Genomic and Epigenomic Changes in the Progeny of Cold-Stressed Arabidopsis thaliana Plants.

Plants are continuously exposed to various environmental stresses. Because they can not escape stress, they have to develop mechanisms of remembering stress exposures somatically and passing it to the progeny. We studied the Arabidopsis thaliana ecotype Columbia plants exposed to cold stress for 25 continuous generations. Our study revealed that multigenerational exposure to cold stress resulted in the changes in the genome and epigenome (DNA methylation) across generations. Main changes in the progeny were due to the high frequency of genetic mutations rather than epigenetic changes; the difference was primarily in single nucleotide substitutions and deletions. The progeny of cold-stressed plants exhibited the higher rate of missense non-synonymous mutations as compared to the progeny of control plants. At the same time, epigenetic changes were more common in the CHG (C = cytosine, H = cytosine, adenine or thymine, G = guanine) and CHH contexts and favored hypomethylation. There was an increase in the frequency of C to T (thymine) transitions at the CHH positions in the progeny of cold stressed plants; because this type of mutations is often due to the deamination of the methylated cytosines, it can be hypothesized that environment-induced changes in methylation contribute to mutagenesis and may be to microevolution processes and that RNA-dependent DNA methylation plays a crucial role. Our work supports the existence of heritable stress response in plants and demonstrates that genetic changes prevail.

Epigenetic aspects of floral homeotic genes in relation to sexual dimorphism in the dioecious plant Mercurialis annua.

In plants, dioecy characterizes species that carry male and female flowers on separate plants and it occurs in about 6% of angiosperms; however, the molecular mechanisms that underlie dioecy are essentially unknown. The ability for sex-reversal by hormone application raises the hypothesis that the genes required for the expression of both sexes are potentially functional but are regulated by epigenetic means. In this study, proteomic analysis of nuclear proteins isolated from flower buds of females, males, and feminized males of the dioecious plant Mercurialis annua revealed differential expression of nuclear proteins that are implicated in chromatin structure and function, including floral homeotic proteins. Focusing on floral genes, we found that class B genes were mainly expressed in male flowers, while class D genes, as well as SUPERMAN-like genes, were mainly expressed in female flowers. Cytokinin-induced feminization of male plants was associated with down-regulation of male-specific genes concomitantly with up-regulation of female-specific genes. No correlation was found between the expression of class B and D genes and the changes in DNA methylation or chromatin conformation of these genes. Thus, we could not confirm DNA methylation or chromatin conformation of floral genes to be the major determinant regulating sexual dimorphisms. Instead, determination of sex in M. annua might be controlled upstream of floral genes by one or more sex-specific factors that affect hormonal homeostasis. A comprehensive model is proposed for sex-determination in M. annua.

Activation of Tag1 transposable elements in Arabidopsis dedifferentiating cells and their regulation by CHROMOMETHYLASE 3-mediated CHG methylation.

Dedifferentiation, that is, the acquisition of stem cell-like state, commonly induced by stress (e.g., protoplasting), is characterized by open chromatin conformation, a chromatin state that could lead to activation of transposable elements (TEs). Here, we studied the activation of the Arabidopsis class II TE Tag1, in which two copies, situated close to each other (near genes) on chromosome 1 are found in Landsberg erecta (Ler) but not in Columbia (Col). We first transformed protoplasts with a construct in which a truncated Tag1 (ΔTag1 non-autonomous) blocks the expression of a reporter gene AtMBD5-GFP and found a relatively high ectopic excision of ΔTag1 accompanied by expression of AtMBD5-GFP in protoplasts derived from Ler compared to Col; further increase was observed in ddm1 (decrease in DNA methylation1) protoplasts (Ler background). Ectopic excision was associated with transcription of the endogenous Tag1 and changes in histone H3 methylation at the promoter region. Focusing on the endogenous Tag1 elements we found low level of excision in Ler protoplasts, which was slightly and strongly enhanced in ddm1 and cmt3 (chromomethylase3) protoplasts, respectively, concomitantly with reduction in Tag1 gene body (GB) CHG methylation and increased Tag1 transcription; strong activation of Tag1 was also observed in cmt3 leaves. Notably, in cmt3, but not in ddm1, Tag1 elements were excised out from their original sites and transposed elsewhere in the genome. Our results suggest that dedifferentiation is associated with Tag1 activation and that CMT3 rather than DDM1 plays a central role in restraining Tag1 activation via inducing GB CHG methylation.

