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Abstract Background Se (selenium) pollution is an emerging environmental concern. Excessive Se induces phytotoxicity. The endogenous H2S (hydrogen sulfide) was involved in plant adaptation to Se stress, but the signaling player of H2S remains unclear. Methods The study was conducted in a hydroponic system with different chemicals added to the treatment solution. Fluorescent tracking was performed to detect endogenous signaling molecules in plant tissues. Physiological changes were determined based on pharmaceutics and histochemical experiments. Gene expression was analyzed using qRT-PCR. The data were summarized using hierarchical cluster and Pearson correlation analysis. Results Se stress inhibited B. rapa growth (e.g. root elongation, shoot height, and seedling fresh weight and dry weight) in both dose- and time-dependent manners, with approximately 50% of root growth inhibition occurred at 20 µM Se. Se stress induced ROS (reactive oxygen species) accumulation and oxidative injury in B. rapa. Se exposure resulted in the upregulation of LCDs (L-cysteine desulfhydrase) and DCDs (D-cysteine desulfhydrase) encoding enzymes for H2S production in B. rapa at early stage of Se exposure, followed by downregulation of these genes at late stage. This was consistent with the change of endogenous H2S in B. rapa. Enhancing endogenous H2S level with NaHS (H2S donor) stimulates endogenous Ca2+ in B. rapa upon Se exposure, accompanied the attenuation of growth inhibition, ROS accumulation, oxidative injury, and cell death. The beneficial effects of H2S on detoxifying Se were blocked by decreasing endogenous Ca2+ level with Ca2+ channel blocker or Ca2+ chelator. Finally, hierarchical cluster combined with correlation analysis revealed that Ca2+ might acted as downstream of H2S to confer Se tolerance in B. rapa. Conclusion Ca2+ was an important player of H2S in the regulation of plant physiological response upon Se stress. Such findings extend our knowledge of the mechanism for Se-induced phytotoxicity. Graphical Abstract

Developing biostimulants is a promising approach for sustainable agriculture under a saline environment. Thymol is a plant-derived compound with a potential antioxidative capacity. However, little is known about whether and how the antioxidative property of thymol plays a role in inducing plant tolerance against abiotic stresses. Here, we find that thymol induces saline tolerance in Chinese cabbage seedlings via enhancing the antioxidative capacity. Treatment with NaCl (100 mM) decreased the seedling fresh weight by 59.9% as compared to a control. Thymol at 20 μM showed the greatest effect on promoting seedling growth under saline stress, with the seedling fresh weight being increased by 71.0% as compared to NaCl treatment. Thymol remarkably decreased the overaccumulation of ROS (hydrogen peroxide and a superoxide radical); cell membrane damage (evaluated by lipid oxidation, membrane integrity, and relative conductivity); and cell death in seedlings under saline stress. Thymol induced three antioxidative systems to lower the ROS level in salt-treated seedlings. First, thymol remarkably activated a set of antioxidative enzymes, such as SOD (superoxide dismutase), APX (ascorbate peroxidase), CAT (catalase), and POD (peroxidase). Second, thymol balanced the cellular redox status by increasing the ratio of AsA/DHA (ascorbic acid/dehydroascorbic acid) and GSH/GSSG (glutathione/oxidized glutathione). Third, thymol significantly enhanced the level-two kinds of antioxidants (total phenol and flavonoid). All of these physiological responses were observed in both the shoots and the roots. In sum, thymol deploys multiple antioxidative systems to help Chinese cabbage seedlings against saline stress. Such findings suggest that thymol has great potential to be developed as a novel biostimulant enhancing crop tolerance against saline stress.

Background: Small-diameter (

An effective HIV vaccine likely requires the elicitation of neutralizing antibodies (NAbs) against multiple HIV-1 clades. The recently developed cleavage-independent native flexibly linked (NFL) envelope (Env) trimers exhibit well-ordered conformation and elicit autologous tier 2 NAbs in multiple animal models. Here, we investigated whether the fusion of molecular adjuvant C3d to the Env trimers can improve B- cell germinal center (GC) formation and antibody responses. To generate Env-C3d trimers, we performed a glycine-serine- based (G4S) flexible peptide linker screening and identified a linker range that allowed native folding. A 30–60- amino- acid- long linker facilitates Env-to-C3d association and achieves the secretion of well-ordered trimers and the structural integrity and functional integrity of Env and C3d. The fusion of C3d did not dramatically affect the antigenicity of the Env trimers and enhanced the ability of the Env trimers to engage and activate B cells in vitro. In mice, the fusion of C3d enhanced germinal center formation, the magnitude of Env-specific binding antibodies, and the avidity of the antibodies in the presence of an adjuvant. The Sigma Adjuvant System (SAS) did not affect the trimer integrity in vitro but contributed to altered immunogenicity in vivo, resulting in increased tier 1 neutralization, likely by increased exposure of variable region 3 (V3). Taken together, the results indicate that the fusion of the molecular adjuvant, C3d, to the Env trimers improves antibody responses and could be useful for Env-based vaccines against HIV.

