Structural basis of antibody inhibition and chemokine activation of the human CC chemokine receptor 8.

The C-C motif chemokine receptor 8 (CCR8) is a class A G-protein coupled receptor that has emerged as a promising therapeutic target in cancer. Targeting CCR8 with an antibody has appeared to be an attractive therapeutic approach, but the molecular basis for chemokine-mediated activation and antibody-mediated inhibition of CCR8 are not fully elucidated. Here, we obtain an antagonist antibody against human CCR8 and determine structures of CCR8 in complex with either the antibody or the endogenous agonist ligand CCL1. Our studies reveal characteristic antibody features allowing recognition of the CCR8 extracellular loops and CCL1-CCR8 interaction modes that are distinct from other chemokine receptor - ligand pairs. Informed by these structural insights, we demonstrate that CCL1 follows a two-step, two-site binding sequence to CCR8 and that antibody-mediated inhibition of CCL1 signaling can occur by preventing the second binding event. Together, our results provide a detailed structural and mechanistic framework of CCR8 activation and inhibition that expands our molecular understanding of chemokine - receptor interactions and offers insight into the development of therapeutic antibodies targeting chemokine GPCRs.

Transcription factors ZmNF-YA1 and ZmNF-YB16 regulate plant growth and drought tolerance in maize.

The identification of drought stress regulatory genes is crucial for the genetic improvement of maize (Zea mays L.) yield. Nuclear factors Y (NF-Ys) are important transcription factors, but their roles in the drought stress tolerance of plants and underlying molecular mechanisms are largely unknown. In this work, we used yeast two-hybrid screening to identify potential interactors of ZmNF-YB16 and confirmed the interaction between ZmNF-YA1 and ZmNF-YB16-YC17 and between ZmNF-YA7 and ZmNF-YB16-YC17. ZmNF-YB16 interacted with ZmNF-YC17 via its histone fold domain to form a heterodimer in the cytoplasm and then entered the nucleus to form a heterotrimer with ZmNF-YA1 or ZmNF-YA7 under osmotic stress. Overexpression of ZmNF-YA1 improved drought and salt stress tolerance and root development of maize, whereas zmnf-ya1 mutants exhibited drought and salt stress sensitivity. ZmNF-YA1-mediated transcriptional regulation, especially in JA signaling, histone modification, and chromatin remodeling, could underlie the altered stress tolerance of zmnf-ya1 mutant plants. ZmNF-YA1 bound to promoter CCAAT motifs and directly regulated the expression of multiple genes that play important roles in stress responses and plant development. Comparison of ZmNF-YB16- and ZmNF-YA1-regulated genes showed that ZmNF-YA1 and ZmNF-YB16 have similar biological functions in stress responses but varied functions in other biological processes. Taken together, ZmNF-YA1 is a positive regulator of plant drought and salt stress responses and is involved in the root development of maize, and ZmNF-Y complexes with different subunits may have discrepant functions.

Microwave-assisted hydrothermal synthesis of copper selenides (Cu2-xSe) thin films for quantum dots-sensitized solar cells.

In this work, copper selenide (Cu2-xSe) thin films were grown on FTO conductive glass substrates using a facile microwave-assisted hydrothermal method. The effects of synthesis parameters such as precursor components and deposition time on the stoichiometry and morphology of the synthesized films were systematically investigated through different techniques including XRD, SEM, and AFM. In order to evaluate the electrochemical catalytic performance of the synthesized copper selenide in electrolyte containing the sulfide/polysulfide redox couple, we assembled liquid-junction quantum dots-sensitized solar cells (QDSSC) using the synthesized copper selenide thin films as counter electrodes and CdSe quantum dots-sensitized mesoporous TiO2as photoanodes. Under the illumination of one Sun (100 mW cm-2), the QDSSC assembled with the optimal copper selenide CEs (Cu:Se = 1:1) exhibited a power conversion efficiency of 2.07%, which is much higher than that of traditional Pt counter electrode (0.76%).

High-fat diet-induced upregulation of exosomal phosphatidylcholine contributes to insulin resistance.