The SbSOS1 gene from the extreme halophyte Salicornia brachiata enhances Na(+) loading in xylem and confers salt tolerance in transgenic tobacco.

BACKGROUND: Soil salinity adversely affects plant growth and development and disturbs intracellular ion homeostasis resulting cellular toxicity. The Salt Overly Sensitive 1 (SOS1) gene encodes a plasma membrane Na(+)/H(+) antiporter that plays an important role in imparting salt stress tolerance to plants. Here, we report the cloning and characterisation of the SbSOS1 gene from Salicornia brachiata, an extreme halophyte. RESULTS: The SbSOS1 gene is 3774 bp long and encodes a protein of 1159 amino acids. SbSOS1 exhibited a greater level of constitutive expression in roots than in shoots and was further increased by salt stress. Overexpressing the S. brachiata SbSOS1 gene in tobacco conferred high salt tolerance, promoted seed germination and increased root length, shoot length, leaf area, fresh weight, dry weight, relative water content (RWC), chlorophyll, K(+)/Na(+) ratio, membrane stability index, soluble sugar, proline and amino acid content relative to wild type (WT) plants. Transgenic plants exhibited reductions in electrolyte leakage, reactive oxygen species (ROS) and MDA content in response to salt stress, which probably occurred because of reduced cytosolic Na(+) content and oxidative damage. At higher salt stress, transgenic tobacco plants exhibited reduced Na(+) content in root and leaf and higher concentrations in stem and xylem sap relative to WT, which suggests a role of SbSOS1 in Na(+) loading to xylem from root and leaf tissues. Transgenic lines also showed increased K(+) and Ca(2+) content in root tissue compared to WT, which reflect that SbSOS1 indirectly affects the other transporters activity. CONCLUSIONS: Overexpression of SbSOS1 in tobacco conferred a high degree of salt tolerance, enhanced plant growth and altered physiological and biochemical parameters in response to salt stress. In addition to Na(+) efflux outside the plasma membrane, SbSOS1 also helps to maintain variable Na(+) content in different organs and also affect the other transporters activity indirectly. These results broaden the role of SbSOS1 in planta and suggest that this gene could be used to develop salt-tolerant transgenic crops.

A novel salt-inducible gene SbSI-1 from Salicornia brachiata confers salt and desiccation tolerance in E. coli.

Salicornia brachiata is one of the extreme salt tolerant plants and grows luxuriantly in coastal areas. Previously we have reported isolation and characterization of ESTs from S. brachiata with large number of unknown gene sequences. Reverse Northern analysis showed upregulation and downregulation of few unknown genes in response to salinity. Some of these unknown genes were made full length and their functional analysis is being tested. In this study, we have selected a novel unknown salt inducible gene SbSI-1 (Salicornia brachiata salt inducible-1) for the functional validation. The SbSI-1 (Gen-Bank accession number JF 965339) was made full length and characterized in detail for its functional validation under desiccation and salinity. The SbSI-1 gene is 917 bp long, and contained 437 bp 3' UTR, and 480 bp ORF region encoding 159 amino acids protein with estimated molecular mass of 18.39 kDa and pI 8.58. The real time PCR analysis revealed high transcript expression in salt, desiccation, cold and heat stresses. However, the maximum expression was obtained by desiccation. The ORF region of SbSI-1 was cloned in pET28a vector and transformed in BL21 (DE3) E. coli cells. The SbSI-1 recombinant E. coli cells showed tolerance to desiccation and salinity stress compared to only vector in the presence of stress.

Multigenerational Exposure to Heat Stress Induces Phenotypic Resilience, and Genetic and Epigenetic Variations in Arabidopsis thaliana Offspring.