Abstract Background Although marginal donor livers expand the donor pool, an ideal method for quantitatively evaluating the quality of donor livers has not been developed. This study aimed to explore the feasibility of indocyanine green (ICG) fluorescence imaging for estimating liver function in an ischemia–reperfusion model. Methods Forty-eight rats were randomly and evenly divided into 8 groups: the control group and the experimental groups (I-VII). The portal vein blocking period was 0 min, 10 min, 20 min, 30 min, 40 min, 50 min and 60 min. After blood flow was reestablished and the hemodynamics stabilized, ICG was injected through the dorsal penile vein as a bolus, and the fluorescence signal was recorded for 30 min in real time. The fluorescence intensity (FI) curve of the liver was fitted with an asymptotic regression model. Fresh liver tissues and serum were obtained from the middle lobe of the liver on postoperative day (POD) 1 and POD 7 for histopathological evaluation and liver function tests. Results The growth rate of the FI curve, parameter b3, decreased from groups I to VII. According to the two sudden changes in b3 (20 min, 50 min), the experimental groups could be classified into 3 groups (A, B and C). Hepatocytes in groups I-II showed slight edema, group III began to show obvious hepatocyte edema and vacuolar degeneration, and in groups VI-VII, severe hepatocyte degeneration, necrosis and large inflammatory cell infiltration were observed. Suzuki’s scores in the 3 groups were also significantly different (P

Gray mold infected with Botrytis cinerea frequently appears on fruits and vegetables throughout the supply chain after harvest, leading to economic losses. Biological control of postharvest disease with phytochemicals is a promising approach. CA (cinnamaldehyde) is a natural phytochemical with medicinal and antimicrobial activity. This study evaluated the effect of CA in controlling B. cinerea on fresh pepper fruit. CA inhibited B. cinerea growth in vitro significantly in a dose- (0.1–0.8 mM) and time-dependent (6–48 h) manner, with an EC50 (median effective concentration) of 0.5 mM. CA induced the collapse and breakdown of the mycelia. CA induced lipid peroxidation resulting from ROS (reactive oxygen species) accumulation in mycelia, further leading to cell leakage, evidenced by increased conductivity in mycelia. CA induced mycelial glycerol accumulation, resulting in osmotic stress possibly. CA inhibited sporulation and spore germination resulting from ROS accumulation and cell death observed in spores. Spraying CA at 0.5 mM induced a defense response in fresh pepper fruits, such as the accumulation of defense metabolites (flavonoid and total phenols) and an increase in the activity of defense enzymes (PAL, phenylalanine ammonia lyase; PPO, polyphenol oxidase; POD, peroxidase). As CA is a type of environmentally friendly compound, this study provides significant data on the activity of CA in the biocontrol of postharvest gray mold in peppers.

Selenium (Se) is involved in the growth and development of plants. More importantly, Se from plant foods is a primary source of Se intake for humans and animals. Improving the Se content in vegetables through biofortification is an effective way to solve the hidden hunger induced by Se deficiency. This study demonstrated the effect of different exogenous Se application concentrations on the growth, antioxidative capacity, and nutritional quality of purple lettuce (Lactuca sativa var. crispa L. “Purple Rome”) at the seedling stage. The low Se application concentration (≤8 μM) significantly promoted the lettuce seedling growth. Conversely, the high Se application concentration (16 μM) inhibited the seedling growth and overproduced the reactive oxygen species in lettuce root tips, which caused oxidative damage to membrane lipids and cell death. Furthermore, the enzyme activities and gene expression of the antioxidant enzymes, superoxide dismutase-peroxidase, and catalase, were significantly increased under exogenous Se application. The exogenous Se application significantly increased the accumulation of nutrients in purple lettuce at the seedling stage. Remarkably, the exogenous Se application concentrations were significantly positively related to the Se and anthocyanin contents. The gene expression levels of chalcone synthase were positively correlated with the anthocyanin contents under exogenous Se application. This study contributes to the role of Se in lettuce growth and provides a reference for producing high-quality purple lettuce rich in Se and anthocyanins.

