimately, the transcriptome data will be compared to phenotypic measurements, such as the antioxidant yield, end-state biomass, and the production of EtOH, to determine the impacts of microgravity and long-term storage on microbial fermentation. The findings from this research will be instrumental in understanding the challenges and opportunities of microorganism-based food systems in space missions.

BioNutrients↗
DOE OSTI2018

Restoration of hair-inductive activity of cultured human follicular keratinocytes by co-culturing with dermal papilla cells

Highlights: • Monocultured human ORS cells lose hair-inductive activity (trichogenicity). • The trichogenicity of human ORS cells can be restored by co-culturing with human DP cells. • Human DP cells regulate expression of a number of genes in co-cultured ORS cells. Hair follicle outer root sheath (ORS) cells can be expanded in vitro, but often lose receptivity to hair-inducing dermal signals. Recent studies have shown hair-inductive activity (trichogenicity) can be restored in rat ORS cells expanded with a fibroblast feeder by co-culturing with rat vibrissae dermal papilla (DP) cells. In this study, we investigated whether the trichogenicity of human ORS cells can be restored by co-culturing with human DP cells. ORS cells from human scalp hair follicles were cultured independently or with DP cells for 5 days and implanted into nude mice alongside freshly isolated neonatal mouse dermal cells. Although there was no hair induction when monocultured ORS cells were implanted, it was observed in co-cultured ORS cells. We also observed differential regulation of a number of genes in ORS cells co-cultured with DP cells compared to monocultured ORS cells as examined by microarray. Taken together, our data strongly suggest that human DP cells restore the trichogenicity of co-cultured ORS cells by influencing ORS gene expression through paracrine factors.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2024

A phosphorylation-deficient ribosomal protein eS6 is largely functional in Arabidopsis thaliana , rescuing mutant defects from global translation and gene expression to photosynthesis and growth

The eukaryote-specific ribosomal protein of the small subunit eS6 is phosphorylated through the target of rapamycin (TOR) kinase pathway. Although this phosphorylation event responds dynamically to environmental conditions and has been studied for over 50 years, its biochemical and physiological significance remains controversial and poorly understood. Here, we report data from Arabidopsis thaliana, which indicate that plants expressing only a phospho-deficient isoform of eS6 grow essentially normally under laboratory conditions. The eS6z (RPS6A) paralog of eS6 functionally rescued a double mutant in both rps6a and rps6b genes when expressed at approximately twice the wild-type dosage. A mutant isoform of eS6z lacking the major six phosphorylatable serine and threonine residues in its carboxyl-terminal tail also rescued the lethality, rosette growth, and polyribosome loading of the double mutant. This isoform also complemented many mutant phenotypes of rps6 that were newly characterized here, including photosynthetic efficiency, and most of the gene expression defects that were measured by transcriptomics and proteomics. However, compared with plants rescued with a phospho-enabled version of eS6z, the phospho-deficient seedlings retained a mild pointed-leaf phenotype, root growth was reduced, and certain cell cycle-related mRNAs and ribosome biogenesis proteins were misexpressed. The residual defects of the phospho-deficient seedlings could be understood as an incomplete rescue of the rps6 mutant defects. There was little or no evidence for gain-of-function defects. As previously published, the phospho-deficient eS6z also rescued the rps6a and rps6b single mutants; however, phosphorylation of the eS6y (RPS6B) paralog remained lower than predicted, further underscoring that plants can tolerate phospho-deficiency of eS6 well. Our data also yield new insights into how plants cope with mutations in essential, duplicated ribosomal protein isoforms.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2021

A novel bivalent chromatin associates with rapid induction of camalexin biosynthesis genes in response to a pathogen signal in Arabidopsis

Temporal dynamics of gene expression underpin responses to internal and environmental stimuli. In eukaryotes, regulation of gene induction includes changing chromatin states at target genes and recruiting the transcriptional machinery that includes transcription factors. As one of the most potent defense compounds in Arabidopsis thaliana , camalexin can be rapidly induced by bacterial and fungal infections. Though several transcription factors controlling camalexin biosynthesis genes have been characterized, how the rapid activation of genes in this pathway upon a pathogen signal is enabled remains unknown. By combining publicly available epigenomic data with in vivo chromatin modification mapping, we found that camalexin biosynthesis genes are marked with two epigenetic modifications with opposite effects on gene expression, trimethylation of lysine 27 of histone 3 (H3K27me3) (repression) and acetylation of lysine 18 of histone 3 (H3K18ac) (activation), to form a previously uncharacterized type of bivalent chromatin. Mutants with reduced H3K27me3 or H3K18ac suggested that both modifications were required to determine the timing of gene expression and metabolite accumulation at an early stage of the stress response. Our study indicates that the H3K27me3-H3K18ac bivalent chromatin, which we name as kairostat, plays an important role in controlling the timely induction of gene expression upon stress stimuli in plants.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Exploring drought-responsive crucial genes in Sorghum

