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At least 469 records · Page 26

The molecular architecture distinctions between compression, opposite and normal wood of Pinus radiata

In gymnosperms compression wood is a specialised type of structural cell wall formed in response to biomechanical stresses. The differences in terms of gross structure, ultrastructure and chemistry are well-known. However, the differences between compression wood, normal wood, and opposite wood regarding the arrangements and interactions of the various polymers and water within their cell walls still needs to be established. The analysis of 13 C-labelled Pinus radiata by solid-state NMR spectroscopy and other complementary techniques revealed several new aspects of compression and opposite wood molecular architecture. Compared to normal wood, compression wood has a lower water content, its overall nanoporosity is reduced, and the water and matrix polymers have a lower molecular mobility. Galactan, which is a specific marker of compression wood, is broadly distributed within the cell wall, disordered, and not aligned with cellulose, and is found to be in close proximity to xylan. Dehydroabietic acid (a resin acid) is immobilised and close to the H-lignin only in compression wood. Although the overall molecular mobility of normal wood and opposite wood are similar, opposite wood has different arabinose conformations, a large increase in the amount of chain ends, contains significantly more galactan and has additional unassigned mobile components highlighting the different molecular arrangement of cell wall polymers in opposite and normal wood.

59 BASIC BIOLOGICAL SCIENCES↗

Elucidating Drought-Tolerance Mechanisms in Plant Roots through 1H NMR Metabolomics in Parallel with MALDI-MS, and NanoSIMS Imaging Techniques

As direct mediators between plants and soil, roots play an important role in metabolic responses to environmental stresses such as drought, yet these responses are vastly uncharacterized on a plant-specific level, especially for co-occurring species. Here, we aim to examine the effects of drought on root metabolic profiles and carbon allocation pathways of three tropical rainforest species by combining cutting-edge metabolomic and imaging technologies in an in situ position-specific 13C-pyruvate root-labeling experiment. Further, washed (rhizosphere-depleted) and unwashed roots were examined to test the impact of microbial presence on root metabolic pathways. Drought had a species-specific impact on the metabolic profiles and spatial distribution in Piper sp. and Hibiscus rosa sinensis roots, signifying different defense mechanisms; Piper sp. enhanced root structural defense via recalcitrant compounds including lignin, while H. rosa sinensis enhanced biochemical defense via secretion of antioxidants and fatty acids. In contrast, Clitoria fairchildiana, a legume tree, was not influenced as much by drought but rather by rhizosphere presence where carbohydrate storage was enhanced, indicating a close association with symbiotic microbes. This study demonstrates how multiple techniques can be combined to identify how plants cope with drought through different drought-tolerance strategies and the consequences of such changes on below-ground organic matter composition.

Drought, Carbon cycle, MALDI-MSI, Metabolomics, Na↗

Metabolic Interactions between Brachypodium and Pseudomonas fluorescens under Controlled Iron-Limited Conditions

Plant iron (Fe) nutritional status has a significant impact on rhizosphere microbial communities. While this effect has been attributed to alterations in the composition of root exudates, the underlying mechanisms are not known. Here, we investigated the effect of Fe deficiency on the interactions between the grass Brachypodium distachyon and the common soil bacteria Pseudomonas fluorescens SBW25. Increased phytosiderophore production was the most significant response of Brachypodium root exudation to Fe deficiency. We observed upregulated expression of phytosiderophore biosynthesis genes but lower accumulated exudate concentrations in the presence of Pseudomonas, indicating that the bacteria degrade these phytosiderophores. Pseudomonas did not produce endogenous siderophores under the low Fe conditions. Rather, the bacterial transcriptome revealed upregulation of four genes in response to Fe deficiency that contain motifs associated with cellular uptake of small molecules. Collectively, these results suggest that rhizosphere Pseudomonas fluorescens may take up phytosiderophores produced by grasses in response to Fe deficiency, and we propose target genes that may be involved. Our findings provide insight into the molecular basis for the exchange of C, N, and Fe between plants and bacteria under Fe deficient conditions. These interactions may contribute to differences in the rhizosphere microbial community structure across soil pH regimes.