Abstract Background Calmodulin1 (CALM1) has been identified as one of the overexpression genes in a variety of cancers and EGFR inhibitor have been widely used in clinical treatment but it is unknown whether CALM1 and epidermal growth factor receptor (EGFR) have a synergistic effect in esophageal squamous cell carcinoma (ESCC). The aim of the present study was to explore the synergistic effects of knock-out CALM1 combined with EGFR inhibitor (Afatinib) and to elucidate the role of CALM1 in sensitizing the resistance to Afatinib in ESCC. Method Immunohistochemistry (IHC) and qRT-PCR were used to examine the expression of CALM1 and EGFR in ESCC tissues. Kaplan–Meier survival analysis was used to analyze the clinical and prognostic significance of CALM1 and EGFR expression in ESCC. Furthermore, to evaluate the biological function of CALM1 in ESCC, the latest gene editing technique CRISPR/Cas9(Clustered regularly interspaced short palindromic repeats)was applied to knockout CALM1 in ESCC cell lines KYSE150, Eca109 and TE-1. MTT, flow cytometry, Transwell migration, scratch wound-healing and colony formation assays were performed to assay the combined effect of knock-out CALM1 and EGFR inhibitor on ESCC cell proliferation and migration. In addition, nude mice xenograft model was used to observe the synergistic inhibition of knock-out CALM1 and Afatinib. Results Both CALM1 and EGFR were found to be significantly over-expressed in ESCC compared with paired normal control. Over-expressed CALM1 and EGFR were significantly associated with clinical stage, T classification and poor overall prognosis, respectively. In vitro, the combined effect of knock-out CALM1 mediated by the lentivirus and EGFR inhibitor was shown to be capable of inhibiting the proliferation, inducing cell cycle arrest at G1/S stage and increasing apoptosis of KYSE-150 and Eca109 cells; invasion and migration were also suppressed. In vivo, the results of tumor weight and total fluorescence were markedly reduced compared with the sgCtrl-infected group and sgCAML1 group. Conclusion Our data demonstrated that knock-out of CALM1 could sensitize ESCC cells to EGFR inhibitor, and it may exert oncogenic role via promotion of EMT. Taken together, CALM1 may be a tempting target to overcome Afatinib resistance.

Fusarium oxysporum f. sp. niveum (FON) causes watermelon wilt that is one of the major disease-causing yield losses of watermelon. Sustainable development of agriculture requires controlling watermelon wilt disease with good environmental performance. One important approach is to identify environmental-friendly compounds with inhibitory activity against FON. Thymol is a plant-derived compound that is safe for ecology. Little is known about the application of thymol in agriculture. In this study, we studied the inhibitory activity of thymol against FON by using morphological, physiological, and histochemical approaches. Thymol significantly inhibited colony diameter of FON in a dose-dependent manner, with EC50 at 21 µg/mL. Thymol at 10, 21, and 35 µg/mL decreased the fresh weight of FON mycelia by 29.0%, 50.6%, and 69.5%, respectively. Microscopic observation revealed irregular damage and loss of shape of mycelia upon thymol exposure. Thymol induced the accumulation of superoxide radical in mycelial cells and accompanied increased activity of antioxidative enzymes (SOD, superoxide dismutase; CAT, catalase). Thymol induced membrane permeability was indicated by lipid peroxidation and electrolyte leakage (increased by 29–58%) in mycelial cells. These results suggested that thymol induced oxidative damage in mycelia, which may be one of the possible reasons for thymol-induced mycelial cell death observed with fluorescent detection. Thymol decreased the production of conidia and inhibited the germination of conidia. Thymol induced superoxide radical accumulation, lipid peroxidation, and cell death in conidia as well. All of these results revealed the inhibitory activity of thymol against FON, which may have resulted from the superoxide radical-induced oxidative injury in both conidia and mycelia of FON.

Functionalization of experimental HIV-1 virus-like particle vaccines with heterologous T helper epitopes (T helper VLPs) can modulate the humoral immune response via intrastructural help (ISH). Current advances in the conjugation of native-like HIV-1 envelope trimers (Env) onto liposomes and encapsulation of peptide epitopes into these nanoparticles renders this GMP-scalable liposomal platform a feasible alternative to VLP-based vaccines. In this study, we designed and analyzed customizable Env-conjugated T helper liposomes. First, we passively encapsulated T helper peptides into a well-characterized liposome formulation displaying a dense array of Env trimers on the surface. We confirmed the closed pre-fusion state of the coupled Env trimers by immunogold staining with conformation-specific antibodies. These peptide-loaded Env-liposome conjugates efficiently activated Env-specific B cells, which further induced proliferation of CD4+ T cells by presentation of liposome-derived peptides on MHC-II molecules. The peptide encapsulation process was then quantitatively improved by an electrostatically driven approach using an overall anionic lipid formulation. We demonstrated that peptides delivered by liposomes were presented by DCs in secondary lymphoid organs after intramuscular immunization of mice. UFO (uncleaved prefusion optimized) Env trimers were covalently coupled to peptide-loaded anionic liposomes by His-tag/NTA(Ni) interactions and EDC/Sulfo-NHS crosslinking. EM imaging revealed a moderately dense array of well-folded Env trimers on the liposomal surface. The conformation was verified by liposomal surface FACS. Furthermore, anionic Env-coupled T helper liposomes effectively induced Env-specific B cell activation and proliferation in a comparable range to T helper VLPs. Taken together, we demonstrated that T helper VLPs can be substituted with customizable and GMP-scalable liposomal nanoparticles as a perspective for future preclinical and clinical HIV vaccine applications. The functional nanoparticle characterization assays shown in this study can be applied to other systems of synthetic nanoparticles delivering antigens derived from various pathogens.

