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At least 343 records · Page 19
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Defining the Expression, Production, and Signaling Roles of Specialized Metabolites during Bacillus subtilis Differentiation

Bacterial specialized (or secondary) metabolites are structurally diverse molecules that mediate intra- and interspecies interactions by altering growth and cellular physiology and differentiation. Bacillus subtilis, a Gram-positive model bacterium commonly used to study biofilm formation and sporulation, has the capacity to produce more than 10 specialized metabolites. Some of these B. subtilis specialized metabolites have been investigated for their role in facilitating cellular differentiation, but only rarely has the behavior of multiple metabolites been simultaneously investigated. In this study, we explored the interconnectivity of differentiation (biofilm and sporulation) and specialized metabolites in B. subtilis. Specifically, we interrogated how development influences specialized metabolites and vice versa. Using the sporulation-inducing medium DSM, we found that the majority of the specialized metabolites examined are expressed and produced during biofilm formation and sporulation. Additionally, we found that six of these metabolites (surfactin, ComX, bacillibactin, bacilysin, subtilosin A, and plipastatin) are necessary signaling molecules for proper progression of B. subtilis differentiation. This study further supports the growing body of work demonstrating that specialized metabolites have essential physiological functions as cell-cell communication signals in bacteria.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Bacterial Homologs of Progestin and AdipoQ Receptors (PAQRs) Affect Membrane Energetics Homeostasis but Not Fluidity

Membrane potential homeostasis is essential for cell survival. Defects in membrane potential lead to pleiotropic phenotypes, consistent with the central role of membrane energetics in cell physiology. Homologs of the progestin and AdipoQ receptors (PAQRs) are conserved in multiple phyla of Bacteria and Eukarya. In eukaryotes, PAQRs are proposed to modulate membrane fluidity and fatty acid (FA) metabolism. The role of bacterial homologs has not been elucidated. Here, we use Escherichia coli and Bacillus subtilis to show that bacterial PAQR homologs, which we name “TrhA,” have a role in membrane energetics homeostasis. Using transcriptional fusions, we show that E. coli TrhA (encoded by yqfA) is part of the unsaturated fatty acid biosynthesis regulon. Fatty acid analyses and physiological assays show that a lack of TrhA in both E. coli and B. subtilis (encoded by yplQ) provokes subtle but consistent changes in membrane fatty acid profiles that do not translate to control of membrane fluidity. Instead, membrane proteomics in E. coli suggested a disrupted energy metabolism and dysregulated membrane energetics in the mutant, though it grew similarly to its parent. These changes translated into a disturbed membrane potential in the mutant relative to its parent under various growth conditions. Similar dysregulation of membrane energetics was observed in a different E. coli strain and in the distantly related B. subtilis. Together, our findings are consistent with a role for TrhA in membrane energetics homeostasis, through a mechanism that remains to be elucidated.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Optical O 2 Sensors Also Respond to Redox Active Molecules Commonly Secreted by Bacteria

From a metabolic perspective, molecular oxygen (O 2 ) is arguably the most significant constituent of Earth’s atmosphere. Nearly every facet of microbial physiology is sensitive to the presence and concentration of O 2 , which is the most favorable terminal electron acceptor used by organisms and also a dangerously reactive oxidant. As O 2 has such sweeping implications for physiology, researchers have developed diverse approaches to measure O 2 concentrations in natural and laboratory settings. Recent improvements to phosphorescent O 2 sensors piqued our interest due to the promise of optical measurement of spatiotemporal O 2 dynamics. However, we found that our preferred bacterial model, Pseudomonas aeruginosa PA14, secretes more than one molecule that quenches such sen sors, complicating O 2 measurements in PA14 cultures and biofilms. Assaying supernatants from cultures of 9 bacterial species demonstrated that this phenotype is common: all super natants quenched a soluble O 2 probe substantially. Phosphorescent O 2 probes are often embedded in solid support for protection, but an embedded probe called O 2 NS was quenched by most supernatants as well. Measurements using pure compounds indicated that quenching is due to interactions with redox-active small molecules, including phena zines and flavins. Uncharged and weakly polar molecules like pyocyanin were especially potent quenchers of O 2 NS. These findings underscore that optical O 2 measurements made in the presence of bacteria should be carefully controlled to ensure that O 2 , and not bacterial secretions, is measured, and motivate the design of custom O 2 probes for specific organisms to circumvent sensitivity to redox-active metabolites.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Structure of the Core Postfusion Porcine Endogenous Retrovirus Fusion Protein

