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The recretohalophyte Limonium bicolor thrives in high‐salinity environments because salt glands on the above‐ground parts of the plant help to expel excess salt. Here, we characterize a nucleus‐localized C3HC4 (RING‐HC)‐type zinc finger protein of L. bicolor named RING ZINC FINGER PROTEIN 1 (LbRZF1). LbRZF1 was expressed in salt glands and in response to NaCl treatment. LbRZF1 showed no E3 ubiquitin ligase activity. The phenotypes of overexpression and knockout lines for LbRZF1 indicated that LbRZF1 positively regulated salt gland development and salt tolerance in L. bicolor. lbrzf1 mutants had fewer salt glands and secreted less salt than did the wild‐type, whereas LbRZF1‐overexpressing lines had opposite phenotypes, in keeping with the overall salt tolerance of these plants. A yeast two‐hybrid screen revealed that LbRZF1 interacted with LbCATALASE2 (LbCAT2) and the transcription factor LbMYB113, leading to their stabilization. Silencing of LbCAT2 or LbMYB113 decreased salt gland density and salt tolerance. The heterologous expression of LbRZF1 in Arabidopsis thaliana conferred salt tolerance to this non‐halophyte. We also identified the transcription factor LbMYB48 as an upstream regulator of LbRZF1 transcription. The study of LbRZF1 in the regulation network of salt gland development also provides a good foundation for transforming crops and improving their salt resistance. [ABSTRACT FROM AUTHOR]

SUMMARY: Salt glands are the unique epidermal structures present in recretohalophytes, plants that actively excrete excess Na+ by salt secretory structures to avoid salt damage. Here, we describe a transmembrane protein that localizes to the plasma membrane of the recretohalophyte Limonium bicolor. As virus‐induced gene silencing of the corresponding gene LbRSG in L. bicolor decreased the number of salt glands, we named the gene Reduced Salt Gland. We detected LbRSG transcripts in salt glands by in situ hybridization and transient transformation. Overexpression and silencing of LbRSG in L. bicolor pointed to a positive role in salt gland development and salt secretion by interacting with Lb3G16832. Heterologous LbRSG expression in Arabidopsis enhanced salt tolerance during germination and the seedling stage by alleviating NaCl‐induced ion stress and osmotic stress after replacing or deleting the (highly) negatively charged region of extramembranous loop. After screened by immunoprecipitation‐mass spectrometry and verified using yeast two‐hybrid, PGK1 and BGLU18 were proposed to interact with LbRSG to strengthen salt tolerance. Therefore, we identified (highly) negatively charged regions in the extramembrane loop that may play an essential role in salt tolerance, offering hints about LbRSG function and its potential to confer salt resistance. Significance Statement: LbRSG (Reduced Salt Gland) was characterized as a transmembrane protein participating in salt gland development and salt resistance of recretohalophyte Limonium bicolor by interacting with Lb3G16832, and the negatively charged regions in the extramembranous loop were verified to play an essential role in salt tolerance of transgenic Arabidopsis. This study facilitated research on the molecular mechanisms of salt gland development and the design of salt‐tolerant crops for agricultural production in saline soils. [ABSTRACT FROM AUTHOR]

Halophytes complete their life cycles in saline environments. The recretohalophyte Limonium bicolor has evolved a specialized salt secretory structure, the salt gland, which excretes Na+ to avoid salt damage. Typical L. bicolor salt glands consist of 16 cells with four fluorescent foci and four secretory pores. Here, we describe a special type of salt gland at the base of the L. bicolor leaf petiole named bracelet salt glands due to their beaded‐bracelet‐like shape of blue auto‐fluorescence. Bracelet salt glands contain more than 16 cells and more than four secretory pores. Leaf disc secretion measurements and non‐invasive micro‐test techniques indicated that bracelet salt glands secrete more salt than normal salt glands, which helps maintain low Na+ levels at the leaf blade to protect the leaf. Cytokinin treatment induced bracelet salt gland differentiation, and the developed ones showed no further differentiation when traced with a living fluorescence microscopy imager, even though new salt gland development and leaf expansion were observed. Transcriptome revealed a NAC transcription factor gene that participates in bracelet salt gland development, as confirmed by its genome editing and overexpression in L. bicolor. These findings shed light on bracelet salt gland development and may facilitate the engineering of salt‐tolerant crops. [ABSTRACT FROM AUTHOR]

