RtNAC055 promotes drought tolerance via a stomatal closure pathway linked to methyl jasmonate/hydrogen peroxide signaling in Reaumuria trigyna.

The stomata regulate CO2 uptake and efficient water usage, thereby promoting drought stress tolerance. NAC proteins (NAM, ATAF1/2, and CUC2) participate in plant reactions following drought stress, but the molecular mechanisms underlying NAC-mediated regulation of stomatal movement are unclear. In this study, a novel NAC gene from Reaumuria trigyna, RtNAC055, was found to enhance drought tolerance via a stomatal closure pathway. It was regulated by RtMYC2 and integrated with jasmonic acid signaling and was predominantly expressed in stomata and root. The suppression of RtNAC055 could improve jasmonic acid and H2O2 production and increase the drought tolerance of transgenic R. trigyna callus. Ectopic expression of RtNAC055 in the Arabidopsis atnac055 mutant rescued its drought-sensitive phenotype by decreasing stomatal aperture. Under drought stress, overexpression of RtNAC055 in poplar promoted ROS (H2O2) accumulation in stomata, which accelerated stomatal closure and maintained a high photosynthetic rate. Drought upregulated the expression of PtRbohD/F, PtP5CS2, and PtDREB1.1, as well as antioxidant enzyme activities in heterologous expression poplars. RtNAC055 promoted H2O2 production in guard cells by directly binding to the promoter of RtRbohE, thus regulating stomatal closure. The stress-related genes RtDREB1.1/P5CS1 were directly regulated by RtNAC055. These results indicate that RtNAC055 regulates stomatal closure by maintaining the balance between the antioxidant system and H2O2 level, reducing the transpiration rate and water loss, and improving photosynthetic efficiency and drought resistance.

Concise Enantioselective Total Synthesis of Daphenylline Enabled by an Intramolecular Oxidative Dearomatization.

Daphenylline is a structurally unique member of the triterpenoid Daphniphyllum natural alkaloids, which exhibit intriguing biological activities. Six total syntheses have been reported, five of which utilize aromatization approaches. Herein, we report a concise protecting-group-free total synthesis by means of a novel intramolecular oxidative dearomatization reaction, which concurrently generates the critical seven-membered ring and the quaternary-containing vicinal stereocenters. Other notable transformations include a tandem reductive amination/amidation double cyclization reaction, to assemble the cage-like architecture, and installation of the other two chiral stereocenters via a highly enantioselective rhodium-catalyzed challenging hydrogenation of the diene intermediate (90% e.e.) and an unprecedented remote acid-directed Mukaiyama-Michael reaction of the complex benzofused cyclohexanone (13:1 d.r.).

RtNAC100 involved in the regulation of ROS, Na+ accumulation and induced salt-related PCD through MeJA signal pathways in recretohalophyte Reaumuria trigyna.

NAM, ATAF1/2, and CUC2 (NAC) proteins regulate plant responses to salt stress. However, the molecular mechanisms by which NAC proteins regulate salt-induced programmed cell death (PCD) are unclear. We identified 56 NAC genes, 35 of which had complete open reading frames with complete NAM domain, in the R. trigyna transcriptome. Salt stress and methyl jasmonate (MeJA) mediated PCD-induced leaf senescence in R. trigyna seedlings. Salt stress accelerated endogenous JA biosynthesis, upregulating RtNAC100 expression. This promoted salt-induced leaf senescence in R. trigyna by regulating RtRbohE and RtSAG12/20 and enhancing ROS accumulation. Transgenic assays showed that RtNAC100 overexpression aggravated salt-induced PCD in transgenic lines by promoting ROS and Na+ accumulation, ROS-Ca2+ hub activation, and PCD-related gene expression. Therefore, RtNAC100 induces PCD via the MeJA signaling pathway in R. trigyna under salt stress.

A leucoanthocyanidin dioxygenase gene (RtLDOX2) from the feral forage plant Reaumuria trigyna promotes the accumulation of flavonoids and improves tolerance to abiotic stresses.