Retroviral elements from endogenous retroviruses have functions in mammalian physiology. The best-known examples are the envelope proteins that function in placenta development and immune suppression. Porcine endogenous retroviruses (PERVs) are an understudied class of endogenous retroviruses that infect cultured human cells, raising concern regarding porcine xenografts. The PERV envelope glycoprotein has also been proposed as a possible swine syncytin with a role in placental development. Despite the growing interest in PERVs, their envelope glycoproteins remain poorly characterized. Here, we successfully determined the postfusion crystal structure of the PERV core fusion ectodomain. The PERV fusion protein structure reveals a conserved class I viral fusion protein six-helix bundle. Biophysical experiments demonstrated that the thermodynamic stability of the PERV fusion protein secondary structure was the same at physiological and acidic pHs. A conserved surface analysis highlights the high degree of sequence conservation among retroviral fusogens in the chain reversal region that facilitates the large-scale conformational change required for membrane fusion. Further structural alignment of class I viral fusogens revealed a phylogenetic clustering that shows evolution into various lineages that correlate with virus mechanisms of cell entry. Our work indicates that structural dendrograms can be used to qualitatively infer insights into the fusion mechanisms of newly discovered class I viral fusogen structures.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

A Gene Cluster That Encodes Histone Deacetylase Inhibitors Contributes to Bacterial Persistence and Antibiotic Tolerance in Burkholderia thailandensis

Persister cells are genetically identical variants in a bacterial population that have phenotypically modified their physiology to survive environmental stress. In bacterial pathogens, persisters are able to survive antibiotic treatment and reinfect patients in a frustrating cycle of chronic infection. To better define core persistence mechanisms for therapeutics development, we performed transcriptomics analyses of Burkholderia thailandensis populations enriched for persisters via three methods: flow sorting for low proton motive force, meropenem treatment, and culture aging. Although the three persister-enriched populations generally displayed divergent gene expression profiles that reflect the multimechanistic nature of stress adaptations, there were several common gene pathways activated in two or all three populations. These include polyketide and nonribosomal peptide synthesis, Clp proteases, mobile elements, enzymes involved in lipid metabolism, and ATP binding cassette (ABC) transporter systems. In particular, identification of genes that encode polyketide synthases (PKSs) and fatty acid catabolism factors indicates that generation of secondary metabolites, natural products, and complex lipids could be part of the metabolic program that governs the persistence state. We also found that loss-of-function mutations in the PKS-encoding gene locus BTH_I2366, which plays a role in biosynthesis of histone deacetylase (HDAC) inhibitors, resulted in increased sensitivity to antibiotics targeting DNA replication. Furthermore, treatment of multiple bacterial pathogens with a fatty acid synthesis inhibitor, CP-640186, potentiated the efficacy of meropenem against the persister populations. Altogether, our results suggest that bacterial persisters may exhibit an outwardly dormant physiology but maintain active metabolic processes that are required to maintain persistence.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Disrupting the ArcA Regulatory Network Amplifies the Fitness Cost of Tetracycline Resistance in Escherichia coli

