NADPH oxidase-dependent H2O2 production mediates salicylic acid-induced salt tolerance in mangrove plant Kandelia obovata by regulating Na+/K+ and redox homeostasis.

Salicylic acid (SA) is known to enhance salt tolerance in plants. However, the mechanism of SA-mediated response to high salinity in halophyte remains unclear. Using electrophysiological and molecular biological methods, we investigated the role of SA in response to high salinity in mangrove species, Kandelia obovata, a typical halophyte. Exposure of K. obovata roots to high salinity resulted in a rapid increase in endogenous SA produced by phenylalanine ammonia lyase pathway. The application of exogenous SA improved the salt tolerance of K. obovata, which depended on the NADPH oxidase-mediated H2O2. Exogenous SA and H2O2 increased Na+ efflux and reduced K+ loss by regulating the transcription levels of Na+ and K+ transport-related genes, thus reducing the Na+/K+ ratio in the salt-treated K. obovata roots. In addition, exogenous SA-enhanced antioxidant enzyme activity and its transcripts, and the expressions of four genes related to AsA-GSH cycle as well, then alleviated oxidative damages in the salt-treated K. obovata roots. However, the above effects of SA could be reversed by diphenyleneiodonium chloride (the NADPH oxidase inhibitor) and paclobutrazol (a SA biosynthesis inhibitor). Collectively, our results demonstrated that SA-induced salt tolerance of K. obovata depends on NADPH oxidase-generated H2O2 that affects Na+/K+ and redox homeostasis in response to high salinity.

PlantC2U: deep learning of cross-species sequence landscapes predicts plastid C-to-U RNA editing in plants.

In plants, C-to-U RNA editing mainly occurs in plastid and mitochondrial transcripts, which contributes to a complex transcriptional regulatory network. More evidence reveals that RNA editing plays critical roles in plant growth and development. However, accurate detection of RNA editing sites using transcriptome sequencing data alone is still challenging. In the present study, we develop PlantC2U, which is a convolutional neural network, to predict plastid C-to-U RNA editing based on the genomic sequence. PlantC2U achieves >95% sensitivity and 99% specificity, which outperforms the PREPACT tool, random forests, and support vector machines. PlantC2U not only further checks RNA editing sites from transcriptome data to reduce possible false positives, but also assesses the effect of different mutations on C-to-U RNA editing based on the flanking sequences. Moreover, we found the patterns of tissue-specific RNA editing in the mangrove plant Kandelia obovata, and observed reduced C-to-U RNA editing rates in the cold stress response of K. obovata, suggesting their potential regulatory roles in plant stress adaptation. In addition, we present RNAeditDB, available online at https://jasonxu.shinyapps.io/RNAeditDB/. Together, PlantC2U and RNAeditDB will help researchers explore the RNA editing events in plants and thus will be of broad utility for the plant research community.

Structural, developmental and functional analyses of leaf salt glands of mangrove recretohalophyte Aegiceras corniculatum.

Salt secretion is an important strategy used by the mangrove plant Aegiceras corniculatum to adapt to the coastal intertidal environment. However, the structural, developmental and functional analyses on the leaf salt glands, particularly the salt secretion mechanism, are not well documented. In this study, we investigated the structural, developmental and degenerative characteristics and the salt secretion mechanisms of salt glands to further elucidate the mechanisms of salt tolerance of A. corniculatum. The results showed that the salt gland cells have a large number of mitochondria and vesicles, and plenty of plasmodesmata as well, while chloroplasts were found in the collecting cells. The salt glands developed early and began to differentiate at the leaf primordium stage. We observed and defined three stages of salt gland degradation for the first time in A. corniculatum, where the secretory cells gradually twisted and wrinkled inward and collapsed downward as the salt gland degeneration increased and the intensity of salt gland autofluorescence gradually diminished. In addition, we found that the salt secretion rate of the salt glands increased when the treated concentration of NaCl increased, reaching the maximum at 400 mM NaCl. The salt-secreting capacity of the salt glands of the adaxial epidermis is significantly greater than that of the abaxial epidermis. The real-time quantitative PCR results indicate that SAD2, TTG1, GL2 and RBR1 may be involved in regulating the development of the salt glands of A. corniculatum. Moreover, Na+/H+ antiporter, H+-ATPase, K+ channel and Cl- channel may play important roles in the salt secretion of salt glands. In sum mary, this study strengthens the understanding of the structural, developmental and degenerative patterns of salt glands and salt secretion mechanisms in mangrove recretohalophyte A. corniculatum, providing an important reference for further studies at the molecular level.

Inventory of cadmium-transporter genes in the root of mangrove plant Avicennia marina under cadmium stress.