Plants are sedentary organisms that constantly sense changes in their environment and react to various environmental cues. On a short-time scale, plants respond through alterations in their physiology, and on a long-time scale, plants alter their development and pass on the memory of stress to the progeny. The latter is controlled genetically and epigenetically and allows the progeny to be primed for future stress encounters, thus increasing the likelihood of survival. The current study intended to explore the effects of multigenerational heat stress in Arabidopsis thaliana. Twenty-five generations of Arabidopsis thaliana were propagated in the presence of heat stress. The multigenerational stressed lineage F25H exhibited a higher tolerance to heat stress and elevated frequency of homologous recombination, as compared to the parallel control progeny F25C. A comparison of genomic sequences revealed that the F25H lineage had a three-fold higher number of mutations [single nucleotide polymorphisms (SNPs) and insertions and deletions (INDELs)] as compared control lineages, suggesting that heat stress induced genetic variations in the heat-stressed progeny. The F25H stressed progeny showed a 7-fold higher number of non-synonymous mutations than the F25C line. Methylome analysis revealed that the F25H stressed progeny showed a lower global methylation level in the CHH context than the control progeny. The F25H and F25C lineages were different from the parental control lineage F2C by 66,491 and 80,464 differentially methylated positions (DMPs), respectively. F25H stressed progeny displayed higher frequency of methylation changes in the gene body and lower in the body of transposable elements (TEs). Gene Ontology analysis revealed that CG-DMRs were enriched in processes such as response to abiotic and biotic stimulus, cell organizations and biogenesis, and DNA or RNA metabolism. Hierarchical clustering of these epimutations separated the heat stressed and control parental progenies into distinct groups which revealed the non-random nature of epimutations. We observed an overall higher number of epigenetic variations than genetic variations in all comparison groups, indicating that epigenetic variations are more prevalent than genetic variations. The largest difference in epigenetic and genetic variations was observed between control plants comparison (F25C vs. F2C), which clearly indicated that the spontaneous nature of epigenetic variations and heat-inducible nature of genetic variations. Overall, our study showed that progenies derived from multigenerational heat stress displayed a notable adaption in context of phenotypic, genotypic and epigenotypic resilience.

Paclobutrazol improves surface water use efficiency by regulating allometric trait behavior in maize.

Paclobutrazol (PBZ) role in drought management of maize is least understood. In maize, root traits are linked with surface water management. Over three years, early and terminal deficit irrigation (EDI and TDI) with or without PBZ were imposed on DKC-9144 and PG-2475 maize varieties. Several allometric parameters viz. stem height, stem diameter, leaf area and root traits along with physiological processes were measured. Implication of these parameters in the management of soil surface irrigation in terms of water use efficiency (WUE) was demonstrated in maize. Increased number of lateral roots and root number density in DKC-9144 provided more surface area for water absorption for better management of EDI. Root growth rates showed a similar pattern with root length, root surface areas, and root numbers in EDI. Elevated expressions of ZmRTCL, ZmRTCS and ZmARF34 in EDI and EDI plus PBZ were associated with seminal roots and root laterals initiation. Under TDI alone or in combination with PBZ, root lengths (BRL, CRL, SRL) and root surface areas varied in DKC-9144 and PG-2475 over control. Furthermore, correlation analysis showed that decrease in WUE under TDI was significantly associated with a reduction in stem thickness and leaf surface area. For WUE_N in TDI and PBZ plus TDI, structural equation modelling proposed, brace root surface area (BRSA_N) as a positive contributor, while a negative contributor was seminal root surface area (SRSA_N). Present study explained the importance of specific root traits and their association with other allometric parameters for improving WUE in DKC-9144 variety of maize and the crop in general.

Cloning and characterization of the Salicornia brachiata Na(+)/H(+) antiporter gene SbNHX1 and its expression by abiotic stress.

Salinity causes multifarious adverse effects to plants. Plants response to salt stress involves numerous processes that function in coordination to alleviate both cellular hyperosmolarity and ion disequilibrium. A Na(+)/H(+) antiporter NHX1 gene has been isolated from a halophytic plant Salicornia brachiata in this study. Predicted amino acid sequence similarity, protein topology and the presence of functional domains conserved in SbNHX1 classify it as a plant vacuolar NHX gene. The SbNHX1 cDNA has an open reading frame of 1,683 bp, encoding a polypeptide of 560 amino acid residues with an estimated molecular mass 62.44 kDa. The SbNHX1 shows high amino acid similarity with other halophytic NHX gene and belongs to Class-I type NHXs. TMpred suggests that SbNHX1 contains 11 strong transmembrane (TM). Real time PCR analysis revealed that SbNHX1 transcript expresses maximum at 0.5 M. Transcript increases gradually by increasing the treatment duration at 0.5 M NaCl, however, maximum expression was observed at 48 h. The overexpression of SbNHX1 gene in tobacco plant showed NaCl tolerance. This study shows that SbNHX1 is a potential gene for salt tolerance, and can be used in future for developing salt tolerant crops.