Soil salinization is one of the major factors restricting crop growth and agricultural production worldwide. Recretohalophytes have developed unique epidermal structures in their aboveground tissues, such as salt glands or salt bladders, to secrete excess salt out of the plant body as a protective mechanism from ion damage. Three hypotheses were proposed to explain how salt glands secrete salts: the osmotic hypothesis, a hypothesis similar to animal fluid transport, and vesicle-mediated exocytosis. However, there is no direct evidence to show whether the salt gland–secreted liquid contains landmark proteins or peptides which would elucidate the salt secretion mechanism. In this study, we collected the secreted liquid of salt glands from Limonium bicolor, followed by extraction and identification of its constituent proteins and peptides by SDS-PAGE and mass spectrometry. We detected 214 proteins and 440 polypeptides in the salt gland-secreted droplets of plants grown under control conditions. Unexpectedly, the proportion of energy metabolism-related proteins increased significantly though only 16 proteins and 35 polypeptides in the droplets of salt-treated plants were detected. In addition, vesicle transport proteins such as the Golgi marker enzyme glycosyltransferase were present in the secreted sap of salt glands from both control and salt-treated plants. These results suggest that trans-Golgi network-mediated vesicular transport and energy production contributes to salt secretion in salt glands. [ABSTRACT FROM AUTHOR]

With global increases in saline soil, it has become increasingly important to decipher salt-tolerance mechanisms and identify strategies to improve salt tolerance in crops. Halophytes complete their life cycles in environments containing ≥200 mM NaCl; these remarkable plants provide a potential source of genes for improving crop salt tolerance. Recretohalophytes such as Limonium bicolor have salt glands that secrete Na+ on their leaf epidermis. Here, we identified Lb1G04202, an uncharacterized gene with no conserved domains, from L. bicolor, which was highly expressed after NaCl treatment. We confirmed its expression in the salt gland by in situ hybridization, and then heterologously expressed Lb1G04202 in Arabidopsis thaliana. The transgenic lines had a higher germination rate, greater cotyledon growth percentage, and longer roots than the wild type (WT) under NaCl treatments (50, 100 and 150 mM). At the seedling stage, the transgenic lines grew better than the WT and had lower Na+ and malonyldialdehyde accumulation, and higher K+ and proline contents. This corresponded with the high expression of the key proline biosynthesis genes AtP5CS1 and AtP5CS2 under NaCl treatment. Isotonic mannitol treatment showed that Lb1G04202 overexpression significantly relieved osmotic stress. Therefore, this novel gene provides a potential target for improving salt tolerance. [ABSTRACT FROM AUTHOR]

Salinized soil is a major environmental stress affecting plant growth and development. Excessive salt in the soil inhibits the growth of most plants and even threatens their survival. Halophytes are plants that can grow and develop normally on saline-alkali soil due to salt tolerance mechanisms that emerged during evolution. For this reason, halophytes are used as pioneer plants for improving and utilizing saline land. Tamarisk, a family of woody halophytes, is highly salt tolerant and has high economic value. Understanding the mechanisms of salt tolerance in tamarisk and identifying the key genes involved are important for improving saline land and increasing the salt tolerance of crops. Here, we review recent advances in our understanding of the salt tolerance mechanisms of tamarisk and the economic and medicinal value of this halophyte. [ABSTRACT FROM AUTHOR]

