Integrated metabolomic and metagenomic strategies shed light on interactions among planting environments, rhizosphere microbiota, and metabolites of tobacco in Yunnan, China.

Changes in climatic factors and rhizosphere microbiota led plants to adjust their metabolic strategies for survival under adverse environmental conditions. Changes in plant metabolites can mediate crop growth and development and interact with rhizosphere microbiota of the plant rhizosphere. To understand the interactions among environmental factors, rhizosphere microbiota, and metabolites of tobacco, a study was conducted by using integrated metagenomic and metabolomic strategies at four typical representative tobacco planting sites in Yunnan, China. The results showed that the agronomical and biochemical traits were significantly affected by temperature, precipitation (PREP), soil pH, and altitude. Correlation analyses revealed a significant positive correlation of temperature with length, width, and area of the leaf, while PREP correlated with plant height and effective leaf numbers. Furthermore, total sugar and reducing sugar contents of baked leaves were significantly higher, while the total nitrogen and total alkaloid levels were lower in tobacco leaves at site with low PREP. A total of 770 metabolites were detected with the highest number of different abundant metabolites (DMs) at Chuxiong (CX) with low PREP as compared to the other three sites, in which secondary metabolites were more abundant in both leaves and roots of tobacco. A total of 8,479 species, belonging to 2,094 genera with 420 individual bins (including 13 higher-quality bins) harboring 851,209 CDSs were detected. The phyla levels of microorganisms such as Euryarchaeota, Myxococcota, and Deinococcota were significantly enriched at the CX site, while Pseudomonadota was enriched at the high-temperature site with good PREP. The correlation analyses showed that the metabolic compounds in low-PREP site samples were positively correlated with Diaminobutyricimonas, Nissabacter, Alloactinosynnema, and Catellatospora and negatively correlated with Amniculibacterium, Nordella, Noviherbaspirillum, and Limnobacter, suggesting that the recruitment of Diaminobutyricimonas, Nissabacter, Alloactinosynnema, and Catellatospora in the rhizosphere induces the production and accumulation of secondary metabolites (SMs) (e.g., nitrogen compounds, terpenoids, and phenolics) for increasing drought tolerance with an unknown mechanism. The results of this study may promote the production and application of microbial fertilizers and agents such as Diaminobutyricimonas and Alloactinosynnema to assemble synthetic microbiota community or using their gene resources for better cultivation of tobacco as well as other crops in drought environments.

Unexpected omega-3 activities in intracellular lipolysis and macrophage foaming revealed by fluorescence lifetime imaging.

Omega-3 polyunsaturated fatty acids (PUFA) found primarily in fish oil have been a popular supplement for cardiovascular health because they can substantially reduce circulating triglyceride levels in the bloodstream to prevent atherosclerosis. Beyond this established extracellular activity, here, we report a mode of action of PUFA, regulating intracellular triglyceride metabolism and lipid droplet (LD) dynamics. Real-time imaging of the subtle and highly dynamic changes of intracellular lipid metabolism was enabled by a fluorescence lifetime probe that addressed the limitations of intensity-based fluorescence quantifications. Surprisingly, we found that among omega-3 PUFA, only docosahexaenoic acid (DHA) promoted the lipolysis in LDs and reduced the overall fat content by approximately 50%, and consequently helped suppress macrophage differentiation into foam cells, one of the early steps responsible for atherosclerosis. Eicosapentaenoic acid, another omega-3 FA in fish oil, however, counteracted the beneficial effects of DHA on lipolysis promotion and cell foaming prevention. These in vitro findings warrant future validation in vivo.

Systematic review and meta-analysis of current evidence in endograft therapy for spontaneous isolated superior mesenteric artery dissection.