Boiteau, Rene M.↗

Serendipita indica Drives Sulfur-Related Microbiota in Enhancing Growth of Hyperaccumulator Sedum alfredii and Facilitating Soil Cadmium Remediation

Endophytic fungus Serendipita indica can bolster plant growth and confer protection against various biotic and abiotic stresses. However, S. indica -reshaped rhizosphere microecology interactions and root-soil interface processes in situ at the submicrometer scale remain poorly understood. Here, we combined amplicon sequencing and high-resolution nano X-ray fluorescence (nano-XRF) imaging of the root-soil interface to reveal cadmium (Cd) rhizosphere processes. S. indica can successfully colonize the roots of Sedum alfredii Hance, which induces a remarkable increase in shoot biomass by 211.32% and Cd accumulation by 235.72%. Nano-XRF images showed that S. indica colonization altered the Cd distribution in the rhizosphere and facilitated the proximity of more Cd and sulfur (S) to enter the roots and transport to the shoot. Furthermore, the rhizosphere-enriched microbiota demonstrated a more stable network structure after the S. indica inoculation. Keystone species were strongly associated with growth promotion and Cd absorption. For example, Comamonadaceae are closely related to the organic acid cycle and S bioavailability, which could facilitate Cd and S accumulation in plants. Meanwhile, Sphingomonadaceae could release auxin and boost plant biomass. In summary, we construct a mutualism system for beneficial fungi and hyperaccumulation plants, which facilitates high-efficient remediation of Cd-contaminated soils by restructuring the rhizosphere microbiota.

60 APPLIED LIFE SCIENCES↗

Orthogonal chemical genomics approaches reveal genomic targets for increasing anaerobic chemical tolerance in Zymomonas mobilis

Genetically engineered microbes have the potential to increase efficiency in the bioeconomy by overcoming growth-limiting production stress. Screens of gene perturbation libraries against production stressors can identify high-value engineering targets, but follow-up experiments needed to guard against false positives are slow and resource-intensive. In principle, the use of orthogonal gene perturbation approaches could increase recovery of true positives over false positives because the strengths of one technique compensate for the weaknesses of the other, but, in practice, two parallel screens are rarely performed at the genome scale. Here, we screen genome-scale CRISPRi (CRISPR interference) knockdown and transposon insertion libraries of the bioenergy-relevant Alphaproteobacterium, Zymomonas mobilis, against growth inhibitors commonly found in deconstructed plant material. Integrating data from the two gene perturbation techniques, we established an approach for defining engineering targets with high specificity. This allowed us to identify all known genes in the cytochrome bc1 and cytochrome c synthesis pathway as potential targets for engineering resistance to phenolic acids under anaerobic conditions, a subset of which we validated using precise gene deletions. Strikingly, this finding is specific to the cytochrome bc1 and cytochrome c pathway and does not extend to other branches of the electron transport chain. We further show that exposure of Z. mobilis to ferulic acid causes substantial remodeling of the cell envelope proteome, as well as the downregulation of TonB-dependent transporters. Our work provides a generalizable strategy for identifying high-value engineering targets from gene perturbation screens that is broadly applicable.

CRISPRi↗

Fc-receptor-mediated phagocytosis is regulated by mechanical properties of the target

Phagocytosis is an actin-based process used by macrophages to clear particles greater than 0.5 microm in diameter. In addition to its role in immunological responses, phagocytosis is also necessary for tissue remodeling and repair. To prevent catastrophic autoimmune reactions, phagocytosis must be tightly regulated. It is commonly assumed that the recognition/selection of phagocytic targets is based solely upon receptor-ligand binding. Here we report an important new criterion, that mechanical parameters of the target can dramatically affect the efficiency of phagocytosis. When presented with particles of identical chemical properties but different rigidity, macrophages showed a strong preference to engulf rigid objects. Furthermore, phagocytosis of soft particles can be stimulated with the microinjection of constitutively active Rac1 but not RhoA, and with lysophosphatidic acid, an agent known to activate the small GTP-binding proteins of the Rho family. These data suggest a Rac1-dependent mechanosensory mechanism for phagocytosis, which probably plays an important role in a number of physiological and pathological processes from embryonic development to autoimmune diseases.