High-fat diet (HFD) decreases insulin sensitivity. How high-fat diet causes insulin resistance is largely unknown. Here, we show that lean mice become insulin resistant after being administered exosomes isolated from the feces of obese mice fed a HFD or from patients with type II diabetes. HFD altered the lipid composition of exosomes from predominantly phosphatidylethanolamine (PE) in exosomes from lean animals (L-Exo) to phosphatidylcholine (PC) in exosomes from obese animals (H-Exo). Mechanistically, we show that intestinal H-Exo is taken up by macrophages and hepatocytes, leading to inhibition of the insulin signaling pathway. Moreover, exosome-derived PC binds to and activates AhR, leading to inhibition of the expression of genes essential for activation of the insulin signaling pathway, including IRS-2, and its downstream genes PI3K and Akt. Together, our results reveal HFD-induced exosomes as potential contributors to the development of insulin resistance. Intestinal exosomes thus have potential as broad therapeutic targets.

Antigen-specific B cell depletion for precision therapy of mucosal pemphigus vulgaris.

Desmoglein 3 chimeric autoantibody receptor T cells (DSG3-CAART) expressing the pemphigus vulgaris (PV) autoantigen DSG3 fused to CD137-CD3ζ signaling domains, represent a precision cellular immunotherapy approach for antigen-specific B cell depletion. Here, we present definitive preclinical studies enabling a first-in-human trial of DSG3-CAART for mucosal PV. DSG3-CAART specifically lysed human anti-DSG3 B cells from PV patients and demonstrated activity consistent with a threshold dose in vivo, resulting in decreased target cell burden, decreased serum and tissue-bound autoantibodies, and increased DSG3-CAART engraftment. In a PV active immune model with physiologic anti-DSG3 IgG levels, DSG3-CAART inhibited antibody responses against pathogenic DSG3 epitopes and autoantibody binding to epithelial tissues, leading to clinical and histologic resolution of blisters. DSG3 autoantibodies stimulated DSG3-CAART IFN-γ secretion and homotypic clustering, consistent with an activated phenotype. Toxicology screens using primary human cells and high-throughput membrane proteome arrays did not identify off-target cytotoxic interactions. These preclinical data guided the trial design for DSG3-CAART and may help inform CAART preclinical development for other antibody-mediated diseases.

Frontline Science: Activation of metabolic nuclear receptors restores periodontal tissue homeostasis in mice with leukocyte adhesion deficiency-1.

ß2 Integrins mediate neutrophil-endothelial adhesion and recruitment of neutrophils to sites of inflammation. The diminished expression of ß2 integrins in patients with mutations in the ITGB2 (CD18) gene (leukocyte adhesion deficiency-Type 1; LAD1) results in few or no neutrophils in peripheral tissues. In the periodontium, neutrophil paucity is associated with up-regulation of IL-23 and IL-17, which drive inflammatory bone loss. Using a relevant mouse model, we investigated whether diminished efferocytosis (owing to neutrophil scarcity) is associated with LAD1 periodontitis pathogenesis and aimed to develop approaches to restore the missing efferocytosis signals. We first showed that CD18-/- mice phenocopied human LAD1 in terms of IL-23/IL-17-driven inflammatory bone loss. Ab-mediated blockade of c-Mer tyrosine kinase (Mer), a major efferocytic receptor, mimicked LAD1-associated up-regulation of gingival IL-23 and IL-17 mRNA expression in wild-type (WT) mice. Consistently, soluble Mer-Fc reversed the inhibitory effect of efferocytosis on IL-23 expression in LPS-activated Mϕs. Adoptive transfer of WT neutrophils to CD18-/- mice down-regulated IL-23 and IL-17 expression to normal levels, but not when CD18-/- mice were treated with blocking anti-Mer Ab. Synthetic agonist-induced activation of liver X receptors (LXR) and peroxisome proliferator-activated receptors (PPAR), which link efferocytosis to generation of homeostatic signals, inhibited the expression of IL-23 and IL-17 and favorably affected the bone levels of CD18-/- mice. Therefore, our data link diminished efferocytosis-associated signaling due to impaired neutrophil recruitment to dysregulation of the IL-23-IL-17 axis and, moreover, suggest LXR and PPAR as potential therapeutic targets for treating LAD1 periodontitis.

Effects of maize organ-specific drought stress response on yields from transcriptome analysis.