The role of endovascular stent therapy (EST) in the treatment of spontaneous isolated superior mesenteric artery dissection (SISMAD) has gained momentum in recent years but remains controversial. We gathered research examining the advantages and disadvantages of EST for SISMAD patients. Primary outcomes involved both immediate and long-term results. Random or fixed effect models were used for effect size (ES) calculation with 95% confidence interval (CI) based on 50% heterogeneity threshold. Our analysis incorporated data from 21 studies including 611 SISMAD cases treated by EST. Our findings show a complication rate of approximately 1% following EST (95%CI 0.01-0.02, I2 = 0%, P = 0.97), with a bare minimum mortality rate of < 1% (95%CI 0.00-0.01, I2 = 0%, P > 0.05) and a reintervention rate of < 1% (95%CI 0.00-0.01, I2 = 0%, P = 0.89). We also found technique success and symptom resolution approaching 94% and 99%, respectively, in the immediate postoperative phase. In the long run, we observed a recurrence of symptoms at 3% (95%CI 0.00-0.06, I2 = 58.6%, P < 0.01), creation of new dissections at 1% (95%CI 0.00-0.02, I2 = 0%, P = 0.73), aneurysm progression at 2% (95%CI 0.00-0.03, I2 = 42.7%, P = 0.12), reintervention due to complications at 3% (95%CI 0.00-0.05, I2 = 0%, P = 0.43) and stenotic stents at 12% (95%CI 0.04-0.23, I2 = 77.5%, P < 0.01). Nevertheless, high levels of stent patency 98% (95% CI 0.97-1.00, I2 = 0%, P = 0.51) and complete remodeling 88% (95% CI 0.82-0.94, I2 = 65.5%, P < 0.05) were observed postoperatively. Overall, EST presents minimal complications and promising long-term outcomes for SISMAD, although the prevalence of stent stenosis requires further attention.

Tumor-Targeted Fluorescent/Photoacoustic Imaging of Legumain Activity In Vivo.

Legumain has been identified as a target for diagnosis and treatment of associated cancers. Therefore, real-time imaging of legumain activity in vivo is helpful in diagnosing and evaluating therapeutic efficacy of associated cancers. Fluorescent/photoacoustic (FL/PA) dual-modal imaging developed rapidly because of its good sensitivity and spatial resolution. As far as we know, a tumor-targeted probe for FL/PA imaging of legumain activity in vivo has not been reported. Hence, we intended to develop a tumor-targeted hemicyanine (HCy) probe (HCy-AAN-Bio) for FL/PA imaging of legumain in vivo. The control probe HCy-AAN does not have tumor-targeting ability. Legumain can specifically cleave HCy-AAN-Bio or HCy-AAN with the generation of FL/PA signal while more HCy-AAN-Bio could be recognized by legumain than HCy-AAN with higher sensitivity in vitro. Due to the tumor-targeting ability, HCy-AAN-Bio could image 4T1 cells with an additional 1.3-fold FL enhancement and 1.9-fold PA enhancement than HCy-AAN. In addition, HCy-AAN-Bio could image legumain activity in vivo with an additional 1.5-fold FL enhancement and 1.9-fold PA enhancement than HCy-AAN. We expected that HCy-AAN-Bio will be a powerful tool for early diagnosis of associated cancer.

Caspase-3-Responsive Fluorescent/Photoacoustic Imaging of Tumor Apoptosis.

Caspase-3 is an essential executor in apoptosis, and its activation has been regarded as a biomarker of cell apoptosis. The development of Caspase-3-responsive multimodal probes is a promising research prospect. Fluorescent/photoacoustic (FL/PA) imaging has attracted considerable attention due to the high sensitivity of FL as well as the high spatial resolution and penetration depth of PA. To our knowledge, there has been no tumor-targeted FL/PA probe to monitor the activity of Caspase-3 in vivo. Therefore, we developed a tumor-targeted FL/PA probe (Bio-DEVD-HCy) for Caspase-3-responsive imaging of tumor apoptosis. Ac-DEVD-HCy without tumor-targeted biotin is used as a control probe. In vitro experiments indicated that Bio-DEVD-HCy is superior to Ac-DEVD-HCy because of the higher kinetic parameter of Bio-DEVD-HCy in comparison to Ac-DEVD-HCy. Cell and tumor imaging results suggested that Bio-DEVD-HCy could enter and accumulate in tumor cells with higher FL/PA signal with the help of tumor-targeted biotin. In detail, Bio-DEVD-HCy or Ac-DEVD-HCy could image apoptotic tumor cells with 4.3-fold or 3.5-fold FL enhancement and 3.4-fold or 1.5-fold PA enhancement. Bio-DEVD-HCy or Ac-DEVD-HCy could image tumor apoptosis with 2.5-fold or 1.6-fold FL enhancement and 4.1-fold or 1.9-fold PA enhancement. We envision that Bio-DEVD-HCy will be applied for FL/PA imaging of tumor apoptosis in clinical settings.