NASA Program Fundamental Space Biology↗

High-Throughput Directed Evolution of Marine Microalgae and Phototrophic Consortia for Improved Biomass Yields (Final Report)

Primary project achievements include using selective pressures (O 2 , light, temperature) and developing culturing regimes for the diatom Nitzschia inconspicua str. hildebrandi to attain enrichments with an ~90% increase in areal biomass productivity relative to the parental strain under pond-mimicking conditions with high O 2 stress in laboratory bioreactors. The resulting strain (GAI-337) was tested further for dilution time, culture density, CO 2 supplementation, pH, temperature, and dissolved O 2 concentration under outdoor pond-mimicking conditions to improve areal productivities. These experiments yielded an optimum harvest and dilution time just after sunset, ~0.45 g AFDW L -1 initial culture density for maximal productivities, no requirement for CO 2 supplementation or pH control, maximal performance under a diel temperature curve going from 24 °C at night to 36 °C during the day, and benefits from some O 2 removal from the culture by bubbling with air. Using pond-mimicking laboratory bioreactors, N. inconspicua GAI-337 achieved ~42 g AFDW m -2 d -1 . Nutrient limitation experiments resulted in a biomass composition that equated to ~160 Gallons of Gasoline Equivalent energy per ton AFDW, highlighting the potential of GAI-337 as a promising renewable fuel feedstock strain. Genome resequencing has revealed genome alterations potentially contributing to the improved growth of GAI-337 in the laboratory. Based on the comparative analyses of the GAI-337 and GAI-229 (reference) strains, we identified 144 single nucleotide substitutions that resulted in amino acid change, 7 single nucleotide substitutions that resulted in protein truncation; 5 deletions; and 1 frameshift mutation. From the mutations that potentially affect expression of functionally annotated genes, particular interest was noted for an interferon-induced 6-16 family protein that may be involved in the host immune response against microbe invasion; the chaperone protein DnaK, which may function to protect the folding of proteins within the cell; and SPRY domain protein that is found in many eukaryotic proteins important in cell signaling pathways. Transcriptome analysis revealed over 1000 genes with increased transcript levels. Many of these and many of the genes with mutations are not yet functionally annotated and an increased bioinformatics effort is necessary to more completely analyze the Nitzschia inconspicua genome. Adaptive laboratory evolution (ALE) was performed for over 300 days using consecutive 0.5°C temperature increases in a constant temperature incubator to attain greater thermal tolerance in Nitzschia inconspicua. The adapted strain was able to grow at a constant temperature of 37.5°C; whereas this constant temperature was lethal to the parental control, which had an upper temperature boundary of 35.5°C prior to adaptive evolution. Several high-temperature clonal isolates were obtained from the evolved population following ALE, and increased temperature tolerance was observed in clonal adapted cultures. The final temperature adaptation was maintained through cryopreservation and was observed in multiple clonal isolates, including multiple clonal isolates with significantly increased cell size, indicating the potential occurrence of a sexual cycle during the clonal isolation process. A survey of Nannochloropsis strains was conducted for tolerances to high pH and high bicarbonate media. Nannochloropsis granulata showed promising growth in diel bioreactors and was successfully grown at the GAI Kauai farm site in long-term growth campaigns. Co-culturing using Nitzschia inconspicua, Nannochloropsis and a cyanobacterium were assembled in the laboratory to determine if productivity synergies could be attained. Although all strains grew well in the laboratory high-bicarbonate media individually, the cyanobacterium quickly outgrew the other strains in the laboratory consortium pushing the co-culture away from a diverse (and potentially synergistic assemblage) phototroph culture towards a monoculture dominated by the cyanobacterium. Several outdoor growth campaigns were conducted, with productivities ranging between 10-20 g/m 2 /d of biomass. The best performing strain in the laboratory (GAI-337) did not outperform reference strains at the Kauai farm under the conditions used. Addition growth campaigns are necessary under conditions that result in higher biomass (>20 g/m 2 /d) and that attain higher O 2 levels are likely necessary. Initial data indicate that the thermally adapted strain did slightly better than the control strain at higher temperatures; however, additional campaigns are necessary to establish statistical significance. In summary, Nitzschia inconspicua is able to attain exemplary biomass and lipid yields in the laboratory bioreactors. Strain evolution to both O 2 and temperature resulted in targeted strain improvements. Additional outdoor campaigns are necessary to determine if laboratory improvements translate to the field.