Special Issue Editorial: Isolation and Analysis of Characteristic Compounds from Herbal and Plant Extracts.

Herbal and plant extracts exhibit various types of properties and activities that have been applied in the medicinal field to treat diseases and achieve better health [...].

Development and Standardization of Rapid and Efficient Seed Germination Protocol for Cannabis sativa.

Cannabis seed germination is an important process for growers and researchers alike. Many biotechnological applications require a reliable sterile method for seed germination. This protocol outlines a seed germination procedure for Cannabis sativa using a hydrogen peroxide (H2O2) solution as liquid germination media. In this protocol, all three steps including seed sterilization, germination, and seedlings development were carried out in an H2O2 solution of different concentrations; 1% H2O2 solution showed the fastest and the most efficient germination. This protocol also exhibited high germination efficiency for very old cannabis seeds with lower viability. Overall, this protocol demonstrates superior germination compared to water control and reduces the risk of contamination, making it suitable for tissue culture and other sensitive applications.

Plants Metabolome Study: Emerging Tools and Techniques.

Metabolomics is now considered a wide-ranging, sensitive and practical approach to acquire useful information on the composition of a metabolite pool present in any organism, including plants. Investigating metabolomic regulation in plants is essential to understand their adaptation, acclimation and defense responses to environmental stresses through the production of numerous metabolites. Moreover, metabolomics can be easily applied for the phenotyping of plants; and thus, it has great potential to be used in genome editing programs to develop superior next-generation crops. This review describes the recent analytical tools and techniques available to study plants metabolome, along with their significance of sample preparation using targeted and non-targeted methods. Advanced analytical tools, like gas chromatography-mass spectrometry (GC-MS), liquid chromatography mass-spectroscopy (LC-MS), capillary electrophoresis-mass spectrometry (CE-MS), fourier transform ion cyclotron resonance-mass spectrometry (FTICR-MS) matrix-assisted laser desorption/ionization (MALDI), ion mobility spectrometry (IMS) and nuclear magnetic resonance (NMR) have speed up precise metabolic profiling in plants. Further, we provide a complete overview of bioinformatics tools and plant metabolome database that can be utilized to advance our knowledge to plant biology.

Nickel excess affects phenology and reproductive attributes of Asterella wallichiana and Plagiochasma appendiculatum growing in natural habitats.

Bryophytes are potent metal absorbers, thriving well on heavy metal (HM)-polluted soils. Mechanisms controlling uptake, compartmentalization and impacts of HMs on bryophytes life cycle are largely unknown. The current study is an effort to decipher mechanisms of nickel (Ni) excess-induced effects on the phenological events of two bryophytes, Asterella wallichiana and Plagiochasma apendiculatum growing in natural habitats. Observations revealed Ni-excess induced negative impacts on abundance, frequency of occurrence of reproductive organs, population viability and morphological traits, spore viability and physiological attributes of both the liverworts. Results led us conclude that P. appendiculatum survived better with the lowest impact on its life cycle events than A. wallichiana under Ni excess in natural habitats. Our findings collectively provide insights into the previously unknown mechanisms of Ni-induced responses in liverworts with respect to phenological attributes, as well as demonstrate the potential of P. appendiculatum to survive better in Ni excess habitats.

OMICs, Epigenetics, and Genome Editing Techniques for Food and Nutritional Security.

The incredible success of crop breeding and agricultural innovation in the last century greatly contributed to the Green Revolution, which significantly increased yields and ensures food security, despite the population explosion. However, new challenges such as rapid climate change, deteriorating soil, and the accumulation of pollutants require much faster responses and more effective solutions that cannot be achieved through traditional breeding. Further prospects for increasing the efficiency of agriculture are undoubtedly associated with the inclusion in the breeding strategy of new knowledge obtained using high-throughput technologies and new tools in the future to ensure the design of new plant genomes and predict the desired phenotype. This article provides an overview of the current state of research in these areas, as well as the study of soil and plant microbiomes, and the prospective use of their potential in a new field of microbiome engineering. In terms of genomic and phenomic predictions, we also propose an integrated approach that combines high-density genotyping and high-throughput phenotyping techniques, which can improve the prediction accuracy of quantitative traits in crop species.