Reaumuria trigyna, a Tamaricaceae archaic recretohalophyte, is an important feral forage plant in the desert steppe of northwestern China. We identified two significantly differentially expressed leucoanthocyanidin dioxygenase genes (RtLDOX/RtLDOX2) and investigated the function and characteristics of RtLDOX2. RtLDOX2 from R. trigyna was rapidly upregulated by salt, drought, and abscisic acid, consistent with the stress-related cis-regulatory elements in the promoter region. Recombinant RtLDOX2 converted dihydrokaempferol to kaempferol in vitro, and was thus interchangeable with flavonol synthase, a dioxygenase in the flavonoid pathway. Transgenic plants overexpressing RtLDOX2 accumulated more anthocyanin and flavonols under abiotic stresses, speculating that RtLDOX2 may act as a multifunctional dioxygenase in the synthesis of anthocyanins and flavonols. Overexpression of RtLDOX2 enhanced the primary root length, biomass accumulation, and chlorophyll content of salt-, drought-, and ultraviolet-B-stressed transgenic Arabidopsis. Antioxidant enzyme activity; proline content; and expression of antioxidant enzyme, proline biosynthesis, and ion-transporter genes were increased in transgenic plants. Therefore, RtLDOX2 confers tolerance to abiotic stress on transgenic Arabidopsis by promoting the accumulation of anthocyanins and flavonols. This in turn increases reactive oxygen species scavenging and activates other stress responses, such as osmotic adjustment and ion transport, and so improves tolerance to abiotic stresses.

Exogenous methyl jasmonate promotes salt stress-induced growth inhibition and prioritizes defense response of Nitraria tangutorum Bobr.

Jasmonates (JAs) play a key role in the regulation of growth and the defense response to environmental stresses. JAs inhibit plant growth and promote defense response. However, their roles in desert halophyte in the response to salt stress remain poorly understood. The effects of the combination of methyl jasmonate (MeJA) and NaCl treatment (the "MeN" condition) on the growth regulation and defense response of Nitraria tangutorum seedlings were investigated. Compared with NaCl treatment alone, exogenous MeJA aggravated the growth inhibition of seedlings by antagonizing to growth-related hormones and suppressing the transcript levels of these hormones-responsive genes, including gibberellin (GA)-responsive NtPIF3, NtGAST1, NtGSAT4, and cytokinin (CYT)-responsive NtARR1, NtARR11, NtARR12. Meanwhile, exogenous MeJA enhanced defense response and alleviated the stress damage by increasing antioxidase activity and antioxidant content, accumulating more osmolytes, maintaining lower Na+ /K+ ratios in shoots and higher Na+ efflux rates in roots of plants. In addition, exogenous MeJA increased the contents of endogenous JA and ABA, and the transcript levels of genes involved in their biosynthesis and responsiveness, thereby further regulating the transcript levels of defense response genes. These findings suggest that exogenous MeJA increases salt stress-induced growth inhibition and prioritizes the defensive responses (e.g. antioxidant defense, osmotic adjustment, and ion homeostasis) of N. tangutorum. These effects may be related to the amplification of jasmonic acid (JA) and abscisic acid (ABA) signals.

Glutaredoxin like protein (RtGRL1) regulates H2O2 and Na+ accumulation by maintaining the glutathione pool during abiotic stress.

Reaumuria trigyna, an endangered recretohalophyte, is a small archaic wild shrub endemic to arid and semiarid plateau regions of Inner Mongolia, China. Based on salt-related transcriptomic data, we isolated a GRX family gene, glutaredoxin like protein (RtGRL1), from R. trigyna that is associated with the removal of active oxygen and regulation of redox status. RtGRL1 encodes a plasma membrane and chloroplast-localized protein induced by salt, cold, drought stress, ABA, and H2O2. In Arabidopsis thaliana, ectopically expressed RtGRL1 positively regulated biomass accumulation, chlorophyll content, germination rate, and primary root length under salt and drought stress. Overexpression of RtGRL1 induced expression of genes related to antioxidant enzymes and proline biosynthesis, thus increasing glutathione biosynthesis, glutathione-dependent detoxification of reactive oxygen species (ROS), and proline content under stress. Changes in RtGRL1 expression consistently affected glutathione/oxidizedglutathione and ascorbate/dehydroascorbate ratios and H2O2 concentrations. Furthermore, RtGRL1 promoted several GSH biosynthesis gene transcripts, decreased leaf Na+ content, and maintained lower Na+/K+ ratios in transgenic A. thaliana compared to wild type plants. These results suggest a critical link between RtGRL1 and ROS modulation, and contribute to a better understanding of the mechanisms governing plant responses to drought and salt stress.

Optically Active Flavaglines-Inspired Molecules by a Palladium-Catalyzed Decarboxylative Dearomative Asymmetric Allylic Alkylation.