09 BIOMASS FUELS↗

Quantitating Iron in Serum Ferritin by Use of ICP-MS

A laboratory method has been devised to enable measurement of the concentration of iron bound in ferritin from small samples of blood (serum). Derived partly from a prior method that depends on large samples of blood, this method involves the use of an inductively-coupled-plasma mass spectrometer (ICP-MS). Ferritin is a complex of iron with the protein apoferritin. Heretofore, measurements of the concentration of serum ferritin (as distinguished from direct measurements of the concentration of iron in serum ferritin) have been used to assess iron stores in humans. Low levels of serum ferritin could indicate the first stage of iron depletion. High levels of serum ferritin could indicate high levels of iron (for example, in connection with hereditary hemochromatosis an iron-overload illness that is characterized by progressive organ damage and can be fatal). However, the picture is complicated: A high level of serum ferritin could also indicate stress and/or inflammation instead of (or in addition to) iron overload, and low serum iron concentration could indicate inflammation rather than iron deficiency. Only when concentrations of both serum iron and serum ferritin increase and decrease together can the patient s iron status be assessed accurately. Hence, in enabling accurate measurement of the iron content of serum ferritin, the present method can improve the diagnosis of the patient s iron status. The prior method of measuring the concentration of iron involves the use of an atomic-absorption spectrophotometer with a graphite furnace. The present method incorporates a modified version of the sample- preparation process of the prior method. First, ferritin is isolated; more specifically, it is immobilized by immunoprecipitation with rabbit antihuman polyclonal antibody bound to agarose beads. The ferritin is then separated from other iron-containing proteins and free iron by a series of centrifugation and wash steps. Next, the ferritin is digested with nitric acid to extract its iron content. Finally, a micronebulizer is used to inject the sample of the product of the digestion into the ICPMS for analysis of its iron content. The sensitivity of the ICP-MS is high enough to enable it to characterize samples smaller than those required in the prior method (samples can be 0.15 to 0.60 mL).

Smith, Scott M.↗

Degradation science: Integrating modeling and experiments to predict localized corrosion processes (Annual Progress Report)