Salt-resistant plants have different mechanisms to limit the deleterious effects of high salt in soil; for example, recretohalophytes secrete salt from unique structures called salt glands. Salt glands are the first differentiated epidermal structure of the recretohalophyte sea lavender (Limonium bicolor), followed by stomata and pavement cells. While salt glands and stomata develop prior to leaf expansion, it is not clear whether these steps are connected. Here, we explored the effects of the five phytohormones salicylic acid, brassinolide, methyl jasmonate, gibberellic acid, and abscisic acid on the development of the first expanded leaf of L. bicolor and its potential connection to salt gland, stomata, and pavement cell differentiation. We calculated the total number of salt glands, stomata, and pavement cells, as well as leaf area and pavement cell area, and assessed the correlations between these parameters. We detected strong and positive correlations between salt gland number and pavement cell area, between stomatal number and pavement cell area, and between salt gland number and stomatal number. We observed evidence of coupling between the development of salt glands, stomata, and pavement cells in L. bicolor , which lays the foundation for further investigation of the mechanism behind salt gland development. [ABSTRACT FROM AUTHOR]

Aims: This study aimed to better understanding the mechanism underlying the interaction between boron and NaCl and may provide methods to improve sugar beet production in saline soils. Methods: Sugar beet seedlings were treated with different concentrations of boron and NaCl. The flame photometer and ion chromatography were used to determine the ion content. And the uptake of Cl− by beet roots, was measured by non-invasive micro-test technology (NMT) and in vivo uptake system. The qPCR was used to measure the relative expression of the genes encoding ion transporters. Results: In this study, we demonstrate that application of 750 µM boron markedly mitigated the growth inhibition of sugar beet seedlings caused by 300 mM NaCl, as reflected in dry and fresh weight, leaf area, chlorophyll content, and net photosynthetic rates. Both Na+ and K+ in the plants were not affected by treatment with boron. However, boron markedly reduced the Cl− content of sugar beet seedlings. Boron profoundly upregulated the expression of CLC-b, CLC-c, CLC-f and downregulated the expression of NIP5, NIP6 and AKT1 under NaCl stress. Conclusions: Our results indicate that exogenous boron reduced Cl− content of sugar beet seedlings by regulation of Cl− transport under NaCl stress, which is possibly involved in upregulation of BvCLC-b, BvCLC-c and BvCLC-f genes encoding Cl− transporters in a cell wall-dependent Ca2+ signal transduction process. [ABSTRACT FROM AUTHOR]

Background: Identifying genes involved in salt tolerance in the recretohalophyte Limonium bicolor could facilitate the breeding of crops with enhanced salt tolerance. Here we cloned the previously uncharacterized gene LbHLH and explored its role in salt tolerance. Results: The 2,067-bp open reading frame of LbHLH encodes a 688-amino-acid protein with a typical helix-loop-helix (HLH) domain. In situ hybridization showed that LbHLH is expressed in salt glands of L. bicolor. LbHLH localizes to the nucleus, and LbHLH is highly expressed during salt gland development and in response to NaCl treatment. To further explore its function, we heterologously expressed LbHLH in Arabidopsis thaliana under the 35S promoter. The overexpression lines showed significantly increased trichome number and reduced root hair number. LbHLH might interact with GLABRA1 to influence trichome and root hair development, as revealed by yeast two-hybrid analysis. The transgenic lines showed higher germination percentages and longer roots than the wild type under NaCl treatment. Analysis of seedlings grown on medium containing sorbitol with the same osmotic pressure as 100 mM NaCl demonstrated that overexpressing LbHLH enhanced osmotic resistance. Conclusion: These results indicate that LbHLH enhances salt tolerance by reducing root hair development and enhancing osmotic resistance under NaCl stress. [ABSTRACT FROM AUTHOR]