Transient expression of the ß-glucuronidase gene in Cannabis sativa varieties.

In plant biology, transient expression analysis plays a vital role to provide a fast method to study the gene of interest. In this study, we report a rapid and efficient method for transient expression in Cannabis sativa seedlings using Agrobacterium tumefaciens-mediated transformation. A. tumefaciens strain EHA105 carrying the pCAMBIA1301 construct with uidA gene was used to transform cannabis seedlings and the GUS assay (a measurement of ß-glucuronidase activity) was used to detect the uidA expression. In the current study, we have also established a rapid germination protocol for cannabis seeds. The all three steps seed sterilization, germination and seedlings development were carried out in a 1% H2O2 solution. Transient transformation revealed that both cotyledons and young true leaves are amenable to transformation. Compared with tobacco (Nicotiana benthamiana), cannabis seedlings were less susceptible to transformation with A. tumefaciens. Susceptibility to Agrobacterium transformation also varied with the different cannabis varieties. The method established in this study has the potential to be an important tool for gene function studies and genetic improvement in cannabis.

Development and Optimization of a Germination Assay and Long-Term Storage for Cannabis sativa Pollen.

Pollen viability and storage is of great interest to cannabis breeders and researchers to maintain desirable germplasm for future use in breeding or for biotechnological and gene editing applications. Here, we report a simple and efficient cryopreservation method for long-term storage of Cannabis sativa pollen. Additionally, the bicellular nature of cannabis pollen was identified using DAPI (4',6-diamidino-2-phenylindole) staining. A pollen germination assay was developed to assess cannabis pollen viability and used to demonstrate that pollen collected from different principal growth stages exhibited differential longevity. Finally, a simple and efficient method that employs pollen combined with baked whole wheat flour and subsequent desiccation under vacuum was developed for the long-term cryopreservation of C. sativa pollen. Using this method, pollen viability was maintained in liquid nitrogen after four months, suggesting long-term preservation of cannabis pollen.

Arabidopsis mutants may represent recombinant introgression lines.

OBJECTIVES: It is a common practice in Arabidopsis to transfer a mutation generated in one genetic background to other genetic background via crossing. However, the drawback of this methodology is unavoidable presence of genomic fragments from the donor parent being often replacing desirable genomic fragments of the recurrent parent. Here, we highlighted problem of Arabidopsis mutants being recombinant introgression lines that can lead to unreliable and misinterpreted results. RESULTS: We studied the regulation of low copy number transposable elements Tag1 and Evelknievel (EK), located at the end of the bottom arm of chromosome 1 and both are present in the Arabidopsis Landsberg erecta (Ler) but not in Columbia (Col) ecotype. Using various epigenetic mutants (cmt3, ddm1, kyp2, ago4, rdr2 hen1 etc.), we found that certain mutants in the Ler background are deficient of Tag1 or EK or both and represent recombinant introgression lines whereby chromosomal regions from Col have been recombined into the Ler genome. Our data support a recent proposal calling for formulating standards for authentication of plant lines that are used in plant research. Most important is to verify that a given trait or genomic locus under study is correctly identified, particularly when using mutants generated by crossing.

CMT3 and SUVH4/KYP silence the exonic Evelknievel retroelement to allow for reconstitution of CMT1 mRNA.