Nanochannel Electro-Injection as a Versatile Platform for Efficient RNA/DNA Programming on Dendritic Cells.

The utilization of dendritic cell (DC) vaccines is a promising approach in cancer immunotherapy, and the modification of DCs for the expression of tumor-associated antigens is critical for successful cancer immunotherapy. A safe and efficient method for delivering DNA/RNA into DCs without inducing maturation is beneficial to achieve successful DC transformation for cell vaccine applications, yet remains challenging. This work presents a nanochannel electro-injection (NEI) system for the safe and efficient delivery of a variety of nucleic acid molecules into DCs. The device is based on track-etched nanochannel membrane as key components, where the nano-sized channels localize the electric field on the cell membrane, enabling lower voltage (<30 V) for cell electroporation. The pulse conditions of NEI are examined so that the transfection efficiency (>70%) and biosafety (viability >85%) on delivering fluorescent dyes, plasmid DNA, messenger RNA, and circular RNA (circRNA) into DC2.4 are optimized. Primary mouse bone marrow DC can also be transfected with circRNA with 68.3% efficiency, but without remarkably affecting cellular viability or inducing DC maturation. These results suggest that NEI can be a safe and efficient transfection platform for in vitro transformation of DCs and possesses a promising potential for developing DC vaccines against cancer.

AoSAP8-P encoding A20 and/or AN1 type zinc finger protein in asparagus officinalis L. Improving stress tolerance in transgenic Nicotiana sylvestris.

The full length CDS of an A20 and AN1 type zinc finger gene (named AoSAP8-P), located nearby the male specific Y chromosome (MSY) region of Asparagus officinalis (garden asparagus) was amplified by RT-PCR from purple passion. This gene, predicted as the stress associated protein (SAPs) gene families, encodes 172 amino acids with multiple cis elements including light, stress response box, MYB and ERF binding sites on its promoter. To analyze its function, the gene expression of different organs in different asparagus gender were analyzed and the overexpressed transgenic Nicotiana sylvestris lines were generated. The results showed the gene was highly expressed in both flower and root of male garden asparagus; the germination rate of seeds of the T2 transgenic lines (T2-5-4 and T2-7-1) under the stress conditions of 125 mM NaCl and 150 mM mannitol were significantly higher than the wild type (WT) respectively. When the potted T2-5-4, T2-7-1 lines and WT were subjected to drought stress for 24 days and the leaf discs immerged into 20 % PEG6000 and 300 mM NaCl solution for 48 h respectively, the T2-5-4 and T2-7-1 with AoSAP8-P expression showed stronger drought, salt and osmotic stress tolerance. When compared, the effects of AoSAP8-P overexpression on productive development showed that the flowering time of transgenic lines, were âˆ¼ 9 day earlier with larger but fewer pollens than its WT counterparts. However, there were no significant differences in anthers, stigmas and pollen viability between the transgenic lines and WT. Our results suggested that, the AoSAP8-P gene plays a role in improving the stress resistance and shortening seeds generation time for perianal survival during the growth and development of garden asparagus.

Fluorescence imaging of intracellular telomerase activity for tumor cell identification by oligonucleotide-functionalized gold nanoparticles.

As a specific biological marker for the occurrence and progression of tumor cells, detection of telomerase activity is of great importance for the physiological research of tumors. However, in situ measurement of telomerase activity in living cells still remains a challenge. Herein, we report a precisely designed oligonucleotide-functionalized gold nanoparticle probe that has realized high-efficiency detection of telomerase activity for cellular imaging toward the identification of tumors. Our method has achieved intracellular imaging of telomerase activity and shows good performance towards the distinction of tumor cells from normal ones. Moreover, the method reported here for tracking tumor cells in blood has wide applications in cancer diagnosis. This strategy offers an opportunity for cancer diagnosis, guiding therapy and evaluating prognosis.

Cathepsin B-Activated Fluorescent and Photoacoustic Imaging of Tumor.