There is an urgent need for strategies to discover secondary drugs to prevent or disrupt antimicrobial resistance (AMR), which is causing >700,000 deaths annually. Here, we demonstrate that tetracycline-resistant (Tet R ) Escherichia coli undergoes global transcriptional and metabolic remodeling, including downregulation of tricarboxylic acid cycle and disruption of redox homeostasis, to support consumption of the proton motive force for tetracycline efflux. Using a pooled genome-wide library of single-gene deletion strains, at least 308 genes, including four transcriptional regulators identified by our network analysis, were confirmed as essential for restoring the fitness of Tet R E. coli during treatment with tetracycline. Targeted knockout of ArcA, identified by network analysis as a master regulator of this new compensatory physiological state, significantly compromised fitness of Tet R E. coli during tetracycline treatment. A drug, sertraline, which generated a similar metabolome profile as the arcA knockout strain, also resensitized Tet R E. coli to tetracycline. We discovered that the potentiating effect of sertraline was eliminated upon knocking out arcA, demonstrating that the mechanism of potential synergy was through action of sertraline on the tetracycline-induced ArcA network in the Tet R strain. Our findings demonstrate that therapies that target mechanistic drivers of compensatory physiological states could resensitize AMR pathogens to lost antibiotics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Proteomic Analysis of Methanococcus voltae Grown in the Presence of Mineral and Nonmineral Sources of Iron and Sulfur

Iron sulfur (Fe-S) proteins are essential and ubiquitous across all domains of life, yet the mechanisms underpinning assimilation of iron (Fe) and sulfur (S) and biogenesis of Fe-S clusters are poorly understood. This is particularly true for anaerobic methanogenic archaea, which are known to employ more Fe-S proteins than other prokaryotes. Here, we utilized a deep proteomics analysis of Methanococcus voltae A3 cultured in the presence of either synthetic pyrite (FeS 2 ) or aqueous forms of ferrous iron and sulfide to elucidate physiological responses to growth on mineral or nonmineral sources of Fe and S. The liquid chromatography-mass spectrometry (LCMS) shotgun proteomics analysis included 77% of the predicted proteome. Through a comparative analysis of intra- and extracellular proteomes, candidate proteins associated with FeS 2 reductive dissolution, Fe and S acquisition, and the subsequent transport, trafficking, and storage of Fe and S were identified. The proteomic response shows a large and balanced change, suggesting that M. voltae makes physiological adjustments involving a range of biochemical processes based on the available nutrient source. Among the proteins differentially regulated were members of core methanogenesis, oxidoreductases, membrane proteins putatively involved in transport, Fe-S binding ferredoxin and radical S-adenosylmethionine proteins, ribosomal proteins, and intracellular proteins involved in Fe-S cluster assembly and storage. This work improves our understanding of ancient biogeochemical processes and can support efforts in biomining of minerals. Clusters of iron and sulfur are key components of the active sites of enzymes that facilitate microbial conversion of light or electrical energy into chemical bonds. The proteins responsible for transporting iron and sulfur into cells and assembling these elements into metal clusters are not well understood. Using a microorganism that has an unusually high demand for iron and sulfur, we conducted a global investigation of cellular proteins and how they change based on the mineral forms of iron and sulfur. Understanding this process will answer questions about life on early earth and has application in biomining and sustainable sources of energy.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Building hypotheses to understand the synergistic effects of heat and pollution exposure in a changing climate

The increasing intensity, frequency, and duration of extreme weather events due to climate change poses a broad range of health risks, and the synergistic effects of extreme temperature co-occurring with other hazardous exposures such as air pollution may result in higher risks of all-cause mortality and cardiovascular and respiratory morbidity than exposure to either event alone. There are a number of hypothesized mechanisms for this interaction effect, including increased susceptibility to heat effects on chronic conditions affected by air pollution exposure, exacerbation of pollution effects due to temperature-induced stress, and shared pathophysiological pathways (e.g., systemic inflammation), but specific physiological mechanisms are not well understood. In this editorial, the authors discuss this aspect of a recently published study examining ambient formaldehyde combined with high temperature exposure and respiratory disease admissions among children. Here, the observation that the health effects of pollutants such as formaldehyde are amplified following periods of high temperature can help inform risk warning systems even without knowledge of the underlying physiological mechanisms, but a deeper biological understanding of the interaction effects of heat and ambient air pollution may better inform interventions. This is even more important in view of increases in both extreme temperature and pollution events tied to climate change.