With the aid of a class of newly discovered Trost-type bisphosphine ligands bearing a chiral cycloalkane framework, the Pd-catalyzed decarboxylative dearomative asymmetric allylic alkylation (AAA) of benzofurans was achieved with high efficiency [0.2-1.0 mol% Pd2(dba)3/L], good generality, and high enantioselectivity (>30 examples, 82-99% yield and 90-96% ee). Moreover, a diversity-oriented synthesis (DOS) of previously unreachable flavaglines is disclosed. It features a reliable and scalable sequence of the freshly developed Tsuji-Trost-Stoltz AAA, a Wacker-Grubbs-Stoltz oxidation, an intra-benzoin condensation, and a conjugate addition, which allows the efficient construction of the challenging and compact cyclopenta[b]benzofuran scaffold with contiguous stereocenters. This strategy offers a new avenue for developing flavagline-based drugs.

Reaumuria trigyna transcription factor RtWRKY23 enhances salt stress tolerance and delays flowering in plants.

Reaumuria trigyna (Reaumuria Linn genus, family Tamaricaceae), an endangered dicotyledonous shrub with the features of a recretohalophyte, is endemic to the Eastern Alxa-Western Ordos area of China. Based on R. trigyna transcriptome data and expression pattern analysis of RtWRKYs, RtWRKY23, a Group II WRKY transcription factor, was isolated from R. trigyna cDNA. RtWRKY23 was mainly expressed in the stem and was induced by salt, drought, cold, ultraviolet radiation, and ABA treatments, but suppressed by heat treatment. Overexpression of RtWRKY23 in Arabidopsis increased chlorophyll content, root length, and fresh weight of the transgenic lines under salt stress. Real-time quantitative PCR (qPCR) analysis and yeast one-hybrid analysis demonstrated that RtWRKY23 protein directly or indirectly modulated the expression levels of downstream genes, including stress-related genes AtPOD, AtPOD22, AtPOD23, AtP5CS1, AtP5CS2, and AtPRODH2, and reproductive development-related genes AtMAF5, AtHAT1, and AtANT. RtWRKY23 transgenic Arabidopsis had higher proline content, peroxidase activity, and superoxide anion clearance rate, and lower H2O2 and malondialdehyde content than WT plants under salt stress conditions. Moreover, RtWRKY23 transgenic Arabidopsis exhibited later flowering and shorter pods, but little change in seed yield, compared with WT plants under salt stress. Our study demonstrated that RtWRKY23 not only enhanced salt stress tolerance through maintaining the ROS and osmotic balances in plants, but also participated in the regulation of flowering under salt stress.

Functional Analysis of Ion Transport Properties and Salt Tolerance Mechanisms of RtHKT1 from the Recretohalophyte Reaumuria trigyna.

Reaumuria trigyna is an endangered recretohalophyte and a small archaic feral shrub that is endemic to arid and semi-arid plateau regions of Inner Mongolia, China. Based on transcriptomic data, we isolated a high-affinity potassium transporter gene (RtHKT1) from R. trigyna, which encoded a plasma membrane-localized protein. RtHKT1 was rapidly up-regulated by high Na+ or low K+ and exhibited different tissue-specific expression patterns before and after stress treatment. Transgenic yeast showed tolerance to high Na+ or low K+, while transgenic Arabidopsis exhibited tolerance to high Na+ and sensitivity to high K+, or high Na+-low K+, confirming that Na+ tolerance in transgenic Arabidopsis depends on a sufficient external K+ concentration. Under external high Na+, high K+ and low K+ conditions, transgenic yeast accumulated more Na+-K+, Na+ and K+, while transgenic Arabidopsis accumulated less Na+-more K+, more Na+ and more Na+-K+, respectively, indicating that the ion transport properties of RtHKT1 depend on the external Na+-K+ environment. Salt stress induced up-regulation of some ion transporter genes (AtSOS1/AtHAK5/AtKUP5-6), as well as down-regulation of some genes (AtNHX1/AtAVP1/AtKUP9-12), revealing that multi-ion-transporter synergism maintains Na+/K+ homeostasis under salt stress in transgenic Arabidopsis. Overexpression of RtHKT1 enhanced K+ accumulation and prevented Na+ transport from roots to shoots, improved biomass accumulation and Chl content in salt-stressed transgenic Arabidopsis. The proline content and relative water content increased significantly, and some proline biosynthesis genes (AtP5CS1 and AtP5CS2) were also up-regulated in salt-stressed transgenic plants. These results suggest that RtHKT1 confers salt tolerance on transgenic Arabidopsis by maintaining Na+/K+ homeostasis and osmotic homeostasis.