Additively manufactured (AM) eutectic high-entropy alloys (EHEAs), such as nano-lamellar AlCoCrFeNi 2.1 , have excellent strength, ductility, and wear resistance even at elevated temperatures, but their corrosion behavior in aggressive acids at different length scales remain poorly understood. This work investigates the corrosion behavior of laser powder bed–fused (L-PBF) AlCoCrFeNi 2.1 as a function of annealing temperatures, probing degradation mechanisms from nanoscopic to macroscopic length scales. The alloy is dual phase consisting of a ductile FCC L1 2 phase and a high-strength BCC B2 phase. Rapid solidification during L-PBF produces a far-from-equilibrium nano-lamellar structure with nearly homogeneous elemental distribution, which tends to evolve upon annealing toward Cr/Co/Fe-enriched FCC and Al/Ni-enriched B2. Three conditions were studied: as-printed, 600 °C/5 h, and 1000 °C/1 h, over which B2 lamellae coarsen, lamellar spacing increases, and elemental segregation becomes more prominent. Microstructure and chemistry were characterized by scanning electron microscopy (SEM) and energy-dispersive spectroscopy (EDS), while in-situ electrochemical atomic force microscopy (EC-AFM) was used to link early (<5 h) local dissolution to microstructure after exposure in sulfuric acid. EC-AFM highlights preferential dissolution of the BCC/B2 phase where the surrounding matrix is Cr-depleted and directly quantifies the dissolution rates within individual phases, tracks the transition from early nano-scale attack to partial repassivation, to correlate height differences with current and impedance responses. To monitor longer-term behavior (up to 96 h), ex-situ AFM, SEM, and confocal imaging were combined with conventional bulk electrochemical tests, bridging nanoscale observations to micro/meso-scale damage morphologies. At the meso-scale, the deepest dissolution channels align with the build-direction lamellae and melt-pool boundaries, indicating that printing directionality guides the propagation of these localized corrosion sites. Annealing modifies corrosion by restructuring BCC/FCC phase fractions, lamellar spacing, and Cr/Al segregation, thereby changing the cathode/anode ratio and passive film stability. The results clarify how as-printed and annealed nano-lamellar architectures differ in their susceptibility to selective dissolution; how elemental segregation competes with residual stresses along the build direction. With these insights, future work will use CALPHAD-guided alloy modification to stabilize higher Cr contents in the B2 phase while retaining a dominant FCC+B2/BCC microstructure, with the goal of designing mechanically robust, corrosion-resistant EHEAs for safety-critical applications to leverage the LLNL’s broader national and global security mission.

36 MATERIALS SCIENCE↗

Pangenomes suggest ecological-evolutionary responses to experimental soil warming

ABSTRACT Below-ground carbon transformations that contribute to healthy soils represent a natural climate change mitigation, but newly acquired traits adaptive to climate stress may alter microbial feedback mechanisms. To better define microbial evolutionary responses to long-term climate warming, we study microorganisms from an ongoing in situ soil warming experiment where, for over three decades, temperate forest soils are continuously heated at 5°C above ambient. We hypothesize that across generations of chronic warming, genomic signatures within diverse bacterial lineages reflect adaptations related to growth and carbon utilization. From our bacterial culture collection isolated from experimental heated and control plots, we sequenced genomes representing dominant taxa sensitive to warming, including lineages of Actinobacteria, Alphaproteobacteria, and Betaproteobacteria. We investigated genomic attributes and functional gene content to identify signatures of adaptation. Comparative pangenomics revealed accessory gene clusters related to central metabolism, competition, and carbon substrate degradation, with few functional annotations explicitly associated with long-term warming. Trends in functional gene patterns suggest genomes from heated plots were relatively enriched in central carbohydrate and nitrogen metabolism pathways, while genomes from control plots were relatively enriched in amino acid and fatty acid metabolism pathways. We observed that genomes from heated plots had less codon bias, suggesting potential adaptive traits related to growth or growth efficiency. Codon usage bias varied for organisms with similar 16S rrn operon copy number, suggesting that these organisms experience different selective pressures on growth efficiency. Our work suggests the emergence of lineage-specific trends as well as common ecological-evolutionary microbial responses to climate change. IMPORTANCE Anthropogenic climate change threatens soil ecosystem health in part by altering below-ground carbon cycling carried out by microbes. Microbial evolutionary responses are often overshadowed by community-level ecological responses, but adaptive responses represent potential changes in traits and functional potential that may alter ecosystem function. We predict that microbes are adapting to climate change stressors like soil warming. To test this, we analyzed the genomes of bacteria from a soil warming experiment where soil plots have been experimentally heated 5°C above ambient for over 30 years. While genomic attributes were unchanged by long-term warming, we observed trends in functional gene content related to carbon and nitrogen usage and genomic indicators of growth efficiency. These responses may represent new parameters in how soil ecosystems feedback to the climate system.