BACKGROUND: Drought is a serious causal factor of reduced crop yields than any other abiotic stresses. As one of the most widely distributed crops, maize plants frequently suffer from drought stress, which causes great losses in the final kernel yield. Drought stress response in plants showed tissue- and developmental stage-specific characteristics. RESULTS: In this study, the ears at the V9 stage, kernels and ear leaf at the 5DAP (days after pollination) stage of maize were used for morphological, physiological and comparative transcriptomics analysis to understand the different features of "sink" or "source" organs and the effects on kernel yield under drought stress conditions. The ABA-, NAC-mediate signaling pathway, osmotic protective substance synthesis and protein folding response were identified as common drought stress response in the three organs. Tissue-specific drought stress responses and the regulators were identified, they were highly correlated with growth, physiological adaptation and yield loss under drought stress. For ears, drought stress inhibited ear elongation, led to the abnormal differentiation of the paired spikelet, and auxin signaling involved in the regulation of cell division and growth and primordium development changes. In the kernels, reduced kernel size caused by drought stress was observed, and the obvious differences of auxin, BR and cytokine signaling transduction appeared, which indicated the modification in carbohydrate metabolism, cell differentiation and growth retardation. For the ear leaf, dramatically and synergistically reduced the expression of photosynthesis genes were observed when suffered from drought stress, the ABA- and NAC- mediate signaling pathway played important roles in the regulation of photosynthesis. CONCLUSIONS: Transcriptomic changes caused by drought were highly correlated with developmental and physiological adaptation, which was closely related to the final yield of maize, and a sketch of tissue- and developmental stage-specific responses to drought stress in maize was drafted.

The bHLH family member ZmPTF1 regulates drought tolerance in maize by promoting root development and abscisic acid synthesis.

Drought stress is the most important environmental stress limiting maize production. ZmPTF1, a phosphate starvation-induced basic helix-loop-helix (bHLH) transcription factor, contributes to root development and low-phosphate tolerance in maize. Here, ZmPTF1 expression, drought tolerance, and the underlying mechanisms were studied by using maize ZmPTF1 overexpression lines and mutants. ZmPTF1 was found to be a positive regulator of root development, ABA synthesis, signalling pathways, and drought tolerance. ZmPTF1 was also found to bind to the G-box element within the promoter of 9-cis-epoxycarotenoid dioxygenase (NCED), C-repeat-binding factor (CBF4), ATAF2/NAC081, NAC30, and other transcription factors, and to act as a positive regulator of the expression of those genes. The dramatically upregulated NCEDs led to increased abscisic acid (ABA) synthesis and activation of the ABA signalling pathway. The up-regulated transcription factors hierarchically regulate the expression of genes involved in root development, stress responses, and modifications of transcriptional regulation. The improved root system, increased ABA content, and activated ABA-, CBF4-, ATAF2-, and NAC30-mediated stress responses increased the drought tolerance of the ZmPTF1 overexpression lines, while the mutants showed opposite trends. This study describes a useful gene for transgenic breeding and helps us understand the role of a bHLH protein in plant root development and stress responses.

Auxin and GA signaling play important roles in the maize response to phosphate deficiency.

Phytohormone signaling is involved in the low-phosphate (LP) response and causes root system changes. To understand the roles of auxin and gibberellic acid (GA) in the maize response to LP stress, inbred line Q319 was used to identify the changes in root morphology and the gene expression response to LP stress with or without exogenous auxin, GA or their inhibitors. The root morphology, IAA and GAs concentration and genes related to the LP response, cell elongation and division, auxin transport and signaling, and GA synthesis and signaling were analyzed. The LP-induced maize root morphological adaption was dependent on changes in the expression of related genes, like IPS1, pht1;1 LPR1b, KRPs, and EXPB1-4. The altered local auxin concentration and signaling were involved in promoting axial root elongation and reducing lateral root density and length under LP conditions, which were regulated by PID and PP2A activity and the auxin signaling pathway. The upregulation of the GA synthesis genes AN1, GA20ox1, and GA20ox2 and the downregulation of the GA inactive genes GA2ox1 and GA2ox2 were observed in maize roots subjected to LP stress, and the increased GA biosynthesis and signaling were involved in root growth. Both hormones participate in LP stress response and jointly regulated root modification and LP acclimation in maize.

Macrophage ß2-Integrins Regulate IL-22 by ILC3s and Protect from Lethal Citrobacter rodentium-Induced Colitis.