Abstract Background Sepsis remains a major health issue without an effective therapy. Ferroptosis, an iron‐dependent programmed cell death, has been proposed to be related to the pathogenesis of sepsis. Irisin, a myokine released during exercise, improves mitochondrial function under various conditions. Ferroptosis is closely related to mitochondrial function. However, the role of irisin in sepsis‐induced ferroptosis and mitochondrial dysfunction in the liver remained unknown. Thus, we hypothesize that irisin treatment suppresses ferroptosis and improves mitochondrial function in sepsis. Methods To study this, we first explored the role of serum irisin levels in patients with sepsis, and then determined the effect of irisin administration on ferroptosis and mitochondrial function in the liver of septic mice. Results Serum irisin levels were decreased and negatively correlated with the APACHE II scores in patients with sepsis. In mice subjected to cecal ligation and puncture (CLP), exogenous irisin administration suppressed ferroptosis, inhibited inflammatory response, decreased reactive oxygen species (ROS) production, restored abnormal mitochondrial morphology, and increased mtDNA copy number and adenosine triphosphate (ATP) content. The effect of irisin on ferroptosis was confirmed in LPS‐treated hepatocytes and CLP‐induced septic mice. Inhibition of glutathione peroxidase 4 (GPX4), a central regulator of ferroptosis, reduced irisin's protective effects in LPS‐treated hepatocytes and CLP‐induced septic mice, while blocking the irisin receptor with RGD peptide or Echistain decreased irisin‐induced GPX4 expression. Conclusions Serum irisin levels are decreased and negatively correlated with disease severity in patients with sepsis, and irisin treatment suppresses ferroptosis and restores mitochondrial function in experimental sepsis. Irisin may offer therapeutic potential in the management of sepsis.

Current management of liver ischemia-reperfusion (I/R) injury is mainly based on supportive care and no specific treatment is available. Irisin, a recently identified hormone, plays pivotal roles in energy expenditure and oxidative metabolism; however, it remains unknown whether irisin has any protective effects on hepatic I/R injury. In this study, we found that serum and liver irisin levels were markedly decreased at 24 h after hepatic I/R. Treatment with exogenous irisin improved liver function, reduced liver necrosis and cell apoptosis, and relieved inflammatory response after hepatic I/R. Meanwhile, exogenous irisin markedly inhibited mitochondrial fission related protein dynamin related protein 1 (drp-1) and fission 1 (Fis-1) expression in hepatic I/R. Additionally, treatment with exogenous irisin increased mitochondrial content and increased mitochondrial biogenesis related peroxisome proliferative activated receptor-γ (PPARγ) co-activator 1α (PGC-1α) and mitochondrial transcription factor (TFAM) expression. Furthermore, irisin decreased oxidative stress by upregulating uncoupling proteins (UCP) 2 expression in hepatic I/R. The results reveal that treatment with exogenous irisin alleviated hepatic I/R injury by restraining mitochondrial fission, promoting mitochondrial biogenesis and relieving oxidative stress. Irisin treatment appears to be a novel and promising therapeutic approach for hepatic I/R injury. Keywords: Hepatic I/R, Irisin, Mitochondrial homeostasis, Oxidative stress

Summary: Although intramuscular (i.m.) administration is the most commonly used route for licensed vaccines, subcutaneous (s.c.) delivery is being explored for several new vaccines under development. Here, we use rhesus macaques, physiologically relevant to humans, to identify the anatomical compartments and early immune processes engaged in the response to immunization via the two routes. Administration of fluorescently labeled HIV-1 envelope glycoprotein trimers displayed on liposomes enables visualization of targeted cells and tissues. Both s.c. and i.m. routes induce efficient immune cell infiltration, activation, and antigen uptake, functions that are tightly restricted to the skin and muscle, respectively. Antigen is also transported to different lymph nodes depending on route. However, these early differences do not translate into significant differences in the magnitude or quality of antigen-specific cellular and humoral responses over time. Thus, although some distinct immunological differences are noted, the choice of route may instead be motivated by clinical practicality. : Route of immunization, especially intramuscular versus subcutaneous administration, is often debated. Ols et al. use a rhesus macaque model to determine the tissues targeted by a nanoparticle vaccine administered by either route. The authors demonstrate that tissue dissemination is route dependent, but innate and adaptive immune responses develop comparably. Keywords: vaccination, intramuscular, subcutaneous, antigen transport, HIV envelope glycoprotein, dendritic cell, follicular dendritic cell, monocytes, lymph node, B cell follicle