Total Synthesis of an Atropisomer of the Schisandra Triterpenoid Schiglautone†A.

A diastereoselective approach for the total synthesis of an unusual atropisomer of the Schisandra triterpenoid (±)-schiglautone A is described. The efficient synthetic strategy features three key transformations: 1) two sequential titanium(III)-catalyzed radical cyclization/homologation reactions to construct the trans-fused [6,7] bicycle as well as install the quaternary carbons at C10 and C14 with the desired stereochemistry; 2) a Claisen rearrangement followed by a ring-closing metathesis to forge the strained nine-membered ring; and 3) a substrate-controlled Michael addition to enable the introduction of the C17 side-chain with good diastereoselectivity.

Diastereoselective Synthesis of the Hydroperoxide-Keto Form of (±)-Steenkrotin B.

A diastereoselective approach for the synthesis of the hydroperoxide-keto form of (±)-steenkrotin B (2') is described. The key features of the strategy involve a Diels-Alder cycloaddition, a titanium(III)-catalyzed reductive annulation, and a regio- and diastereoselective hydroperoxidation.

Expression of a Na+/H+ antiporter RtNHX1 from a recretohalophyte Reaumuria trigyna improved salt tolerance of transgenic Arabidopsis thaliana.

Reaumuria trigyna is an endangered recretohalophyte and a small xeric shrub that is endemic to the eastern Alxa and western Ordos areas of Inner Mongolia, China. Using transcriptome data, we identified a 1662-bp open reading frame encoding a 553-amino-acid protein corresponding to a Na+/H+ antiporter (RtNHX1) from R. trigyna. RtNHX1 was rapidly up-regulated by NaCl and exogenous abscisic acid treatment and had different tissue-specific expression patterns before and after salt-stress treatment. Overexpression of RtNHX1 enhanced seed germination, biomass accumulation, chlorophyll content, and root elongation in transgenic Arabidopsis plants under salt stress and rescued the salt-sensitive deficiencies of the nhx1 mutant. POD and CAT enzyme activities, proline content, and RWC all increased significantly in salt-stressed transgenic Arabidopsis plants, whereas MDA content did not. Additionally, there was a corresponding upregulation of some antioxidant-enzyme, proline biosynthesis and other stress responsive genes (AtPOD1, AtCAT1, AtP5CS1, AtP5CS2, AtRD29A, AtRD29B, AtKIN1, and AtABI2). The transgenic Arabidopsis plants accumulated more K+ and less Na+ in their leaves and had lower Na+/K+ ratios than WT plants. This was reflected in the upregulation of some ion transport-related genes (AtAVP1, AtSOS1, AtKUP6, and AtKUP8). When RtNHX1 was expressed in the AXT3 yeast strain, the accumulation of Na+ and K+ in the vacuole increased and the Na+/K+ ratio decreased. These results reveal that R. trigyna RtNHX1 is a functional antiporter that sequesters Na+ and K+ in the vacuole and could confer salt tolerance on transgenic Arabidopsis plants by maintaining Na+/K+ homeostasis and enhancing osmotic and antioxidant regulatory capacity. These results suggest that RtNHX1 may be a good target for improving salt tolerance in plants.

Small molecule-folic acid modification on nanopatterned PDMS and investigation on its surface property.

Folic acid (or folate, FA) has attracted considerable attention for cancer therapy. As one small molecule, its receptor (folate receptor, FR) is significantly overexpressed on the surface of many human tumor cells compared with normal cells. In this work, physical and chemical coupled modification method, that is the combination of nanoimprinting technique and graft polymerization, was adopted to modify FA on nanopatterned polydimethylsiloxane (PDMS) surface for possible application in micro-nanofluidic cytology. The surface property of differently treated PDMS was characterized by FTIR, AFM and contact angle measurement. AO/PI double staining, cell counting and MTT method were performed to examine the potential influence of FA modified nanopatterned PDMS on human cervical carcinoma (HeLa) cell behavior. Both FA modification and nanostructure have positive effect on the growth and viability of HeLa cells. It is the first time that the small molecule-folic acid was used to immobilize on the surface of PDMS in order to improve its surface property.