Early diagnosis is crucial to the treatment of cancer. Cathepsin B (CTB) plays an important role in numerous cancers, which is a promising biomarker for early diagnosis of cancer. It is necessary to exploit new probes for visualization of CTB in vivo. Fluorescent/photoacoustic (FL/PA) imaging is a powerful tool for in vivo study which possesses both excellent sensitivity and spatial resolution. To our knowledge, there has been no FL/PA probe to image CTB in vitro or in vivo. Therefore, we developed two CTB-activated FL/PA probes HCy-Cit-Val and HCy-Gly-Leu-Phe-Gly, which could successfully monitor CTB activity in vivo. Both two probes had excellent sensitivity and selectivity in vitro. Cell imaging showed that HCy-Cit-Val or HCy-Gly-Leu-Phe-Gly could image endogenous CTB in lysosome with 6.8-fold or 5.1-fold enhancement of the FL signal and 5.8-fold or 3.4-fold enhancement of the PA signal compared to their inhibitor contrast groups. Tumor imaging in vivo further confirmed the good applicability of these two probes to monitor CTB activity with high sensitivity and spatial resolution. Moreover, the property of HCy-Cit-Val is superior to HCy-Gly-Leu-Phe-Gly due to the higher catalytic efficiency of CTB toward HCy-Cit-Val than HCy-Gly-Leu-Phe-Gly. We envision that our FL/PA probe HCy-Cit-Val will be suitable for clinical early diagnosis of CTB-related cancer in the near future.

In situ imaging of intracellular human telomerase RNA with molecular beacon-functionalized gold nanoparticles.

Since the expression level of human telomerase RNA (hTR) in tumor cells is much higher than that in normal cells, the determination of hTR is of prime importance in biological research of tumors. In this work, we report molecular beacon-functionalized gold nanoparticles for hTR imaging in live cells. The molecular beacon has a loop-and-stem structure with hTR recognition sequences and a red fluorophore Cy5. In the presence of hTR, the hTR sequence could be hybridized with the loop of molecular beacon to form a duplex DNA chain and thus the fluorescence state switched from "off" to "on". After co-incubation with cells, the probe could readily permeate into cells, leading to the in situ imaging of intracellular hTR. The proposed approach could be used to differentiate tumor cells from normal ones and assess hTR expression levels in different tumor cells. Furthermore, the proposed approach allowed us to dynamically monitor the expression level of hTR in live cells and holds great potential for application in tumor diagnosis and hTR-related drug delivery.

A Multi-responsive Fluorescent Probe Reveals Mitochondrial Nucleoprotein Dynamics with Reactive Oxygen Species Regulation through Super-resolution Imaging.

Understanding the biomolecular interactions in a specific organelle has been a long-standing challenge because it requires super-resolution imaging to resolve the spatial locations and dynamic interactions of multiple biomacromolecules. Two key difficulties are the scarcity of suitable probes for super-resolution nanoscopy and the complications that arise from the use of multiple probes. Herein, we report a quinolinium derivative probe that is selectively enriched in mitochondria and switches on in three different fluorescence modes in response to hydrogen peroxide (H2 O2 ), proteins, and nucleic acids, enabling the visualization of mitochondrial nucleoprotein dynamics. STED nanoscopy reveals that the proteins localize at mitochondrial cristae and largely fuse with nucleic acids to form nucleoproteins, whereas increasing H2 O2 level leads to disassociation of nucleic acid-protein complexes.

Silica-anchored cadmium sulfide nanocrystals for the optical detection of copper(II).

A fluorometric assay was developed for the determination of copper(II) ion based on its quenching effect on the green fluorescent probe of SiO2-anchored CdS nanocrystals (SiO2/CdS NCs). The fluorescent probe was prepared by a surface ion-directing strategy for homogeneous precipitation of CdS NCs onto the carboxyl-capped SiO2 core surfaces. In comparison to CdS NCs, the SiO2/CdS NCs has strong fluorescence emission and good photostability. Moreover, SiO2/CdS NCs show higher fluorescence selectivity for copper(II) ions than for other common metal ions because copper(II) ions have a strong fluorescence quenching effect on SiO2/CdS NCs. At excitation/emission wavelengths of 300/516 nm, the resulting fluorescent probe shows wide linear ranges from 0.01 to 2 µM with a detection limit of 6.3 nM for copper(II) ions. The method has been applied to the determination of trace copper(II) ions in tea infusions with satisfactory results. Graphical abstract.

Chemical characterization, antioxidant properties and anticancer activity of exopolysaccharides from Floccularia luteovirens.