Zircon Hf-isotopic mapping can be regarded as a useful tool for evaluating the coupling relationship between lithospheric structure and metallic mineralization. Hence, this method shows important significance for mineral prediction. To explore this potential, the published granite zircon Hf isotope data from the Sanjiang Tethyan Orogen were systematically compiled. This study uses the Kriging weighted interpolation in the Mapgis software system to contour Hf isotopes, revealing a relation between the crustal structure and metallogenesis. The mapping results suggest that the Changning–Menglian suture zone is the boundary between ancient and juvenile crust, viz., the western terranes have ancient crust attributes, whereas the eastern terranes exhibit the properties of new juvenile crust. In addition, this study also found that the mineralization and element types in the Sanjiang Tethyan Orogen have a coupling relationship with the crustal structure. The distribution of porphyry Cu-Mo-Au deposits is mainly controlled by the new juvenile crust, whereas the magmatic-hydrothermal Sn-W and porphyry Mo-W(-Cu) deposits are closely related to the reworked ancient crust. The results of zircon Hf isotope mapping prove that the formation and spatial distribution of deposits are related to the composition and properties of the crust. Hf isotope mapping can reveal the regional metallogenic rules and explore metallogenic prediction and metallogenic potential evaluation.

The Oligocene to Miocene Baiyangping Cu–Co and Pb–Zn mineralization was deposited during the collision of the India and Asia continents. The mineralization is hosted in veins in faulted carbonate and clastic rocks, which act as fluid pathways. A detailed fluid-inclusion analysis and C–O–S isotopic study are reported, focusing on the origin of the mineralizing fluids and mineralizing processes. The microthermometry of fluid inclusions hosted in quartz and carbonates indicate that the fluid peak temperature for both Cu–Co and Pb–Zn mineralizations ranges from 160 to 200 °C and from 120 to 160 °C, respectively. During these metallogenic hydrothermal events, salinity evolved from 2.7 wt% NaCl equiv to 24.8 wt% NaCl equiv. The δ34S values measured on sphalerite and tennantite range from +5.2‰ to +9.5‰, which indicates that sulfur originated from a reservoir characterized by thermochemical sulfate reduction (TSR). The C–O isotopic values obtained from the carbonate samples suggest that the fluid mainly originates from basinal brines. The combination of C and S isotopic values of the Cu–Co and Pb–Zn mineralization indicate the sedimentary source of the basin. Mineralization is controlled by faults, thermochemical sulfate reduction, and the physicochemical conditions of the fluid.

Abstract Background Arabidopsis pumila is native to the desert region of northwest China and it is extraordinarily well adapted to the local semi-desert saline soil, thus providing a candidate plant system for environmental adaptation and salt-tolerance gene mining. However, understanding of the salt-adaptation mechanism of this species is limited because of genomic sequences scarcity. In the present study, the transcriptome profiles of A. pumila leaf tissues treated with 250 mM NaCl for 0, 0.5, 3, 6, 12, 24 and 48 h were analyzed using a combination of second-generation sequencing (SGS) and third-generation single-molecule real-time (SMRT) sequencing. Results Correction of SMRT long reads by SGS short reads resulted in 59,328 transcripts. We found 8075 differentially expressed genes (DEGs) between salt-stressed tissues and controls, of which 483 were transcription factors and 1157 were transport proteins. Most DEGs were activated within 6 h of salt stress and their expression stabilized after 48 h; the number of DEGs was greatest within 12 h of salt stress. Gene annotation and functional analyses revealed that expression of genes associated with the osmotic and ionic phases rapidly and coordinately changed during the continuous salt stress in this species, and salt stress-related categories were highly enriched among these DEGs, including oxidation–reduction, transmembrane transport, transcription factor activity and ion channel activity. Orphan, MYB, HB, bHLH, C3H, PHD, bZIP, ARF and NAC TFs were most enriched in DEGs; ABCB1, CLC-A, CPK30, KEA2, KUP9, NHX1, SOS1, VHA-A and VP1 TPs were extensively up-regulated in salt-stressed samples, suggesting that they play important roles in slat tolerance. Importantly, further experimental studies identified a mitogen-activated protein kinase (MAPK) gene MAPKKK18 as continuously up-regulated throughout salt stress, suggesting its crucial role in salt tolerance. The expression patterns of the salt-responsive 24 genes resulted from quantitative real-time PCR were basically consistent with their transcript abundance changes identified by RNA-Seq. Conclusion The full-length transcripts generated in this study provide a more accurate depiction of gene transcription of A. pumila. We identified potential genes involved in salt tolerance of A. pumila. These data present a genetic resource and facilitate better understanding of salt-adaptation mechanism for ephemeral plants.