ß2-integrins promote neutrophil recruitment to infected tissues and are crucial for host defense. Neutrophil recruitment is defective in leukocyte adhesion deficiency type-1 (LAD1), a condition caused by mutations in the CD18 (ß2-integrin) gene. Using a model of Citrobacter rodentium (CR)-induced colitis, we show that CD18-/- mice display increased intestinal damage and systemic bacterial burden, compared to littermate controls, ultimately succumbing to infection. This phenotype is not attributed to defective neutrophil recruitment, as it is shared by CXCR2-/- mice that survive CR infection. CR-infected CD18-/- mice feature prominent upregulation of IL-17 and downregulation of IL-22. Exogenous IL-22 administration, but not endogenous IL-17 neutralization, protects CD18-/- mice from lethal colitis. ß2-integrin expression on macrophages is mechanistically linked to Rac1/ROS-mediated induction of noncanonical-NLRP3 (nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing-3) inflammasome-dependent IL-1ß production, which promotes ILC3-derived IL-22. Therefore, ß2-integrins are required for protective IL-1ß-dependent IL-22 responses in colitis, and the identified mechanism may underlie the association of human LAD1 with colitis.

DEL-1 promotes macrophage efferocytosis and clearance of inflammation.

Resolution of inflammation is essential for tissue homeostasis and represents a promising approach to inflammatory disorders. Here we found that developmental endothelial locus-1 (DEL-1), a secreted protein that inhibits leukocyte-endothelial adhesion and inflammation initiation, also functions as a non-redundant downstream effector in inflammation clearance. In human and mouse periodontitis, waning of inflammation was correlated with DEL-1 upregulation, whereas resolution of experimental periodontitis failed in DEL-1 deficiency. This concept was mechanistically substantiated in acute monosodium-urate-crystal-induced inflammation, where the pro-resolution function of DEL-1 was attributed to effective apoptotic neutrophil clearance (efferocytosis). DEL-1-mediated efferocytosis induced liver X receptor-dependent macrophage reprogramming to a pro-resolving phenotype and was required for optimal production of at least certain specific pro-resolving mediators. Experiments in transgenic mice with cell-specific overexpression of DEL-1 linked its anti-leukocyte-recruitment action to endothelial cell-derived DEL-1 and its efferocytic/pro-resolving action to macrophage-derived DEL-1. Thus, the compartmentalized expression of DEL-1 facilitates distinct homeostatic functions in an appropriate context that can be harnessed therapeutically.

ZmbZIP4 Contributes to Stress Resistance in Maize by Regulating ABA Synthesis and Root Development.

In plants, bZIP (basic leucine zipper) transcription factors regulate diverse processes such as development and stress responses. However, few of these transcription factors have been functionally characterized in maize (Zea mays). In this study, we characterized the bZIP transcription factor gene ZmbZIP4 from maize. ZmbZIP4 was differentially expressed in various organs of maize and was induced by high salinity, drought, heat, cold, and abscisic acid treatment in seedlings. A transactivation assay in yeast demonstrated that ZmbZIP4 functioned as a transcriptional activator. A genome-wide screen for ZmbZIP4 targets by immunoprecipitation sequencing revealed that ZmbZIP4 could positively regulate a number of stress response genes, such as ZmLEA2, ZmRD20, ZmRD21, ZmRab18, ZmNHX3, ZmGEA6, and ZmERD, and some abscisic acid synthesis-related genes, including NCED, ABA1, AAO3, and LOS5 In addition, ZmbZIP4 targets some root development-related genes, including ZmLRP1, ZmSCR, ZmIAA8, ZmIAA14, ZmARF2, and ZmARF3, and overexpression of ZmbZIP4 resulted in an increased number of lateral roots, longer primary roots, and an improved root system. Increased abscisic acid synthesis by overexpression of ZmbZIP4 also can increase the plant's ability to resist abiotic stress. Thus, ZmbZIP4 is a positive regulator of plant abiotic stress responses and is involved in root development in maize.

Incorporation of DDR2 clusters into collagen matrix via integrin-dependent posterior remnant tethering.