Aims: To elucidate the genetics underlying salt tolerance in recretohalophytes and assess its relevance to non-halophytes, we cloned the Limonium bicolor homolog of Arabidopsis thaliana (Arabidopsis) SUPER SENSITIVE TO ABA AND DROUGHT2 (AtSAD2) and named it LbSAD2 , an importin-β gene associated with trichome initiation and reduced abscisic acid (ABA) sensitivity, and then we assessed the heterologously expressed LbSAD2 in Arabidopsis. Methods: We examined LbSAD2 expression and assessed the effect of heterologous LbSAD2 expression in Arabidopsis on root hair/trichome induction; the expression levels of possible related genes in trichome/root hair development; some physiological parameters involved in salt tolerance including germination rate, root length, and contents of Na+, proline, and malondialdehyde; and the response of ABA at the germination stage. Results: The LbSAD2 gene is highly expressed in the salt gland development stage and salt treatment, especially located in the salt gland by in situ hybridization, and the LbSAD2 protein contains some special domains compared with AtSAD2, which may suggest the involvement of LbSAD2 in salt tolerance. Compared with the SAD2/GL1 mutant CS65878, which lacks trichomes, CS65878-35S:LbSAD2 had higher trichome abundance but lower root hair abundance. Under 100 mM NaCl treatment, CS65878-35S:LbSAD2 showed enhanced germination and root lengths; improved physiological parameters, including high proline and low contents of Na+ and malondialdehyde; higher expression of the salt-tolerance genes Δ 1-PYRROLINE-5-CARBOXYLATE SYNTHETASE 1 (P5CS1) and GST CLASS TAU 5 (GSTU5); reduced ABA sensitivity; and increased expression of the ABA signaling genes RESPONSIVE TO ABA 18 (RAB18) and SNF1-RELATED PROTEIN KINASE 2 (SRK2E), but not of the ABA biosynthesis gene 9-CIS-EPOXYCAROTENOID DIOXYGENASE 3 (NCED3). Conclusion: LbSAD2 enhances salt tolerance in Arabidopsis by specifically reducing root hair development, Na+ accumulation, and ABA sensitivity. [ABSTRACT FROM AUTHOR]

Background: Salt, a common environmental stress factor, inhibits plant growth and reduces yields. Melatonin is a pleiotropic molecule that regulates plant growth and can alleviate environmental stress in plants. All previous research on this topic has focused on the use of melatonin to improve the relatively low salt tolerance of glycophytes by promoting growth and enhancing antioxidant ability. It is unclear whether exogenous melatonin can increase the salt tolerance of halophytes, particularly recretohalophytes, by enhancing salt secretion from the salt glands. Results: To examine the mechanisms of melatonin-mediated salt tolerance, we explored the effects of exogenous applications of melatonin on the secretion of salt from the salt glands of Limonium bicolor (a kind of recretohalophyte) seedlings and on the expression of associated genes. A pretreatment with 5 μM melatonin significantly improved the growth of L. bicolor seedlings under 300 mM NaCl. Furthermore, exogenous melatonin significantly increased the dry weight and endogenous melatonin content of L. bicolor. In addition, this treatment reduced the content of Na+ and Cl− in leaves, but increased the K+ content. Both the salt secretion rate of the salt glands and the expression level of genes encoding ion transporters (LbHTK1, LbSOS1, LbPMA, and LbNHX1) and vesicular transport proteins (LbVAMP721, LbVAP27, and LbVAMP12) were significantly increased by exogenous melatonin treatment. These results indicate that melatonin improves the salt tolerance of the recretohalophyte L. bicolor via the upregulation of salt secretion by the salt glands. Conclusions: Our results showed that melatonin can upregulate the expression of genes encoding ion transporters and vesicle transport proteins to enhance salt secretion from the salt glands. Combining the results of the current study with previous research, we formulated a novel mechanism by which melatonin increases salt secretion in L. bicolor. Ions in mesophyll cells are transported to the salt glands through ion transporters located at the plasma membrane. After the ions enter the salt glands, they are transported to the collecting chamber adjacent to the secretory pore through vesicle transport and ions transporter and then are secreted from the secretory pore of salt glands, which maintain ionic homeostasis in the cells and alleviate NaCl-induced growth inhibition. [ABSTRACT FROM AUTHOR]