Choudoir, Mallory J. (ORCID:0000000291175150)↗

Hyaloscypha finlandica Metabolome Repository

This repository provides the curated data tables, manuscript figure and table exports, dependency records, and workflow scripts supporting an integrated comparative genomics and untargeted LC-MS/MS metabolomics analysis of Hyaloscypha finlandica strain PMI 746, a root-associated dark septate endophyte of poplar. The repository includes genome-mining summaries from antiSMASH, FunBGCeX, BGC-Prophet, and BiG-SCAPE; processed metabolomics inputs; metabolite annotation evidence; statistical outputs; and publication-facing figures and tables. Raw LC-MS/MS spectra, full genome/protein downloads, and large generated tool outputs are referenced through public archive/accession records and are not stored in Git.

59 BASIC BIOLOGICAL SCIENCES↗

Thermal Conductivity of Ultem(TradeMark)/Carbon Nanofiller Blends

In an effort to improve polymer thermal conductivity (TC), Ultem(TradeMark) 1000 was compounded with nano-fillers of carbon allotropes. Ultem(TradeMark) 1000 was selected since it is both solution and melt processable. As-received and modified multiwalled carbon nanotubes (MWCNTs), vapor grown carbon nanofibers (CNF) and expanded graphite (EG) were investigated. MWCNTs were modified by functionalizing the surface through oxidization with concentrated acids, mixing with an alkyl bromide, and addition of alkyl and phosphorus compounds after initial treatment with n-butyl lithium. Functionalization was performed to improve the TC compatibility between the resin and MWCNTs. It was postulated that this may provide an improved interface between the MWCNT and the polymer which would result in enhanced TC. The nano-fillers were mixed with Ultem(TradeMark) 1000 in the melt and in solution at concentrations ranging from 5 to 40 wt%. Ribbons were extruded from the blends to form samples where the nano-fillers were aligned to some degree in the extrusion direction. Samples were also fabricated by compression molding resulting in random orientation of the nano-fillers. Thermal properties of the samples were evaluated by Differential Scanning Calorimetry (DSC) and Thermal Gravimetric Analyzer (TGA). Tensile properties of aligned samples were determined at room temperature. The specimens were cut from the ribbons in the extrusion direction; hence the nano-fillers are somewhat aligned in the direction of stress. Typically it was observed that melt mixed samples exhibited superior mechanical properties compared to solution mixed samples. As expected, increased filler loading led to increased modulus and decreased elongation with respect to the neat polymer. The degree of dispersion and alignment of the nano-fillers was determined by high-resolution scanning electron microscopy (HRSEM). HRSEM of the ribbons revealed that the MWCNTs and CNFs were predominantly aligned in the flow direction. The TC of the samples was measured using a Nanoflash(TradeMark) instrument. Since the MWCNTs and CNF are anisotropic, the TC was expected to be different in the longitudinal (parallel to the nanotube and fiber axis) and transverse (perpendicular to the nanotube and fiber axis) directions. The extruded ribbons provided samples for transverse TC measurements. However, to determine the TC in the longitudinal direction, the ribbons needed to be stacked and molded under 1.7 MPa and 270 C. Samples were then obtained by cutting the molded block with a diamond saw. The largest TC improvement was achieved for aligned samples when the measurement was performed in the direction of MWCNT and CNF alignment (i.e. longitudinal axis). Unaligned samples also showed a significant improvement in TC and may be potentially useful in applications when it is not possible to align the nano-filler. The results of this study will be presented.