Drought severely affects global food production. Sorghum is a typical drought-resistant model crop. Based on RNA-seq data for Sorghum with multiple time points and the gray correlation coefficient, this paper firstly selects candidate genes via mean variance test and constructs weighted gene differential co-expression networks (WGDCNs); then, based on guilt-by-rewiring principle, the WGDCNs and the hidden Markov random field model, drought-responsive crucial genes are identified for five developmental stages respectively. Enrichment and sequence alignment analysis reveal that the screened genes may play critical functional roles in drought responsiveness. A multilayer differential co-expression network for the screened genes reveals that Sorghum is very sensitive to pre-flowering drought. Furthermore, a crucial gene regulatory module is established, which regulates drought responsiveness via plant hormone signal transduction, MAPK cascades, and transcriptional regulations. The proposed method can well excavate crucial genes through RNA-seq data, which have implications in breeding of new varieties with improved drought tolerance.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

High glucose up-regulates microRNA-34a-5p to aggravate fibrosis by targeting SIRT1 in HK-2 cells

Highlights: • The expression of SIRT1 is declined and the level of miR-34a-5p is up-regulated in the kidneys of DN mice. • SIRT1 prevents TGF-β1-induced pro-fibrotic gene expression. • SIRT1 expression was declined time-dependently under high glucose in HK-2 cells. • MiR-34a-5p aggravate fibrosis through directly suppressing SIRT1. • MiR-34a-5p inhibitor reverse the expressions of downstream genes through SIRT1 in HK-2 cells. Tubulointerstitial fibrosis (TIF) is crucial in the development of renal fibrosis in diabetic nephropathy(DN). Previous data shows that SIRT1 plays an important role on fibrosis, but the effect on TIF in DN and underlying mechanisms remains uncertain. In this study, we evaluated the vital role of SIRT1 and identified SIRT1 as a downstream target gene of microRNA-34a-5p (miR-34a-5p) in TIF of DN. The result revealed that expression of miR-34a-5p, fibronectin(FN),collagen type I (COL1) and transforming growth factor β1 (TGF-β1) were up-regulated accompanied by the corresponding down-regulation of SIRT1 in renal tissues of high fat diet and streptozotocin(HFD/STZ)induced diabetic mice with DN, and that the SIRT1 mRNA level was negatively correlated with miR-34a-5p expression in high glucose stimulated human proximal tubule cell line(HK-2) cells. We then demonstrated that overexpression of SIRT1 reduced, whereas small interfering RNA targeting SIRT1 enhanced the expressions of TGF-β1 and fibrosis-related genes including FN and COL1 in HK-2 cells. Furthermore, we identified that miR-34a-5p directly suppressed SIRT1 to increase the profibrogenic effects of TGFβ1 through targeting the 3'untranslated region of SIRT1. The functional correlation of miR-34a-5p induced SIRT1 decrease was supported by overexpression and inhibition of miR-34a-5p in HK-2 cells. All the results reveal that SIRT1 which is vital in the evolution of renal TIF in DN can be directly suppressed by miR-34a-5p, and suggest that miR-34a-5p is a new target for DN treatment.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Development and use of a switchgrass ( Panicum virgatum L.) transformation pipeline by the BioEnergy Science Center to evaluate plants for reduced cell wall recalcitrance

Here, the mission of the BioEnergy Science Center (BESC) was to enable efficient lignocellulosic-based biofuel production. One BESC goal was to decrease poplar and switchgrass biomass recalcitrance to biofuel conversion while not affecting plant growth. A transformation pipeline (TP), to express transgenes or transgene fragments (constructs) in these feedstocks with the goal of understanding and decreasing recalcitrance, was considered essential for this goal. Centralized data storage for access by BESC members and later the public also was essential. A BESC committee was established to codify procedures to evaluate and accept genes into the TP. A laboratory information management system (LIMS) was organized to catalog constructs, plant lines and results from their analyses. One hundred twenty-eight constructs were accepted into the TP for expression in switchgrass in the first 5 years of BESC. Here we provide information on 53 of these constructs and the BESC TP process. Eleven of the constructs could not be cloned into an expression vector for transformation. Of the remaining constructs, 22 modified expression of the gene target. Transgenic lines representing some constructs displayed decreased recalcitrance in the field and publications describing these results are tabulated here. Transcript levels of target genes and detailed wall analyses from transgenic lines expressing six additional tabulated constructs aimed toward modifying expression of genes associated with wall structure (xyloglucan and lignin components) are provided. Altered expression of xyloglucan endotransglucosylase/hydrolases did not modify lignin content in transgenic plants. Simultaneous silencing of two hydroxycinnamoyl CoA:shikimate hydroxycinnamoyl transferases was necessary to decrease G and S lignin monomer and total lignin contents, but this reduced plant growth. A TP to produce plants with decreased recalcitrance and a LIMS for data compilation from these plants were created. While many genes accepted into the TP resulted in transgenic switchgrass without modified lignin or biomass content, a group of genes with potential to improve lignocellulosic biofuel yields was identified. Results from transgenic lines targeting xyloglucan and lignin structure provide examples of the types of information available on switchgrass lines produced within BESC. This report supplies useful information when developing coordinated, large-scale, multi-institutional reverse genetic pipelines to improve crop traits.

09 BIOMASS FUELS↗