Here, we studied the evolution of salt glands in 11 species of Tamarix and determined their role in adaptation to saline environments by measuring the effect of NaCl on plant growth and salt gland characteristics. Cluster analysis divided Tamarix species into three types (types I–III) according to salt‐gland characteristics. A phylogenetic tree based on ITS sequences indicated an evolutionary relationship consistent with the geographical distribution of Tamarix. We measured growth under different NaCl conditions (0, 100, 200, and 300 mM) for 40 days in three species (T. gallica, T. ramosissima, and T. laxa) representing the three Tamarix types. With increasing NaCl concentration, the biomass of all species was significantly reduced, especially that of T. gallica. Salt secretion ability and salt‐gland density showed similar trends in three types. The order of salt tolerance was type I > type II > type III. We conclude that during Tamarix adaptation to salinity, salt‐gland evolution followed two directions: one increasing salt‐gland density, and the other increasing salt secretion rate per salt‐gland. This study provides a basis for potential mechanisms of recretohalophyte adaptation to salinity. [ABSTRACT FROM AUTHOR]

The Arabidopsis thaliana WD40-repeat protein TRANSPARENT TESTA GLABRA1 (TTG1) controls epidermis development, playing opposite roles in trichome differentiation and root hair formation. We isolated and characterized LbTTG1 (encoding a WD40-repeat protein with high sequence similarity to TTG1) from the recretohalophyte Limonium bicolor , which actively excretes absorbed salt via a salt gland. The complete open reading frame of LbTTG1 was 1,095 bp, encoding a protein of 364 amino acids, and showed highest expression during the salt gland initiation stage. We heterologously expressed LbTTG1 in wild type and ttg1-13 Arabidopsis plants to verify the protein's function, and the copies of LbTTG1 were identified in transgenic strains using southern blotting. Trichomes were extremely induced on the first true leaves of plants heterologously expressing LbTTG1 , whereas no trichomes were produced by ttg1-13 plants. Conversely, plants heterologously expressing LbTTG1 produced fewer root hairs than ttg1-13 plants. In plants heterologously expressing LbTTG1 compared to controls, epidermis differentiation genes (GLABRA1 and GLABRA3) were up-regulated while genes encoding negative regulators of trichome development (TRIPTYCHON and CAPRICE) were down-regulated. Under increased NaCl concentrations, both of the transgenic lines showed enhanced germination and root length, and accumulated less malondialdehyde (MDA) and Na+ and produced more proline, soluble sugar, and higher glutathione S-transferase activity, compared with the ttg1-13 mutant. These results indicate that LbTTG1 participates in epidermis development in Arabidopsis , similarly to other WD40-repeat proteins, and specifically increases salt tolerance of transgenic Arabidopsis by reducing ion accumulation and increasing osmolyte levels. [ABSTRACT FROM AUTHOR]

The effect of NaCl on reproductive development was investigated in euhalophyte Suaeda salsa L. under controlled conditions. Results showed that NaCl promoted the reproductive growth of S. salsa and 200mM NaCl was optimal. This was reflected in the increases of seed yield, seed number, flower number per plant and leaf axil, 1000 seed weight, as well as a decrease in flower abortion percentage with supply of NaCl. NaCl reduced the flower abortion percentage by increasing stigma receptivity instead of pollen viability. The Na+ and Cl- concentration in petals, stems and leaves were increased significantly but slightly in stamen and pistil. In contrast, the K+ concentration decreased markedly in leaves, stems and petals but a little in stamen and pistil. The Na+ and Cl- concentrations also increased significantly in seed from mother plants exposed to NaCl, whereas K+ decreased. However, seed quality was not influenced. Our results showed that high concentration of NaCl markedly increases the seed number and quality of S. salsa primarily via increasing flower number and fertility and S. salsa develops strategy to maintain ion homeostasis in reproductive organs for the generation. These factors play a pivotal role in setting up plant populations in saline environment. [ABSTRACT FROM AUTHOR]