Chambliss, Sarah [University of Texas at Austin, T↗
DOE OSTI2020

Dynamic Construction, Perception, and Remodeling of Plant Cell Walls

Plant cell walls are dynamic structures that are synthesized by plants to provide durable coverings for the delicate cells they encase. They are made of polysaccharides, proteins, and other biomolecules and have evolved to withstand large amounts of physical force and to resist external attack by herbivores and pathogens but can in many cases expand, contract, and undergo controlled degradation and reconstruction to facilitate developmental transitions and regulate plant physiology and reproduction. Recent advances in genetics, microscopy, biochemistry, structural biology, and physical characterization methods have revealed a diverse set of mechanisms by which plant cells dynamically monitor and regulate the composition and architecture of their cell walls, but much remains to be discovered about how the nanoscale assembly of these remarkable structures underpins the majestic forms and vital ecological functions achieved by plants.

biofuels (including algae and biomass), bio-inspir↗
DOE OSTI2023

Acute stress reduces population-level metabolic and proteomic variation

Abstract Background Variation in omics data due to intrinsic biological stochasticity is often viewed as a challenging and undesirable feature of complex systems analyses. In fact, numerous statistical methods are utilized to minimize the variation among biological replicates. Results We demonstrate that the common statistics relative standard deviation (RSD) and coefficient of variation (CV), which are often used for quality control or part of a larger pipeline in omics analyses, can also be used as a metric of a physiological stress response. Using an approach we term Replicate Variation Analysis (RVA), we demonstrate that acute physiological stress leads to feature-wide canalization of CV profiles of metabolomes and proteomes across biological replicates. Canalization is the repression of variation between replicates, which increases phenotypic similarity. Multiple in-house mass spectrometry omics datasets in addition to publicly available data were analyzed to assess changes in CV profiles in plants, animals, and microorganisms. In addition, proteomics data sets were evaluated utilizing RVA to identify functionality of reduced CV proteins. Conclusions RVA provides a foundation for understanding omics level shifts that occur in response to cellular stress. This approach to data analysis helps characterize stress response and recovery, and could be deployed to detect populations under stress, monitor health status, and conduct environmental monitoring.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

De novo transcriptome in roots of switchgrass ( Panicum virgatum L. ) reveals gene expression dynamic and act network under alkaline salt stress

Background: Soil salinization is a major limiting factor for crop cultivation. Switchgrass is a perennial rhizomatous bunchgrass that is considered an ideal plant for marginal lands, including sites with saline soil. Here we investigated the physiological responses and transcriptome changes in the roots of Alamo (alkaline-tolerant genotype) and AM314/MS-155 (alkaline-sensitive genotype) under alkaline salt stress. Results: Alkaline salt stress significantly affected the membrane, osmotic adjustment and antioxidant systems in switchgrass roots, and the ASTTI values between Alamo and AM-314/MS-155 were divergent at different time points. A total of 108,319 unigenes were obtained after reassembly, including 73,636 unigenes in AM-314/MS-155 and 65,492 unigenes in Alamo. A total of 10,219 DEGs were identified, and the number of upregulated genes in Alamo was much greater than that in AM-314/MS-155 in both the early and late stages of alkaline salt stress. The DEGs in AM-314/MS-155 were mainly concentrated in the early stage, while Alamo showed greater advantages in the late stage. These DEGs were mainly enriched in plant-pathogen interactions, ubiquitin-mediated proteolysis and glycolysis/gluconeogenesis pathways. We characterized 1480 TF genes into 64 TF families, and the most abundant TF family was the C2H2 family, followed by the bZIP and bHLH families. A total of 1718 PKs were predicted, including CaMK, CDPK, MAPK and RLK. WGCNA revealed that the DEGs in the blue, brown, dark magenta and light steel blue 1 modules were associated with the physiological changes in roots of switchgrass under alkaline salt stress. The consistency between the qRT-PCR and RNA-Seq results confirmed the reliability of the RNA-seq sequencing data. A molecular regulatory network of the switchgrass response to alkaline salt stress was preliminarily constructed on the basis of transcriptional regulation and functional genes. Conclusions: Alkaline salt tolerance of switchgrass may be achieved by the regulation of ion homeostasis, transport proteins, detoxification, heat shock proteins, dehydration and sugar metabolism. These findings provide a comprehensive analysis of gene expression dynamic and act network induced by alkaline salt stress in two switchgrass genotypes and contribute to the understanding of the alkaline salt tolerance mechanism of switchgrass and the improvement of switchgrass germplasm.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Coupling of high-resolution mass spectrometer and photosynthesis system for comprehensive leaf volatile metabolite profiling