In this work, the tensile-shear mechanical behaviors of friction stir spot weld and adhesive hybrid joint were performed from both numerical and experimental viewpoints. Weld through (WT) and flow in (FI) processes were studied in this research. The focus was to evaluate joint defects, tensile-shear failure load (TSFL), failure energy, failure mode and stress distribution of the joint. In FI joints, keyhole and hook defects appeared in the weld zone and the areas of material removed from the base metal were filled with adhesive. In the WT joints, the adhesive layer close to the weld zone was carbonized due to the welding heat. Meanwhile, under the rotating movement of welding tool, the adhesive impurities entered the stirring zone (SZ) and heat affected zone (HAZ) of the weld, which decreased mechanical performances of WT joints. Compared to the friction stir spot welding (FSSW) joint, the TSFL value, stiffness and failure energy of FI joint were increased by 2.7, 1.1 and 8.14 times, respectively. In order to study the stress distribution of the joints, a finite element (FE) model, which considered the weld structure and mechanical properties of weld regions, was implemented. Moreover, the adhesive layer was simplified by the cohesive zone model (CZM). FE results show that the FI process effectively decreases the stress concentration of the weld edge from 243.09 to 15.5 MPa, under the 2 kN tensile load. The weld can block the adhesive crack propagation, and the adhesive optimized the stress distribution of FI joints through a synergistic effect. So, the use of FI process for aluminum alloy connection is strongly recommended, especially in crucial structure areas.

Soluble HIV-1 envelope glycoprotein (Env) trimers are under active investigation as vaccine candidates in relevant pre-clinical models. Like SOSIPs, the cleavage-independent native flexibly linked (NFL) trimers are faithful mimics of the Env spike. Here, we analyzed multiple new designs to explore alternative modifications, informing tertiary interactions, while maintaining NFL trimer homogeneity and integrity. Accordingly, we performed a proline (P) substitution screen in the gp41 heptad repeat 1 region, identifying other trimer-enhancing Ps, including L555P. This P improved trimer integrity compared to I559P in selected properties. Next, we screened 15 structure-guided potential cysteine pairs in gp140 and found that A501C-L663C (“CC2”) forms an inter-protomer disulfide bond that demonstrably increased NFL trimer thermostability. We combined these two approaches with trimer-derived substitutions, coupled with glycine substitutions at helix-to-coil transitions, developed by our group. To increase the exposure of the fusion peptide (FP) N-terminus, we engineered an enterokinase (EK) cleavage site upstream of the FP for controlled post-expression cleavage. In combination, the redesigns resulted in highly stable and homogeneous NFL mimics derived from different clades. Following recombinant EK cleavage, the NFL trimers retained covalent linkage, maintaining a native-like structure while displaying enhanced stability and favorable antigenic features. These trimers also displayed increased exposure of neutralizing epitopes in the FP and gp120/gp41 interface, while retaining other neutralizing epitopes and occluding non-neutralizing elements. This array of Env-structure-guided designs reveals additional interactive regions in the prefusion state of the HIV Env spike, affording the development of novel antigens and immunogens.

Well-ordered soluble HIV-1 envelope glycoprotein (Env) spike mimetics such as Native Flexibly Linked (NFL) trimers display high homogeneity, desired antigenicity, and high in vitro stability compared to previous generation soluble HIV-1 Env trimers. Glutaraldehyde (GLA) cross-linking was shown to further increase the thermostability of clade C 16055 NFL trimers and enhance the induction of tier 2 autologous neutralizing antibodies in guinea pigs. Here, we investigated if GLA fixation affected other aspects of the Env-specific immune response by performing a comparative immunogenicity study in C57BL/6 mice with non-fixed and GLA-fixed 16055 NFL trimers administered in AbISCO-100 adjuvant. We detected lower Env-specific binding antibody titers and increased skewing toward Th2 responses in mice immunized with GLA-fixed trimers compared to mice immunized with unfixed trimers, as shown by a higher Env-specific IgG1:IgG2b antibody subclass ratio. These results suggest that the presence of GLA adducts on Env influences the quality of the induced antibody response.

Arabidopsis pumila is an ephemeral plant, and a close relative of the model plant Arabidopsis thaliana, but it possesses higher photosynthetic efficiency, higher propagation rate, and higher salinity tolerance compared to those A. thaliana, thus providing a candidate plant system for gene mining for environmental adaption and salt tolerance. However, A. pumila is an under-explored resource for understanding the genetic mechanisms underlying abiotic stress adaptation. To improve our understanding of the molecular and genetic mechanisms of salt stress adaptation, more than 19,900 clones randomly selected from a cDNA library constructed previously from leaf tissue exposed to high-salinity shock were sequenced. A total of 16,014 high-quality expressed sequence tags (ESTs) were generated, which have been deposited in the dbEST GenBank under accession numbers JZ932319 to JZ948332. Clustering and assembly of these ESTs resulted in the identification of 8,835 unique sequences, consisting of 2,469 contigs and 6,366 singletons. The blastx results revealed 8,011 unigenes with significant similarity to known genes, while only 425 unigenes remained uncharacterized. Functional classification demonstrated an abundance of unigenes involved in binding, catalytic, structural or transporter activities, and in pathways of energy, carbohydrate, amino acid, or lipid metabolism. At least seven main classes of genes were related to salt-tolerance among the 8,835 unigenes. Many previously reported salt tolerance genes were also manifested in this library, for example VP1, H+-ATPase, NHX1, SOS2, SOS3, NAC, MYB, ERF, LEA, P5CS1. In addition, 251 transcription factors were identified from the library, classified into 42 families. Lastly, changes in expression of the 12 most abundant unigenes, 12 transcription factor genes, and 19 stress-related genes in the first 24 h of exposure to high-salinity stress conditions were monitored by qRT-PCR. The large-scale EST library obtained in this study provides first-hand information on gene sequences expressed in young leaves of A. pumila exposed to salt shock. The rapid discovery of known or unknown genes related to salinity stress response in A. pumila will facilitate the understanding of complex adaptive mechanisms for ephemerals.