Soil salinization is a serious and growing problem around the world. Some plants, recognized as the recretohalophytes, can normally grow on saline–alkali soil without adverse effects by secreting excessive salt out of the body. The elucidation of the salt secretion process is of great significance for understanding the salt tolerance mechanism adopted by the recretohalophytes. Between the 1950s and the 1970s, three hypotheses, including the osmotic potential hypothesis, the transfer system similar to liquid flow in animals, and vesicle-mediated exocytosis, were proposed to explain the salt secretion process of plant salt glands. More recently, increasing evidence has indicated that vesicular transport plays vital roles in salt secretion of recretohalophytes. Here, we summarize recent findings, especially regarding the molecular evidence on the functional roles of vesicular trafficking in the salt secretion process of plant salt glands. A model of salt secretion in salt gland is also proposed. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) plays an important role in regulating plant stress responses. However, there are currently no reports on ABA in regulating the salt tolerance of Limonium bicolor. Here, we analyzed the effects of exogenous ABA on growth, photosynthesis, salt secretion, and physiological and transcriptome changes of L. bicolor seedlings under 300 mM NaCl. Exogenous 10 µM ABA treatment in L. bicolor seedlings treated with salt upregulated the expression of genes related to stomatal development, increased the density and number of stomata, and improved plant gas exchange capacity, thereby increasing the net photosynthesis rate. Exogenous ABA treatment in L. bicolor seedlings under salt stress also upregulated the expression of genes related to salt gland development, promoted salt glands development, increased salt glands density, increased the salt secretion capacity of leaves, and reduced the accumulation of Na+ in the plant. ABA application enhanced salt tolerance of L. bicolor seedlings by increasing content of soluble solutes, enhancing antioxidant enzymes activities, decreasing accumulation of reactive oxygen species, and increasing ABA and indole acetic acid contents. Transcriptome sequencing revealed that genes differentially expressed in L. bicolor seedlings during ABA-induced salt tolerance are involved in many metabolic pathways, including plant hormone signal transduction, proline metabolism, endocytosis and protein processing in the endoplasmic reticulum. Furthermore, some transcription factors, MYB, bHLH, and WRKY, might contribute to the improved salt stress tolerance of L. bicolor conferred by ABA. These results provide a basis for further clarifying the salt tolerance mechanism mediated by exogenous ABA in L. bicolor seedlings. [Display omitted] • Exogenous ABA significantly increased salt tolerance of Limonium bicolor. • Exogenous ABA improved photosynthesis via regulating stomatal development. • Exogenous ABA increased leaves Na+ secretion via regulating salt gland development. [ABSTRACT FROM AUTHOR]

• AtSIZ1 is a salt-induced C2H2 type zinc finger protein. • AtSIZ1 positively improves salt tolerance of Arabidopsis. • AtSIZ1 enhances salt tolerance in Arabidopsis by maintaining ionic and osmotic homeostasis. C2H2-type zinc finger proteins play important roles in plant growth, development, and abiotic stress tolerance. Here, we explored the role of the C2H2-type zinc finger protein SALT INDUCED ZINC FINGER PROTEIN1 (AtSIZ1; At3G25910) in Arabidopsis thaliana under salt stress. AtSIZ1 expression was induced by salt treatment. During the germination stage, the germination rate, germination energy, germination index, cotyledon growth rate, and root length were significantly higher in AtSIZ1 overexpression lines than in the wild type under various stress treatments, whereas these indices were significantly reduced in AtSIZ1 loss-of-function mutants. At the mature seedling stage, the overexpression lines maintained higher levels of K+, proline, and soluble sugar, lower levels of Na+ and MDA, and lower Na+/K+ ratios than the wild type. Stress-related marker genes such as SOS1 , AtP5CS1 , AtGSTU5 , COR15A , RD29A , and RD29B were expressed at higher levels in the overexpression lines than the wild type and loss-of-function mutants under salt treatment. These results indicate that AtSIZ1 improves salt tolerance in Arabidopsis by helping plants maintain ionic homeostasis and osmotic balance. [ABSTRACT FROM AUTHOR]