Ghose, S.↗

Reformulation of dicamba herbicide: Impacts on offsite transport and soybean damage

Abstract The herbicide dicamba (3,6‐dichloro‐2‐methoxybenzoic acid) is commonly used to control broadleaf weeds in soybeans. Dicamba, however, is susceptible to volatilization and drift, thereby causing significant plant damage to nontarget crops downwind. Dicamba was reformulated to reduce volatility and off‐target movement. The effectiveness of the dicamba reformulation was assessed by quantifying dicamba emissions following spray application and investigated how meteorological factors influenced the off‐target movement. The experiments were conducted at the University of Minnesota Agricultural Experiment Station (UMORE Park) during the growing season of 2018, 2019, 2021, and 2022. Multiple high‐flow polyurethane foam air samplers were used to measure dicamba concentrations downwind from a 4‐ha soybean field sprayed with dicamba. Dicamba emissions were estimated using backward Lagrangian modeling constrained by the air sample observations. The results indicate that dicamba emissions and downwind transport were significant for several days following application. Further, non‐traited soybeans located within 15–45 m showed substantial dicamba‐related damage. In warmer, drier seasons, increased dicamba emissions caused more severe damage to downwind soybeans, likely worsened by drought stress preventing recovery. Favorable atmospheric conditions that reduced potential drift can be difficult to achieve in terms of the typical weather experienced over agricultural sites in the Upper Midwest. These results indicate that the dicamba reformulation has not adequately prevented significant post‐spray volatilization losses and downwind transport.

Agriculture↗

Building Muscles, Keeping Muscles: Protein Turnover During Space Flight

As we age we lose muscle mass and strength. The problem is a matter of use it or lose it and more - a fact to which any active senior can attest. An imbalance in the natural cycle of protein turnover may be a contributing factor to decreased muscle mass. But the answer is not so simple, since aging is associated with changes in hormones, activity levels, nutrition, and often, disease. The human body constantly uses amino acids to build muscle protein, which then breaks down and must be replaced. When protein turnover gets out of balance, so that more protein breaks down than the body can replace, the result is muscle loss. This is not just the bane of aging, however. Severely burned people may have difficulty building new muscle long after the burned skin has been repaired. Answers to why we lose muscle mass and strength - and how doctors can fix it - may come from space. Astronauts usually eat a well-balanced diet and maintain an exercise routine to stay in top health. During long-duration flight, they exercise regularly to reduce the muscle loss that results from being in a near-weightless environment. Despite these precautions, astronauts lose muscle mass and strength during most missions. They quickly recover after returning to Earth - this is a temporary condition in an otherwise healthy population. Members of the STS-107 crew are participating in a study of the effects of space flight, hormone levels, and stress on protein turnover. When we are under stress, the body responds with a change in hormone levels. Researchers hypothesize that this stress-induced change in hormones along with the near-weightlessness might result in the body synthesizing less muscle protein, causing muscles to lose their strength and size. Astronauts, who must perform numerous duties in a confined and unusual environment, experience some stress during their flight, making them excellent candidates for testing the researchers' hypothesis.

Arny Ferrando↗

A Systems Approach to Increasing Carbon Flux to Seed Oil for Biofuels and Bioproducts Production in Camelina sativa (Final Report)

To combat climate change and alleviate the dependency of the United States on fossil fuels, the transition to biofuel crops has long been proposed as a crucial part of the long-term solution. Camelina sativa has emerged as one of the leading commercially viable options for biofuel and bioproduct production for the U.S. Camelina has the advantages of low agronomic inputs and natural resistance to diverse biotic and abiotic stresses relative to other oilseed crops, and Camelina oil-based blends have been tested and approved as liquid transportation fuels. A major limitation in the widespread adoption of Camelina as a viable industrial oilseed crop is its modest oil yields. This project directly investigated possible paths towards increasing oil, by employing tissue-specific and whole plant systems approaches to identify major regulatory bottlenecks. We developed a new high-throughput method for the identification of multi-gene transformants in polyploid species like Camelina sativa and used this to determine that, contrary to what mathematical models suggested, introduction of the microbial Entner-Doudoroff (ED) pathway into Camelina sativa did not result in significant seed oil increases. We developed flux maps providing accurate and statistically robust models of central carbon metabolism in Camelina, including of cultured Camelina embryos. This knowledge was used in the capacitation of a number of junior researchers in gaining knowledge on approaches used for quantitative flux map analyses. We identified several novel Camelina transcription factor genes regulating fatty acid biosynthesis that showed variable effects on seed oil accumulation in transgenic plants. Among a number of community resources, we also developed a knowledge base web resource (CamRegBase) to integrate Camelina gene regulatory information. Together, research outcomes from this project have contributed to a much better understanding of the regulation and bottlenecks to engineer seed oil production in Camelina.