Graphene-silicon (Gr-Si) Schottky solar cell has attracted much attention recently as promising candidate for low-cost photovoltaic application. For the fabrication of Gr-Si solar cell, the Gr film is usually transferred onto the Si substrate by wet transfer process. However, the impurities induced by this process at the graphene/silicon (Gr/Si) interface, such as H2O and O2, degrade the photovoltaic performance of the Gr-Si solar cell. We found that the thermal annealing process can effectively improve the photovoltaic performance of the Gr-Si solar cell by removing these impurities at the Gr/Si interface. More interestingly, the photovoltaic performance of the Gr-Si solar cell can be improved, furthermore, when exposed to air environment after the thermal annealing process. Through investigating the characteristics of the Gr-Si solar cell and the properties of the Gr film (carrier density and sheet resistance), we point out that this phenomenon is caused by the natural doping effect of the Gr film.

Forest landowner cooperatives have emerged as a formal institution capable of fostering professional forest management. There are many reasons to motivate participation in forest cooperatives. For instance, cooperatives could ease nonindustrial family forest landowners (NIFFLs) access to financial assistance and enhance market entrance opportunities. In China, various forms of forest cooperatives supported by government agencies are developing rapidly in an effort to facilitate forest management at a large scale. However, the development of forest cooperatives is still in its early stages. An understanding of NIFFLs' attitudes towards the development of cooperatives is instrumental for crafting policies and organizational structures that motivate landowners' participation. Three hundred and ten NIFFLs from Fujian and Jiangxi provinces in Southern China were interviewed to elicit their attitudes and willingness-to-participate in forest cooperatives. Logistic regression was used to evaluate landowner, household, forest resource, and institutional factors associated with NIFFLs' stated decision to participate in a forest cooperative. Results show that NIFFLs' stated willingness-to-participate in forest cooperatives was associated with demographic characteristics like education, forest-related income, size of forestland, and attitudes toward land tenure reform and cooperatives. This paper outlines suggestions on how to further develop forest cooperatives in Southern China.

An effective HIV vaccine likely requires the elicitation of neutralizing antibodies (NAbs) against multiple HIV-1 clades. The recently developed cleavageindependent native flexibly linked (NFL) envelope (Env) trimers exhibit wellordered conformation and elicit autologous tier 2 NAbs in multiple animal models. Here, we investigated whether the fusion of molecular adjuvant C3d to the Env trimers can improve B-cell germinal center (GC) formation and antibody responses. To generate Env-C3d trimers, we performed a glycineserine-based (G4S) flexible peptide linker screening and identified a linker range that allowed native folding. A 30-60-amino-acid-long linker facilitates Env-to-C3d association and achieves the secretion of well-ordered trimers and the structural integrity and functional integrity of Env and C3d. The fusion of C3d did not dramatically affect the antigenicity of the Env trimers and enhanced the ability of the Env trimers to engage and activate B cells in vitro. In mice, the fusion of C3d enhanced germinal center formation, the magnitude of Env-specific binding antibodies, and the avidity of the antibodies in the presence of an adjuvant. The Sigma Adjuvant System (SAS) did not affect the trimer integrity in vitro but contributed to altered immunogenicity in vivo, resulting in increased tier 1 neutralization, likely by increased exposure of variable region 3 (V3). Taken together, the results indicate that the fusion of the molecular adjuvant, C3d, to the Env trimers improves antibody responses and could be useful for Env-based vaccines against HIV. [ABSTRACT FROM AUTHOR]

Well-ordered soluble HIV-1 envelope glycoprotein (Env) spike mimetics such as Native Flexibly Linked (NFL) trimers display high homogeneity, desired antigenicity, and high in vitro stability compared to previous generation soluble HIV-1 Env trimers. Glutaraldehyde (GLA) cross-linking was shown to further increase the thermostability of clade C 16055 NFL trimers and enhance the induction of tier 2 autologous neutralizing antibodies in guinea pigs. Here, we investigated if GLA fixation affected other aspects of the Env-specific immune response by performing a comparative immunogenicity study in C57BL/6 mice with non-fixed and GLA-fixed 16055 NFL trimers administered in AbISCO-100 adjuvant. We detected lower Env-specific binding antibody titers and increased skewing toward Th2 responses in mice immunized with GLA-fixed trimers compared to mice immunized with unfixed trimers, as shown by a higher Env-specific IgG1:IgG2b antibody subclass ratio. These results suggest that the presence of GLA adducts on Env influences the quality of the induced antibody response. [ABSTRACT FROM AUTHOR]