Halophytes are organisms that can grow and reproduce in high-salinity environments, making them an important resource for identifying and studying salt-tolerant genes. As the acreage of saline soils increases worldwide, it is becoming more urgent to understand and exploit the mechanism(s) of salt tolerance from halophytes, with the goal of improving the salt tolerance of crops. Limonium bicolor is a halophyte in the Plumbaginaceae family that presents salt glands on its leaf epidermis that secrete excess Na+. Here, we show that the LbCPC gene encodes a protein that participates in salt gland development and salt secretion. Transgenic lines overexpressing LbCPC showed inhibited salt gland development and decreased salt gland density, whereas lines with LbCPC silenced by virus-induced gene silencing (VIGS) showed enhanced salt gland development and an increase in the number of salt glands and the amount of salt secretion by the plant, demonstrating that LbCPC has an negative effect on salt gland development of L. bicolor. We further investigated the role of LbCPC in salt tolerance by generating transgenic lines in Arabidopsis thaliana. Compared to normal wild type, transgenic Arabidopsis lines overexpressing LbCPC had fewer trichomes and more root hairs. As the NaCl concentration increased, so did the salt sensitivity of these transgenic lines, as evidenced by inhibited germination and root elongation, which may be due to lowered expression of salt-tolerance marker genes. Together, we show that LbCPC negatively regulates the development of salt glands and inhibits the production of salt glands, providing a new avenue to screen for genes promoting salt resistance. • LbCPC gene participates in salt gland development of Limonium bicolor. • LbCPC is involved in salt secretion and salt tolerance of Limonium bicolor. • Transgenic Arabidopsis lines overexpressing LbCPC showed more salt sensitive than the wild type. [ABSTRACT FROM AUTHOR]

Abstract: The strategy of Suaeda salsa adaptation to high salinity was shown in this study. Our results firstly demonstrated that Na+ was a beneficial element rather than toxic ion for S. salsa plants. The biomass of S. salsa seedlings increased significantly after salt-treatment (100–400mmol/L NaCl). Secondly, S. salsa had strong ability to accumulate Na+ under saline condition. Ninety-four percent of Na+ absorbed by salt-treated S. salsa plant accumulated in the shoot, especially in the leaves, and 98% of Na+ in the leaves accumulated in leaf symplast. Thirdly, measurement on isolated tonoplast-enriched membrane vesicles derived from the leaves of S. salsa showed that increased V-H+-ATPase hydrolytic activity, V-H+-ATPase proton pump activity and the tonoplast Na+/H+ antiport activity were found in salt-treated leaves compared with the control leaves. Up-regulation of the tonoplast Na+/H+ antiport activity was due to increase of both transcription and translation. These results suggested that coordinate up-regulation of V-H+-ATPase activity and Na+/H+ antiporter activity play a pivotal role in Na+ sequestering into vacuoles of S. salsa. [Copyright &y& Elsevier]