Cell-matrix interactions play critical roles in cell adhesion, tissue remodeling and cancer metastasis. Discoidin domain receptor 2 (DDR2) is a collagen receptor belonging to receptor tyrosine kinase (RTK) family. It is a powerful regulator of collagen deposition in the extracellular matrix (ECM). Although the oligomerization of DDR extracellular domain (ECD) proteins can affect matrix remodeling by inhibiting fibrillogenesis, it is still unknown how cellular DDR2 is incorporated into collagen matrix. Using 3-dimentional (3D) imaging for migrating cells, we identified a novel mechanism that explains how DDR2 incorporating into collagen matrix, which we named as posterior remnant tethering. We followed the de novo formation of these remnants and identified that DDR2 clusters formed at the retracting phase of a pseudopodium, then these clusters were tethered to fibrillar collagen and peeled off from the cell body to generate DDR2 containing posterior remnants. Inhibition of ß1-integrin or Rac1 activity abrogated the remnant formation. Thus, our findings unveil a special cellular mechanism for DDR2 clusters incorporating into collagen matrix in an integrin-dependent manner.

ZmNF-YB16 Overexpression Improves Drought Resistance and Yield by Enhancing Photosynthesis and the Antioxidant Capacity of Maize Plants.

ZmNF-YB16 is a basic NF-YB superfamily member and a member of a transcription factor complex composed of NF-YA, NF-YB, and NF-YC in maize. ZmNF-YB16 was transformed into the inbred maize line B104 to produce homozygous overexpression lines. ZmNF-YB16 overexpression improves dehydration and drought stress resistance in maize plants during vegetative and reproductive stages by maintaining higher photosynthesis and increases the maize grain yield under normal and drought stress conditions. Based on the examination of differentially expressed genes between the wild-type (WT) and transgenic lines by quantitative real time PCR (qRT-PCR), ZmNF-YB16 overexpression increased the expression of genes encoding antioxidant enzymes, the antioxidant synthase, and molecular chaperones associated with the endoplasmic reticulum (ER) stress response, and improved protection mechanism for photosynthesis system II. Plants that overexpression ZmNF-YB16 showed a higher rate of photosynthesis and antioxidant enzyme activity, better membrane stability and lower electrolyte leakage under control and drought stress conditions. These results suggested that ZmNF-YB16 played an important role in drought resistance in maize by regulating the expression of a number of genes involved in photosynthesis, the cellular antioxidant capacity and the ER stress response.

Modulation of Myelopoiesis Progenitors Is an Integral Component of Trained Immunity.

Trained innate immunity fosters a sustained favorable response of myeloid cells to a secondary challenge, despite their short lifespan in circulation. We thus hypothesized that trained immunity acts via modulation of hematopoietic stem and progenitor cells (HSPCs). Administration of ß-glucan (prototypical trained-immunity-inducing agonist) to mice induced expansion of progenitors of the myeloid lineage, which was associated with elevated signaling by innate immune mediators, such as IL-1ß and granulocyte-macrophage colony-stimulating factor (GM-CSF), and with adaptations in glucose metabolism and cholesterol biosynthesis. The trained-immunity-related increase in myelopoiesis resulted in a beneficial response to secondary LPS challenge and protection from chemotherapy-induced myelosuppression in mice. Therefore, modulation of myeloid progenitors in the bone marrow is an integral component of trained immunity, which to date, was considered to involve functional changes of mature myeloid cells in the periphery.

TsNAC1 Is a Key Transcription Factor in Abiotic Stress Resistance and Growth.

NAC proteins constitute one of the largest families of plant-specific transcription factors, and a number of these proteins participate in the regulation of plant development and responses to abiotic stress. T. HALOPHILA STRESS RELATED NAC1 (TsNAC1), cloned from the halophyte Thellungiella halophila, is a NAC transcription factor gene, and its overexpression can improve abiotic stress resistance, especially in salt stress tolerance, in both T. halophila and Arabidopsis (Arabidopsis thaliana) and retard the growth of these plants. In this study, the transcriptional activation activity of TsNAC1 and RD26 from Arabidopsis was compared with the target genes' promoter regions of TsNAC1 from T. halophila, and the results showed that the transcriptional activation activity of TsNAC1 was higher in tobacco (Nicotiana tabacum) and yeast. The target sequence of the promoter from the target genes also was identified, and TsNAC1 was shown to target the positive regulators of ion transportation, such as T. HALOPHILA H+-PPASE1, and the transcription factors MYB HYPOCOTYL ELONGATION-RELATED and HOMEOBOX12 In addition, TsNAC1 negatively regulates the expansion of cells, inhibits LIGHT-DEPENDENT SHORT HYPOCOTYLS1 and UDP-XYLOSYLTRANSFERASE2, and directly controls the expression of MULTICOPY SUPPRESSOR OF IRA14 Based on these results, we propose that TsNAC1 functions as an important upstream regulator of plant abiotic stress responses and vegetative growth.