Mangrove Avicennia marina has the importantly potential for cadmium (Cd) pollution remediation in coastal wetlands. Unfortunately, the molecular mechanisms and transporter members for Cd uptake by the roots of A. marina are not well documented. In this study, photosynthetic and phenotypic analysis indicated that A. marina is particularly tolerant to Cd. The content and flux analysis indicated that Cd is mainly retained in the roots, with greater Cd influx in fine roots than that in coarse roots, and higher Cd influx in the root meristem zone as well. Using transcriptomic analysis, a total of 5238 differentially expressed genes were identified between the Cd treatment and control group. Moreover, we found that 54 genes were responsible for inorganic ion transport. Among these genes, AmHMA2, AmIRT1, and AmPCR2 were localized in the plasma membrane and AmZIP1 was localized in both plasma membrane and cytoplasm. All above gene encoding transporters showed significant Cd transport activities using function assay in yeast cells. In addition, the overexpression of AmZIP1 or AmPCR2 in Arabidopsis improved the Cd tolerance of transgenic plants. This is particularly significant as it provides insight into the molecular mechanism for Cd uptake by the roots of mangrove plants and a theoretical basis for coastal wetland phytoremediation.

Integration of eQTL and GWAS analysis uncovers a genetic regulation of natural ionomic variation in Arabidopsis.

KEY MESSAGE: This study provided important insights into the genetic architecture of variations in A. thaliana leaf ionome in a cell-type-specific manner. The functional interpretation of traits associated variants by expression quantitative trait loci (eQTL) analysis is usually performed in bulk tissue samples. While the regulation of gene expression is context-dependent, such as cell-type-specific manner. In this study, we estimated cell-type abundances from 728 bulk tissue samples using single-cell RNA-sequencing dataset, and performed cis-eQTL mapping to identify cell-type-interaction eQTL (cis-eQTLs(ci)) in A. thaliana. Also, we performed Genome-wide association studies (GWAS) analyses for 999 accessions to identify the genetic basis of variations in A. thaliana leaf ionome. As a result, a total of 5,664 unique eQTL genes and 15,038 unique cis-eQTLs(ci) were significant. The majority (62.83%) of cis-eQTLs(ci) were cell-type-specific eQTLs. Using colocalization, we uncovered one interested gene AT2G25590 in Phloem cell, encoding a kind of plant Tudor-like protein with possible chromatin-associated functions, which colocalized with the most significant cis-eQTL(ci) of a Mo-related locus (Chr2:10,908,806:A:C; P = 3.27 × 10-27). Furthermore, we prioritized eight target genes associated with AT2G25590, which were previously reported in regulating the concentration of Mo element in A. thaliana. This study revealed the genetic regulation of ionomic variations and provided a foundation for further studies on molecular mechanisms of genetic variants controlling the A. thaliana ionome.

Proteomic analysis reveals differential responsive mechanisms in Solanum nigrum exposed to low and high dose of cadmium.

Cadmium (Cd) contamination is becoming a widespread environmental problem. However, the differential responsive mechanisms of Cd hyperaccumulator Solanum nigrum to low or high dose of Cd are not well documented. In this study, phenotypic and physiological analysis firstly suggested that the seedlings of S. nigrum showed slight leaf chlorosis symptoms under 25 µM Cd and severe inhibition on growth and photosynthesis under 100 µM Cd. Further proteomic analysis identified 105 differentially expressed proteins (DEPs) in the Cd-treated leaves. Under low dose of Cd stress, 47 DEPs are mainly involved in primary metabolic processes, while under high dose of Cd stress, 92 DEPs are mainly involved in photosynthesis, energy metabolism, production of phytochelatin and reactive oxygen species (ROS). Protein-protein interaction (PPI) network analysis of DEPs support above differential responses in the leaves of S. nigrum to low and high dose of Cd treatments. This work provides the differential responsive mechanisms in S. nigrum to low and high dose of Cd, and the theoretical foundation for the application of hyperaccumulating plants in the phytoremediation of Cd-contaminated soils.

Hydrogen sulfide upregulates the alternative respiratory pathway in mangrove plant Avicennia marina to attenuate waterlogging-induced oxidative stress and mitochondrial damage in a calcium-dependent manner.

Hydrogen sulfide (H2 S) is considered to mediate plant growth and development. However, whether H2 S regulates the adaptation of mangrove plant to intertidal flooding habitats is not well understood. In this study, sodium hydrosulfide (NaHS) was used as an H2 S donor to investigate the effect of H2 S on the responses of mangrove plant Avicennia marina to waterlogging. The results showed that 24-h waterlogging increased reactive oxygen species (ROS) and cell death in roots. Excessive mitochondrial ROS accumulation is highly oxidative and leads to mitochondrial structural and functional damage. However, the application of NaHS counteracted the oxidative damage caused by waterlogging. The mitochondrial ROS production was reduced by H2 S through increasing the expressions of the alternative oxidase genes and increasing the proportion of alternative respiratory pathway in the total mitochondrial respiration. Secondly, H2 S enhanced the capacity of the antioxidant system. Meanwhile, H2 S induced Ca2+ influx and activated the expression of intracellular Ca2+ -sensing-related genes. In addition, the alleviating effect of H2 S on waterlogging can be reversed by Ca2+ chelator and Ca2+ channel blockers. In conclusion, this study provides the first evidence to explain the role of H2 S in waterlogging adaptation in mangrove plants from the mitochondrial aspect.