• Limonium bicolor is considered an excellent halophyte to transform saline soil. • LbTRY expression can significantly increase the sensitivity of Arabidopsis. • High development of root hairs and osmotic homeostasis is responsible. Arabidopsis thaliana TRY is a negative regulator of trichome differentiation that promotes root hair differentiation. Here, we established that LbTRY, from the recretohalophyte Limonium bicolor , is a typical MYB transcription factor that exhibits transcriptional activation activity and locates in nucleus. By in situ hybridization in L. bicolor , LbTRY may be specifically positioned in salt gland of the expanded leaves. LbTRY expression was the highest in mature leaves and lowest under NaCl treatment. For functional assessment, we heterologously expressed LbTRY in wild-type and try29760 mutant Arabidopsis plants. Epidermal differentiation was remarkably affected in the transgenic wild-type line, as was increased root hair development. Complementation of try29760 with LbTRY under both 35S and LbTRY specific promoter restored the wild-type phenotype. qRT-PCR analysis suggested that AtGL3 and AtZFP5 promote root hair cell fate in lines heterologously producing LbTRY. In addition, four genes (AtRHD6 , AtRSL1 , AtLRL2 , and AtLRL3) involved in root hair initiation and elongation were upregulated in the transgenic lines. Furthermore, LbTRY specifically increased the salt sensitivity of the transgenic lines. The transgenic and complementation lines showed poor germination rates and reduced root lengths, whereas the mutant unexpectedly fared the best under a range of NaCl treatments. Under salt stress, the transgenic seedlings accumulated more MDA and Na+ and less proline and soluble sugar than try29760. Thus, when heterologously expressed in Arabidopsis, LbTRY participates in hair development, similar to other MYB proteins, and specifically reduces salt tolerance by increasing ion accumulation and reducing osmolytes. The expression of salt-tolerance marker genes (SOS1 , SOS2 , SOS3 and P5CS1) was significant reduced in the transgenic lines. More will be carried by downregulating expression of TRY homologs in crops to improve salt tolerance. [ABSTRACT FROM AUTHOR]

• Salt secretion of L. bicolor salt glands is inhibited after treatment with BFA. • LbSYP61 is significantly upregulated by 200 mM NaCl treatment. • After silencing of LbSYP61 , the salt secretion is obviously reduced. • Vesicular transport participates in salt secretion of salt glands. Soil salinization is a global problem that inhibits plant growth and limits crop production. Recretohalophyte plants secrete excess salt via salt glands, thus avoiding excessive accumulation of salt within their cells and protecting themselves from salt-associated damage. However, the molecular mechanisms underlying salt secretion via salt glands are unclear. Here, we used brefeldin A (BFA), a specific inhibitor of Golgi-mediated cell secretion, to test whether vesicle transport participates in salt secretion in the recretohalophyte Limonium bicolor using leaf disc and non-invasive micrometric assays. Indeed, salt secretion via salt glands was significantly inhibited, the Golgi apparatus in the salt gland was damaged, and acidic phosphatase activity was significantly inhibited following a 200 μg/ml BFA treatment. To further examine the role of vesicle transport in salt secretion, we used virus-induced gene silencing (VIGS) to downregulate the gene encoding a trans Golgi-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein, LbSYP61. After silencing LbSYP61 , the ability of L. bicolor to secrete salt via salt glands was also significantly reduced. These results provide direct evidence that vesicular transport participates in salt secretion via salt glands and the vesicular transport-related protein LbSYP61 plays an important role in this process. [ABSTRACT FROM AUTHOR]

Salt glands are specialized epidermal salt secretion structures on the surfaces of recretohalophytes leaves. In wild-type plants of the recretohalophyte Limonium bicolor (Bag.) Kuntze, salt glands comprise 16 cells each and appear as a blue ring under fluorescence microscopy, with four strong autofluorescence points. Here, we describe the lb23 mutant, which showed four additional types of salt glands, with five, six, seven and eight strong autofluorescence points. These salt glands comprised 20, 24, 28 and 32 cells, respectively, with correspondingly increased numbers of secretory, accessory, inner cup and outer cup cells. The cell arrangement and cell types within the salt glands in the mutant were unaltered from those of the wild type. For example, salt glands with five strong autofluorescence points had five secretory cells, five accessory cells, five inner cup cells and five outer cup cells. Compared with wild type, the abnormal salt glands with five, six, seven and eight strong autofluorescence points showed higher salt secretion rates per salt gland. Thus, the increase in salt gland cell numbers resulted in markedly increased salt secretion. This L. bicolor mutant provides an important resource for research on the molecular mechanism of salt gland development. [ABSTRACT FROM AUTHOR]