Background Leaf-level biogenic volatile organic compounds (BVOCs) emissions represent a major source of organic gases in the atmosphere, influencing both climate and air quality. These emissions are strongly driven by environmental perturbations, which affect individual plant- to ecosystem-level processes. Uncovering all the BVOCs and understanding how their emissions respond to altered environmental conditions provide critical insights into vegetation-driven changes in atmospheric chemistry. We developed a tandem instrumentation setup that integrates a proton transfer reaction time-of-flight mass spectrometer (PTR-ToF-MS) with parts-per-trillion detection limits and a photosynthetic infrared gas exchange system for the untargeted survey of all the BVOCs. This novel system enables simultaneous, real-time monitoring of BVOC emissions and photosynthetic parameters at the leaf level, offering new opportunities to disentangle the physiological and environmental drivers of VOC release. Furthermore, we established the VOC Analysis and Processing Optimization Resource (VAPOR), an open-access software tool designed for rapid data post-processing and the analysis of the variability of hundreds of BVOCs. We assessed the performance of the tandem system under varying background conditions, using standard gas mixtures and a range of environmental factors. Results Blank emissions were substantially lower for major BVOCs (e.g., isoprene) compared to those observed in plant emissions. Despite this, the observation of background-level VOCs highlights the importance of routinely acquiring and accounting for blank measurements in analyses using the coupled instrumentation. Introduction of known VOC concentrations to the system demonstrated a linear response across different compounds with varying molecular compositions, indicating minimal gas loss regardless of chemical moieties within the coupled instrumentation. We applied the optimized system to investigate the physiological mechanisms driving BVOC emissions across different genotypes of poplar and pennycress. The high mass resolution capabilities of the PTR-ToF-MS, coupled with comprehensive VAPOR-driven data analysis, enabled the identification of several important BVOCs, including methanol and methanethiol; these BVOCs displayed substantial variation across pennycress genotypes and showed concentrations ~ 100–350% higher than the blank. Moreover, isoprene emissions varied significantly among poplar genotypes grown in different potting media. Conclusions Tandem instrumentation offers a powerful tool for profiling volatile molecular markers and elucidating their genetic and environmental underpinnings. This approach enhances our ability to predict BVOC emissions in response to genotype by environmental interactions and contributes to a deeper understanding of vegetation responses to environmental changes.

Biogenic volatile organic compounds↗
DOE OSTI2024

Existing evidence on the effects of climate variability and climate change on ungulates in North America: a systematic map