Secreted protein Del-1 regulates myelopoiesis in the hematopoietic stem cell niche.

Hematopoietic stem cells (HSCs) remain mostly quiescent under steady-state conditions but switch to a proliferative state following hematopoietic stress, e.g., bone marrow (BM) injury, transplantation, or systemic infection and inflammation. The homeostatic balance between quiescence, self-renewal, and differentiation of HSCs is strongly dependent on their interactions with cells that constitute a specialized microanatomical environment in the BM known as the HSC niche. Here, we identified the secreted extracellular matrix protein Del-1 as a component and regulator of the HSC niche. Specifically, we found that Del-1 was expressed by several cellular components of the HSC niche, including arteriolar endothelial cells, CXCL12-abundant reticular (CAR) cells, and cells of the osteoblastic lineage. Del-1 promoted critical functions of the HSC niche, as it regulated long-term HSC (LT-HSC) proliferation and differentiation toward the myeloid lineage. Del-1 deficiency in mice resulted in reduced LT-HSC proliferation and infringed preferentially upon myelopoiesis under both steady-state and stressful conditions, such as hematopoietic cell transplantation and G-CSF- or inflammation-induced stress myelopoiesis. Del-1-induced HSC proliferation and myeloid lineage commitment were mediated by ß3 integrin on hematopoietic progenitors. This hitherto unknown Del-1 function in the HSC niche represents a juxtacrine homeostatic adaptation of the hematopoietic system in stress myelopoiesis.

miRNA alterations are important mechanism in maize adaptations to low-phosphate environments.

Maize is a globally important crop, and a low phosphate (LP) supply frequently limits maize yields in many areas. microRNAs (miRNAs) play important roles in plant development and environmental adaptation. In this study, spatio-temporal miRNA transcript profiling and some of the target genes in the roots and leaves of the maize inbred line Q319 were analyzed in response to LP. Complex small RNA populations were detected after LP culture, and they displayed different patterns in the roots and leaves. Differentially expressed miRNAs can be grouped into 'early' miRNAs, which respond rapidly and are often non-specific to phosphate deficiency, and 'late' miRNAs, which alter the morphology, physiology or metabolism of plants upon prolonged phosphate deficiency. miR827 and miR399-mediated posttranscriptional pathway responses to phosphate availability were conserved and species-specific in maize. Abiotic stress-related miRNAs were engaged in interactions with different signaling and/or metabolic pathways. Auxin-related miRNAs and their targets' expression may be involved in root architecture modification and upland growth retardation in maize when subjected to LP. The changes that were found in the expression of miRNAs and their target genes suggested that miRNA regulation/alterations are pivotal mechanisms in maize adaptations to LP environments. A complex regulatory mechanism involving miRNAs in response to the LP environment is present in maize.

Complement inhibition in pre-clinical models of periodontitis and prospects for clinical application.

Periodontitis is a dysbiotic inflammatory disease leading to the destruction of the tooth-supporting tissues. Current therapies are not always effective and this prevalent oral disease continues to be a significant health and economic burden. Early clinical studies have associated periodontitis with elevated complement activity. Consistently, subsequent genetic and pharmacological studies in rodents have implicated the central complement component C3 and downstream signaling pathways in periodontal host-microbe interactions that promote dysbiosis and inflammatory bone loss. This review discusses these mechanistic advances and moreover focuses on the compstatin family of C3 inhibitors as a novel approach to treat periodontitis. In this regard, local application of the current lead analog Cp40 was recently shown to block both inducible and naturally occurring periodontitis in non-human primates. These promising results from non-human primate studies and the parallel development of Cp40 for clinical use highlight the feasibility for developing an adjunctive, C3-targeted therapy for human periodontitis.