Cd (cadmium) stress always alters the homeostasis of ROS (reactive oxygen species) including H2O2 (hydrogen sulfide) and O2·- (superoxide radical), leading to the oxidative injury and growth inhibition in plants. In addition to triggering oxidative injury, ROS has been suggested as important regulators modulating root elongation. However, whether and how Cd stress induces the inhibition of root elongation by differentially regulating endogenous H2O2 and O2·-, rather than by inducing oxidative injury, remains elusive. To address these gaps, histochemical, physiological, and biochemical approaches were applied to investigate the mechanism for Cd to fine-tune the balance between H2O2 and O2·- in the root tip of Brassica rapa. Treatment with Cd at 4 and 16 µM significantly inhibited root elongation, while only 16 µM but not 4 µM of Cd induced oxidative injury and cell death in root tip. Fluorescent and pharmaceutical tests suggested that H2O2 and O2·- played negative and positive roles, respectively, in the regulation of root elongation in the presence of Cd (4 µM) or not. Treatment with Cd at 4 µM led to the increase in H2O2 and the decrease in O2·- in root tip, which may be attributed to the up-regulation of Br_UPB1s and the down-regulation of their predicted targets (four peroxidase genes). Cd at 4 µM resulted in the increase in endogenous H2S in root tip by inducing the up-regulation of LCDs and DCDs. Treatment with H2S biosynthesis inhibitor or H2S scavenger significantly blocked Cd (4 µM)-induced increase in endogenous H2S level, coinciding with the recovery of root elongation, the altered balance between H2O2 and O2·-, and the expression of Br_UPB1s and two peroxidase genes. Taken together, it can be proposed that endogenous H2S mediated the phytotoxicity of Cd at low concentration by regulating Br_UPB1s-modulated balance between H2O2 and O2·- in root tip. Such findings shed new light on the regulatory role of endogenous H2S in plant adaptions to Cd stress. [ABSTRACT FROM AUTHOR]

The development of a successful vaccine against human immunodeficiency virus type 1 (HIV-1) likely requires immunogens that elicit both broadly neutralizing antibodies against envelope spikes and T cell responses that recognize multiple viral proteins. HIV-1 virus-like particles (VLP), because they display authentic envelope spikes on the particle surface, may be developed into such immunogens. However, in one way or the other current systems for HIV-1 VLP production have many limitations. To overcome these, in the present study we developed a novel strategy to produce HIV-1 VLP using stably transfected Drosophila S2 cells. We cotransfected S2 cells with plasmids encoding HIV-1 envelope, Gag, and Rev proteins and a selection marker. After stably transfected S2 clones were established, HIV-1 VLP and their immunogenicity in mice were carefully evaluated. Here, we report that HIV-1 envelope proteins are properly cleaved, glycosylated, and incorporated into VLP with Gag. The amount of VLP released into culture supernatants is comparable to those produced by insect cells infected with recombinant baculoviruses. Moreover, cryo-electron microscopy tomography revealed average 17 spikes per purified VLP, and antigenic epitopes on the spikes were recognized by the broadly neutralizing antibodies 2G12, b12, VRC01, and 4E10 but not by PG16. Finally, mice primed with DNA and boosted with VLP in the presence of CpG exhibited anti-envelope antibody responses, including ELISA-binding, neutralizing, antibody-dependent cell-mediated cytotoxicity and antibody-dependent cell-mediated viral inhibition, as well as envelope and Gag-specific CD8 T cell responses. Thus, we conclude that HIV-1 VLP produced by the S2 expression system has many desirable features to be developed into a vaccine component against HIV-1. [ABSTRACT FROM AUTHOR]

We studied the effects of selenite on growth and anthocyanin accumulation in purple lettuce. The results will ascertain which selenite concentration is more advantageous to the purple lettuce and help study the effect of selenite on the molecular mechanism of plant anthocyanin accumulation using the quantitative reverse transcription polymerase chain reaction (qRT-PCR). In this study, the low selenite concentrations (≤ 8μM) could not only promote purple lettuce growth including plant height, leaf area (LA), and fresh weight (FW), but also the anthocyanin contents which may be due to how the selenite affects the UDPglycose flavonoid glycosyl transferase (UFGT) and flavanone 3-hydroxylase (F3H) genes expression of anthocyanin biosynthesis. Next, on the basis of the obtained results from the preliminary experiments, 8 μM selenite was used to analyze the anthocyanin accumulation with the treatment time prolonged. Although the anthocyanin content of purple lettuce was not reduced on day 24, the control plants were significantly decreased. The reason may be that compared with the control plants, UFGT and F3H genes were markedly up-regulated on day 24. Therefore, the influence of selenium on anthocyanin accumulation and molecular regulation of anthocyanin synthesis is mainly due to the expression levels of the F3H and UFGT gene. It needs to be a further studied. [ABSTRACT FROM AUTHOR]