Abstract Background Climate is an important driver of ungulate life-histories, population dynamics, and migratory behaviors. Climate conditions can directly impact ungulates via changes in the costs of thermoregulation and locomotion, or indirectly, via changes in habitat and forage availability, predation, and species interactions. Many studies have documented the effects of climate variability and climate change on North America’s ungulates, recording impacts to population demographics, physiology, foraging behavior, migratory patterns, and more. However, ungulate responses are not uniform and vary by species and geography. Here, we present a systematic map describing the abundance and distribution of evidence on the effects of climate variability and climate change on native ungulates in North America. Methods We searched for all evidence documenting or projecting how climate variability and climate change affect the 15 ungulate species native to the U.S., Canada, Mexico, and Greenland. We searched Web of Science, Scopus, and the websites of 62 wildlife management agencies to identify relevant academic and grey literature. We screened English-language documents for inclusion at both the title and abstract and full-text levels. Data from all articles that passed full-text review were extracted and coded in a database. We identified knowledge clusters and gaps related to the species, locations, climate variables, and outcome variables measured in the literature. Review findings We identified a total of 674 relevant articles published from 1947 until September 2020. Caribou ( Rangifer tarandus ), elk ( Cervus canadensis ), and white-tailed deer ( Odocoileus virginianus ) were the most frequently studied species. Geographically, more research has been conducted in the western U.S. and western Canada, though a notable concentration of research is also located in the Great Lakes region. Nearly 75% more articles examined the effects of precipitation on ungulates compared to temperature, with variables related to snow being the most commonly measured climate variables. Most studies examined the effects of climate on ungulate population demographics, habitat and forage, and physiology and condition, with far fewer examining the effects on disturbances, migratory behavior, and seasonal range and corridor habitat. Conclusions The effects of climate change, and its interactions with stressors such as land-use change, predation, and disease, is of increasing concern to wildlife managers. With its broad scope, this systematic map can help ungulate managers identify relevant climate impacts and prepare for future changes to the populations they manage. Decisions regarding population control measures, supplemental feeding, translocation, and the application of habitat treatments are just some of the management decisions that can be informed by an improved understanding of climate impacts. This systematic map also identified several gaps in the literature that would benefit from additional research, including climate effects on ungulate migratory patterns, on species that are relatively understudied yet known to be sensitive to changes in climate, such as pronghorn ( Antilocapra americana ) and mountain goats ( Oreamnos americanus ), and on ungulates in the eastern U.S. and Mexico.

Malpeli, Katherine C. (ORCID:000000030780918X)↗
DOE OSTI2022

Global patterns of diversity and metabolism of microbial communities in deep-sea hydrothermal vent deposits

When deep-sea hydrothermal fluids mix with cold oxygenated fluids, minerals precipitate out of solution and form hydrothermal deposits. These actively venting deep-sea hydrothermal deposits support a rich diversity of thermophilic microorganisms which are involved in a range of carbon, sulfur, nitrogen, and hydrogen metabolisms. Global patterns of thermophilic microbial diversity in deep-sea hydrothermal ecosystems have illustrated the strong connectivity between geological processes and microbial colonization, but little is known about the genomic diversity and physiological potential of these novel taxa. Here we explore this genomic diversity in 42 metagenomes from four deep-sea hydrothermal vent fields and a deep-sea volcano collected from 2004 to 2018 and document their potential implications in biogeochemical cycles. Our dataset represents 3635 metagenome-assembled genomes encompassing 511 novel and recently identified genera from deep-sea hydrothermal settings. Some of the novel bacterial (107) and archaeal genera (30) that were recently reported from the deep-sea Brothers volcano were also detected at the deep-sea hydrothermal vent fields, while 99 bacterial and 54 archaeal genera were endemic to the deep-sea Brothers volcano deposits. We report some of the first examples of medium- (≥ 50% complete, ≤ 10% contaminated) to high-quality (> 90% complete, < 5% contaminated) MAGs from phyla and families never previously identified, or poorly sampled, from deep-sea hydrothermal environments. We greatly expand the novel diversity of Thermoproteia, Patescibacteria (Candidate Phyla Radiation, CPR), and Chloroflexota found at deep-sea hydrothermal vents and identify a small sampling of two potentially novel phyla, designated JALSQH01 and JALWCF01. Metabolic pathway analysis of metagenomes provides insights into the prevalent carbon, nitrogen, sulfur, and hydrogen metabolic processes across all sites and illustrates sulfur and nitrogen metabolic “handoffs” in community interactions. We confirm that Campylobacteria and Gammaproteobacteria occupy similar ecological guilds but their prevalence in a particular site is driven by shifts in the geochemical environment. Our study of globally distributed hydrothermal vent deposits provides a significant expansion of microbial genomic diversity associated with hydrothermal vent deposits and highlights the metabolic adaptation of taxonomic guilds. Collectively, our results illustrate the importance of comparative biodiversity studies in establishing patterns of shared phylogenetic diversity and physiological ecology, while providing many targets for enrichment and cultivation of novel and endemic taxa.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Rapid remodeling of the soil lipidome in response to a drying-rewetting event