Cadmium promotes the absorption of ammonium in hyperaccumulator Solanum nigrum L. mediated by ammonium transporters and aquaporins.

Cadmium (Cd) is a toxic heavy metal affecting the normal growth of plants. Nitrate (NO3-) and ammonium (NH4+) are the primary forms of inorganic nitrogen (N) absorbed by plants. However, the mechanism of N absorption and regulation under Cd stress remains unclear. This study found that: (1) Cd treatment affected the biomass, root length, and Cd2+ flux in Solanum nigrum seedling roots. Specifically, 50 µM Cd significantly inhibited NO3- influx while increased NH4+ influx compared with 0 and 5 µM Cd treatments measured by non-invasive micro-test technology. (2) qRT-PCR analysis showed that 50 µM Cd inhibited the expressions of nitrate transporter genes, SnNRT2;4 and SnNRT2;4-like, increased the expressions of ammonium transporter genes, SnAMT1;2 and SnAMT1;3, in the roots. (3) Under NH4+ supply, 50 µM Cd significantly induced the expressions of the aquaporin genes, SnPIP1;5, SnPIP2;7, and SnTIP2;1. Our results showed that 50 µM Cd stress promoted NH4+ absorption by up-regulating the gene expressions of NH4+ transporter and aquaporins, suggesting that high Cd stress can affect the preference of N nutrition in S. nigrum.

Exogenous hydrogen sulfide mediates Na+ and K+ fluxes of salt gland in salt-secreting mangrove plant Avicennia marina.

Hydrogen sulfide (H2S), is a crucial biological player in plants. Here, we primarily explored the interaction between sodium hydrosulfide (NaHS, a H2S donor) and the fluxes of Na+ and K+ from the salt glands of mangrove species Avicennia marina (Forsk.) Vierh. with non-invasive micro-test technology (NMT) and quantitative real-time PCR (qRT-PCR) approaches under salinity treatments. The results showed that under 400-mM NaCl treatment, the addition of 200-µM NaHS markedly increased the quantity of salt crystals in the adaxial epidermis of A. marina leaves, accompanied by an increase in the K+/Na+ ratio. Meanwhile, the endogenous content of H2S was dramatically elevated in this process. The NMT result revealed that the Na+ efflux was increased from salt glands, whereas K+ efflux was decreased with NaHS application. On the contrary, the effects of NaHS were reversed by H2S scavenger hypotaurine (HT), and DL-propargylglycine (PAG), an inhibitor of cystathionine-γ-lyase (CES, a H2S synthase). Moreover, enzymic assay revealed that NaHS increased the activities of plasma membrane and tonoplast H+-ATPase. qRT-PCR analysis revealed that NaHS significantly increased the genes transcript levels of tonoplast Na+/H+ antiporter (NHX1), plasma membrane Na+/H+ antiporter (SOS1), plasma membrane H+-ATPase (AHA1) and tonoplast H+-ATPase subunit c (VHA-c1), while suppressed above-mentioned gene expressions by the application of HT and PAG. Overall, H2S promotes Na+ secretion from the salt glands of A. marina by up-regulating the plasma membrane and tonoplast Na+/H+ antiporter and H+-ATPase.

Ammonium has stronger Cd detoxification ability than nitrate by reducing Cd influx and increasing Cd fixation in Solanum nigrum L.

Cadmium (Cd) is a harmful heavy metal that affects the growth and development of plants. Nitrogen (N) is an essential nutrient for plants, and appropriate N management can improve Cd tolerance. The aim of our study was to explore the effects of different forms of N on the molecular and physiological responses of the hyperaccumulator Solanum nigrum to Cd toxicity. Measurement of biomass, photosynthetic parameters, and Cd2+ fluxes using non-invasive micro-test technique, Cd fluorescent dying, biochemical methods and quantitative real-time PCR analysis were performed in our study. Our results showed that ammonium (NH4+) has stronger Cd detoxification ability than nitrate (NO3-), which are likely attributed to the following three reasons: (1) NH4+ decreased the influx and accumulation of Cd2+ by regulating the transcription of Cd transport-related genes; (2) the ameliorative effects of NH4+ were accompanied by the increased retention of Cd in the cell walls of roots; and (3) NH4+ up-regulated SnExp expression.