09 BIOMASS FUELS↗

Combined Effects of Simulated Microgravity and Radiation Exposure on Osteoclast Cell Fusion

The loss of bone mass and alteration in bone physiology during space flight are one of the major health risks for astronauts. Although the lack of weight bearing in microgravity is considered a risk factor for bone loss and possible osteoporosis, organisms living in space are also exposed to cosmic radiation and other environmental stress factors. As such, it is still unclear as to whether and by how much radiation exposure contributes to bone loss during space travel, and whether the effects of microgravity and radiation exposure are additive or synergistic. Bone is continuously renewed through the resorption of old bone by osteoclast cells and the formation of new bone by osteoblast cells. In this study, we investigated the combined effects of microgravity and radiation by evaluating the maturation of a hematopoietic cell line to mature osteoclasts. RAW 264.7 monocyte/macrophage cells were cultured in rotating wall vessels that simulate microgravity on the ground. Cells under static 1g or simulated microgravity were exposed to γ rays of varying doses, and then cultured in receptor activator of nuclear factor-κB ligand (RANKL) for the formation of osteoclast giant multinucleated cells (GMCs) and for gene expression analysis. Results of the study showed that radiation alone at doses as low as 0.1 Gy may stimulate osteoclast cell fusion as assessed by GMCs and the expression of signature genes such as tartrate resistant acid phosphatase (Trap) and dendritic cell-specific transmembrane protein (Dcstamp). However, osteoclast cell fusion decreased for doses greater than 0.5 Gy. In comparison to radiation exposure, simulated microgravity induced higher levels of cell fusion, and the effects of these two environmental factors appeared additive. Interestingly, the microgravity effect on osteoclast stimulatory transmembrane protein (Ocstamp) and Dcstamp expressions was significantly higher than the radiation effect, suggesting that radiation may not increase the synthesis of adhesion molecules as much as microgravity.

Zhang, Ye↗

Highly durable fuel cell electrocatalyst with low-loading Pt-Co nanoparticles dispersed over single-atom Pt-Co-N-Graphene nanofiber

The limited durability of Pt electrocatalysis toward cathodic oxygen reduction reaction remains challenging, yet crucial for the development of Proton Exchange Membrane Fuel Cell. Here, we present a rational design of a robust catalyst consisting of PtCo nanoparticles supported on Pt-Co-N-graphene nanofiber prepared through electrospun Cobalt-Metal-Organic-Framework. The catalyst delivers unprecedented mass activity of 2.48 A·mgPt -1 , and retains 80% of initial value after 60,000 Accelerated-Stress-Test cycles. Operando X-ray absorption spectroscopies show that the electronic configurations of Pt sites in PtCo and Co sites in Co-N4 in the hybrid catalyst are modified toward high catalytic activities. Density Functional Theory unveils that the enhanced curvature of the substrate induced by the morphology engineering lowers the reaction thermodynamic barrier on Co-N4 sites, favoring the formation of H2O and suppressing that of H2O2. This result along with the strong affinity of PtCo nanoparticles to the Pt-Co-N-graphene fiber endows the catalyst an exceptional durability.

acidic electrolyte↗