Abstract Background Microbiomes contribute to multiple ecosystem services by transforming organic matter in the soil. Extreme shifts in the environment, such as drying-rewetting cycles during drought, can impact the microbial metabolism of organic matter by altering microbial physiology and function. These physiological responses are mediated in part by lipids that are responsible for regulating interactions between cells and the environment. Despite this critical role in regulating the microbial response to stress, little is known about microbial lipids and metabolites in the soil or how they influence phenotypes that are expressed under drying-rewetting cycles. To address this knowledge gap, we conducted a soil incubation experiment to simulate soil drying during a summer drought of an arid grassland, then measured the response of the soil lipidome and metabolome during the first 3 h after wet-up. Results Reduced nutrient access during soil drying incurred a replacement of membrane phospholipids, resulting in a diminished abundance of multiple phosphorus-rich membrane lipids. The hot and dry conditions increased the prevalence of sphingolipids and lipids containing long-chain polyunsaturated fatty acids, both of which are associated with heat and osmotic stress-mitigating properties in fungi. This novel finding suggests that lipids commonly present in eukaryotes such as fungi may play a significant role in supporting community resilience displayed by arid land soil microbiomes during drought. As early as 10 min after rewetting dry soil, distinct changes were observed in several lipids that had bacterial signatures including a rapid increase in the abundance of glycerophospholipids with saturated and short fatty acid chains, prototypical of bacterial membrane lipids. Polar metabolites including disaccharides, nucleic acids, organic acids, inositols, and amino acids also increased in abundance upon rewetting. This rapid metabolic reactivation and growth after rewetting coincided with an increase in the relative abundance of firmicutes, suggesting that members of this phylum were positively impacted by rewetting. Conclusions Our study revealed specific changes in lipids and metabolites that are indicative of stress adaptation, substrate use, and cellular recovery during soil drying and subsequent rewetting. The drought-induced nutrient limitation was reflected in the lipidome and polar metabolome, both of which rapidly shifted (within hours) upon rewet. Reduced nutrient access in dry soil caused the replacement of glycerophospholipids with phosphorus-free lipids and impeded resource-expensive osmolyte accumulation. Elevated levels of ceramides and lipids with long-chain polyunsaturated fatty acids in dry soil suggest that lipids likely play an important role in the drought tolerance of microbial taxa capable of synthesizing these lipids. An increasing abundance of bacterial glycerophospholipids and triacylglycerols with fatty acids typical of bacteria and polar metabolites suggest a metabolic recovery in representative bacteria once the environmental conditions are conducive for growth. These results underscore the importance of the soil lipidome as a robust indicator of microbial community responses, especially at the short time scales of cell-environment reactions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

SAXS studies of X-ray induced disulfide bond damage: Engineering high-resolution insight from a low-resolution technique

A significant problem in biological X-ray crystallography is the radiation chemistry caused by the incident X-ray beam. This produces both global and site-specific damage. Site specific damage can misdirect the biological interpretation of the structural models produced. Cryo-cooling crystals has been successful in mitigating damage but not eliminating it altogether; however, cryo-cooling can be difficult in some cases and has also been shown to limit functionally relevant protein conformations. The doses used for X-ray crystallography are typically in the kilo-gray to mega-gray range. While disulfide bonds are among the most significantly affected species in proteins in the crystalline state at both cryogenic and higher temperatures, there is limited information on their response to low X-ray doses in solution, the details of which might inform biomedical applications of X-rays. In this work we engineered a protein that dimerizes through a susceptible disulfide bond to relate the radiation damage processes seen in cryo-cooled crystals to those closer to physiologic conditions. This approach enables a low-resolution technique, small angle X-ray scattering (SAXS), to detect and monitor a residue specific process. A dose dependent fragmentation of the engineered protein was seen that can be explained by a dimer to monomer transition through disulfide bond cleavage. This supports the crystallographically derived mechanism and demonstrates that results obtained crystallographically can be usefully extrapolated to physiologic conditions. Fragmentation was influenced by pH and the conformation of the dimer, providing information on mechanism and pointing to future routes for investigation and potential mitigation. The novel engineered protein approach to generate a large-scale change through a site-specific interaction represents a promising tool for advancing radiation damage studies under solution conditions.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Data from: "Ecophysiological variation in two provenances of Pinus flexilis seedlings across an elevation gradient from forest to alpine"