Two polysaccharides, ALF1 and ALF2 were obtained from the fermentation liquid of Floccularia luteovirens. These fractions had good performance in scavenging radicals and ALF1 exhibited obvious antioxidant activities. Further, linkage analysis and NMR were used to characterize the structures of ALF1. Linkage and NMR data comprehensively showed that ALF1 mainly contained six kinds of linkage type units as →4)-ß-D-Manp→, 1,3-α-Fucp→, α-L-Araf-C1→, →6)-ß-D-Galp-C1→, →4)-α-D-GlcAp-(1→ and →3)-ß-D-Glcp(1→. In addition, ALF1 had good bioactivities such as anticancer and antioxidant activities. ALF1 was proven to be able to inhibit tumor cells without affecting the normal cells. Besides, ALF1 improved the activities of SOD, GSH-Px and CAT, and decreased the production of MDA which result in protecting PC12 cells against H2O2-induced oxidative stress. ALF1 decreased ROS production, and stabilize mitochondrial membrane potential. The findings indicated that the fermentation liquid of Floccularia luteovirens could be used as a potential natural source of antioxidant.

MicroRNA­671­3p regulates the development of knee osteoarthritis by targeting TRAF3 in chondrocytes.

Osteoarthritis (OA) is a degenerative joint disease characterized by articular cartilage degradation and joint inflammation. A previous study showed that microRNA (miR)­671­3p is involved in the development of OA, however, its function and molecular target in chondrocytes during the pathogenesis of OA remain to be fully elucidated. In the present study, miR­671­3p was significantly downregulated in knee OA cartilage tissues compared with normal cartilage tissues. The expression levels of pro­inflammatory cytokines, including interleukin (IL)­1ß, IL­6, IL­8 and tumor necrosis factor (TNF)­α, in the knee OA cartilage tissues were significantly higher than those in the normal cartilage tissues. Through gain­of­function and loss­of­function experiments, miR­671­3p was shown to significantly affect matrix synthesis gene expression, cell proliferation, apoptosis and inflammation in chondrocytes from patients with OA. Subsequent bioinformatics analysis identified potential target sites of the miR­671­3p located in the 3'untranslated region of TNF receptor­associated factor (TRAF3). The results of a dual­luciferase reporter assay showed that TRAF3 is a target gene of miR­671­3p. Western blot analysis demonstrated that miR­671­3p inhibited the gene expression of TRAF3. Furthermore, the restoration of TRAF3 markedly abrogated the effect of miR­671­3p. Taken together, the present study suggests that miR­671­3p may be important in the pathogenesis of OA through targeting TRAF3 and regulating chondrocyte apoptosis and inflammation, which may be a potential molecular target for OA treatment.

EPEL promotes the migration and invasion of osteosarcoma cells by upregulating ROCK1.

E2F-mediated cell proliferation enhancing long non-coding RNA (lncRNA) (EPEL) is a newly identified lncRNA involved in the regulation of lung cancer cell proliferation. However, its association with other types of cancer is unknown. The present study recruited patients with osteosarcoma and healthy controls. Tumor and adjacent healthy tissues were obtained from patients with osteosarcoma, and whole blood was extracted from patients and healthy controls. The expression levels of EPEL in tissues were detected by reverse transcription-quantitative polymerase chain reaction. The diagnostic value of serum EPEL for osteosarcoma was evaluated by receiver operating characteristic curve analysis. The association between serum levels of EPEL and basic clinical patient information was analyzed by χ2 test. Subsequently, EPEL overexpression in osteosarcoma cell lines was established, and its effects on cell migration and invasion were explored by Transwell assay. The implications of EPEL overexpression on Rho-associated coiled-coil containing protein kinase 1 (ROCK1) expression were investigated by western blotting. The results revealed that EPEL was upregulated in tumor tissues compared with adjacent tissues. In addition, serum levels of EPEL were higher in patients with osteosarcoma compared with healthy controls, and were positively associated with distant tumor metastasis. Furthermore, EPEL overexpression promoted the migration and invasion of osteosarcoma cells and induced overexpression of ROCK1. In conclusion, these results suggested that EPEL may promote the migration and invasion of osteosarcoma cells by upregulating ROCK1.

Visualizing telomerase activity for tumour identification by hybridization-triggered ratiometric fluorescence.