This archive contains data used to support conclusions drawn in “Ecophysiological variation in two provenances of Pinus flexilis seedlings across an elevation gradient from forest to alpine”, by Reinhardt et al., 2011. Data were collected over one summer season in plots within the Alpine Treeline Warming Experiment (ATWE), before climate manipulations began. The experiment was located on Niwot Ridge, in the Front Range of the Colorado Rocky Mountains. This data package includes five comma-separated-values (.csv) files, five Microsoft Excel (.xlsx) files, one .pdf file, and two types of geospatial files: keyhole markup language (.kml), and ESRI shapefiles (.shp). .csv files can be opened using any simple text-editing software (such as Notepad and TextEdit), R, and Microsoft Excel. .xlsx files can only be opened using Microsoft Excel. The .pdf file can be opened using Adobe Acrobat Reader or any other compatible file viewing software. The .kml file can be opened using Google Earth and Google Maps, and shapefiles can be opened using any software compatible with the file type, such as ESRI’s ArcGIS suite and QGIS.Data archived contain gas exchange and plant physiology measurements, non-structural carbohydrate data, among others. Geospatial files are also provided for additional locational context. The files and their contents in this data package are summarized under "Data Summary" in the included Data User's Guide. All files (excluding geospatial) are available in both Microsoft Excel and in .csv format, and are indicated in the Data Summary list as well.-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------Climate change is predicted to cause upward shifts in forest tree distributions, which will require seedling recruitment beyond current forest boundaries. However, predicting the likelihood of successful plant establishment beyond current species’ ranges under changing climate is complicated by the interaction of genetic and environmental controls on seedling establishment. To determine how genetics and climate may interact to affect seedling establishment, we transplanted recently germinated seedlings from high- and low-elevation provenances (HI and LO, respectively) of Pinus flexilis in common gardens arrayed along an elevation and canopy gradient from subalpine forest into the alpine zone and examined differences in physiology and morphology between provenances and among sites. Plant dry mass, projected leaf area and shoot:root ratios were 12–40% greater in LO compared with HI seedlings at each elevation. There were no significant changes in these variables among sites except for decreased dry mass of LO seedlings in the alpine site. Photosynthesis, carbon balance (photosynthesis/respiration) and conductance increased >2× with elevation for both provenances, and were 35–77% greater in LO seedlings compared with HI seedlings. There were no differences in dark-adapted chlorophyll fluorescence (Fv/Fm) among sites or between provenances. Our results suggest that for P. flexilis seedlings, provenances selected for above-ground growth may outperform those selected for stress resistance in the absence of harsh climatic conditions, even well above the species’ range limits in the alpine zone. This indicates that forest genetics may be important to understanding and managing species’ range adjustments due to climate change.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2022

Photosynthesis in aging perennial and leaf nitrogen

Perennial energy grasses are a fundamental component of the emerging bioeconomy. While their yield dynamics and environmental services are being widely studied, little attention has been given to how the physiological changes associated with perenniality (e.g., nutrient cycling) that underpin yield and environmental services, change as a function of age. Across species, perennial grasses show a photosynthetic decline as a function of stand age, however, the physiological drivers of this age-related decline are poorly understood. In this manuscript we ask: by how much does nitrogen dilution in larger older plants contribute to this photosynthetic decline? Our results support the hypothesis that nitrogen dilution did not limit photosynthesis in mature stands. Compared per unit of area-based leaf nitrogen, photosynthesis of fertilized mature stands was still 26% lower than unfertilized juvenile stands. We hypothesize that the lack of late-season carbohydrate sinks may instead drive the age-related decline in photosynthesis.

Tejera, Mauricio↗