Telomerase is repressed stringently in normal cells but highly activated in tumours to maintain their uncontrollable growth. We report a ratiometric fluorescence strategy for visualising telomerase activity for tumour identification. Using our strategy, tumour cells and tissues can be differentiated from normal cells and tissues by the naked eye.

miR-29a inhibits adhesion, migration, and invasion of osteosarcoma cells by suppressing CDC42.

Osteosarcoma is one of the most common tumors of the bone in children and adolescents worldwide. The relapse and metastasis of osteosarcoma are a major therapeutic challenge. Recently, several metastasis regulators, including miRNAs, kinases, and lncRNAs, were reported in osteosarcoma. Identifying novel regulators of metastasis will be useful to explore novel biomarkers for osteosarcoma. The present study showed miR-29a overexpression significantly inhibited HOS and MG-63 cell adhesion, invasion, and migration. About 70% of the wound area was repaired by migrating cells after 24 h in the control group, and only 50% of the wound area was repaired in the miR-29a overexpression group. The numbers of invading cells were decreased by 40% and 50% in HOS and MG-63 cells transfected with miR-29a, respectively, compared with the negative control group. Moreover, the present study validated that CDC42 was a direct target of miR-29a in OS cells. In conclusion, miR-29a may serve as a therapeutic target for osteosarcoma.

SUMO-specific protease 2 (SENP2) functions as a tumor suppressor in osteosarcoma via SOX9 degradation.

Osteosarcoma (OS) is the most common primary bone malignancy in children and adolescents, the pathogenesis of which remain largely unknown. Small ubiquitin-like modifier (SUMO)-Specific Protease 2 (SENP2) has been reported to serve as a tumor suppressor in hepatocellular carcinoma cells. The aim of the present study was to investigate the critical role of SENP2 in OS cells. Using reverse transcription-quantitative polymerase chain reaction and western blot assays, it was observed that SENP2 was significantly downregulated in clinical OS tissues compared with adjacent normal samples. Ectopic expression of SENP2 resulted in the suppression of proliferation, migration and invasion in OS cells, whereas SENP2 knockdown by CRISPR-Cas9-based gene editing had the opposite effect. SENP2 is associated with the proteasome-dependent ubiquitination and degradation of SRY-box-9 (SOX9). SOX9 silencing impaired SENP2-depletion-induced accelerated cell growth and migration. Together, these results suggest that SOX9 is a critical downstream effector of the tumor suppressor SENP2 in OS.

Sticky-flares for in situ monitoring of human telomerase RNA in living cells.

Human telomerase RNA (hTR), a template of telomerase for telomeric repeat synthesis, was used to reflect the telomerase activity and act as a potential target of antitumor therapy. Here, we report a novel DNA-conjugated AuNP probe termed sticky-flares for the in situ detection of intracellular human telomerase RNA. The sticky-flares probe is capable of entering living cells directly without any auxiliary and recognizing the binding domain of human telomerase RNA. On recognition, the fluorophore-modified recognition flares can specifically bind to the target, separate from the sticky-flares and act as a fluorescent reporter to quantify and dynamically profile human telomerase RNA in living cells. We envision that the sticky-flares probe would be a valuable platform to investigate the function and regulation of hTR in antitumor therapy and hTR-related drug invention.

RNA interference of GhPEPC2 enhanced seed oil accumulation and salt tolerance in Upland cotton.

Phosphoenolpyruvate carboxylase (PEPCase) mainly produces oxaloacetic acid for tricarboxylic acid (TCA) cycle. Here we reported that GhPEPC2 silencing with PEPC2-RNAi vector could regulate oil and protein accumulation in cottonseeds. In GhPEPC2 transgenic plants, PEPCase activities in immature embryos were significantly reduced, and the oil content in seed kernel was increased 7.3 percentages, whereas total proteins decreased 5.65 percentages. Compared to wild type, agronomical traits of transgenic plant were obviously unaffected. Furthermore, gene expression profile of GhPEPC2 transgenic seeds were investigated using RNA-seq, most lipid synthesis related genes were up-regulated, but amino acid metabolic related genes were down-regulated. In addition, the GhPEPC2 transgenic cotton seedlings were stressed using sodium salts at seedling stage, and the salt tolerance was significantly enhanced. Our observations of GhPEPC2 in cotton would shade light on understanding the regulation of oil content, protein accumulation and salt tolerance enhancement in other plants.