BACKGROUND: The Chromomethylase 1 (CMT1) has long been considered a nonessential gene because, in certain Arabidopsis ecotypes, the CMT1 gene is disrupted by the Evelknievel (EK) retroelement, inserted within exon 13, or contains frameshift mutations, resulting in a truncated, non-functional protein. In contrast to other transposable elements, no transcriptional activation of EK was observed under stress conditions (e.g., protoplasting). RESULTS: We wanted to explore the regulatory pathway responsible for EK silencing in the Ler ecotype and its effect on CMT1 transcription. Methylome databases confirmed that EK retroelement is heavily methylated and methylation is extended toward CMT1 downstream region. Strong transcriptional activation of EK accompanied by significant reduction in non-CG methylation was found in cmt3 and kyp2, but not in ddm1 or RdDM mutants. EK activation in cmt3 and kyp2 did not interfere with upstream CMT1 expression but abolish transcription through the EK. We identified, in wild-type Ler, three spliced variants in which the entire EK is spliced out; one variant (25% of splicing incidents) facilitates proper reconstitution of an intact CMT1 mRNA. We could recover very low amount of the full-length CMT1 mRNA from WT Ler and Col, but not from cmt3 mutant. CONCLUSIONS: Our findings highlight CMT3-SUVH4/KYP as the major pathway silencing the intragenic EK via inducing non-CG methylation. Furthermore, retroelement insertion within exons (e.g., CMT1) may not lead to a complete abolishment of the gene product when the element is kept silent. Rather the element can be spliced out to bring about reconstruction of an intact, functional mRNA and possibly retrieval of an active protein.

S1-Type Endonuclease 2 in Dedifferentiating Arabidopsis Protoplasts: Translocation to the Nucleus in Senescing Protoplasts Is Associated with De-Glycosylation.

Cell dedifferentiation characterizes the transition of leaf cells to protoplasts and is accompanied by global chromatin decondensation. Here we show that in Arabidopsis, chromocentric chromatin undergoes prompt and gradual decondensation upon protoplasting. We hypothesized that prompt chromatin decondensation is unlikely to be driven solely by epigenetic means and other factors might be involved. We investigated the possibility that S1-type endonucleases are involved in prompt chromatin decondensation via their capability to target and cleave unpaired regions within superhelical DNA, leading to chromatin relaxation. We showed that the expression and activity of the S1-type endonuclease 2 (ENDO2) is upregulated in dedifferentiating protoplasts concomitantly with chromatin decondensation. Mutation of the ENDO2 gene did not block or delay chromocentric chromatin decondensation upon protoplasting. Further study showed that ENDO2 subcellular localization is essentially cytoplasmic (endoplasmic reticulum-associated) in healthy cells, but often localized to the nucleus and in senescing/dying cells it was associated with fragmented nuclei. Using in gel nuclease assays we identified two ENDO2 variants, designated N1 (cytoplasmic variant) and N2 (cytoplasmic and nuclear variant), and based on their capability to bind concanavalin A (ConA), they appear to be glycosylated and de-glycosylated (or decorated with ConA non-binding sugars), respectively. Our data showed that the genome is responding promptly to acute stress (protoplasting) by acquiring decondensation state, which is not dependent on ENDO2 activity. ENDO2 undergoes de-glycosylation and translocation to the nucleus where it is involved in early stages of cell death probably by introducing double strand DNA breaks into superhelical DNA leading to local chromatin relaxation and fragmentation of nuclei.

Nanoformulations of quercetin: a potential phytochemical for the treatment of uv radiation induced skin damages

Abstract The continuous prolonged exposures of sun light especially the ultra violet (UV) radiation present in it, cause not only the risk of skin cancer but also it may cause premature skin aging, photodermatoses and actinic keratoses. Flavonoids (including Flavane, Flavanone, Flavone, Flavonol, Isoflavone, Neoflavone etc.) having potent antioxidant activity, used as topical applications for protection against UV induced skin damages as well as for skin care. Most commonly used flavonoid is quercetin (Flavonol), which is present in fruits, vegetables, and herbs. We aim to review the research focused on development of different novel formulations to treat UV radiations induced skin diseases. In this review, several formulations of flavonoid quercetin were discussed and their outcomes were compiled and compared in context to solubility, stability and efficiency of application. On the basis this comparative analysis we have concluded that three formulations, namely glycerosomes, nanostructured lipid carriers and deformable liposomes hold good applications for future aspects for topical delivery of quercetin. These formulations showed enhanced stability, increased quercetin accumulation in different skin layers, facilitated drug permeation in skin and long-lasting drug release.

A SNARE-Like Superfamily Protein SbSLSP from the Halophyte Salicornia brachiata Confers Salt and Drought Tolerance by Maintaining Membrane Stability, K(+)/Na(+) Ratio, and Antioxidant Machinery.

About 1000 salt-responsive ESTs were identified from an extreme halophyte Salicornia brachiata. Among these, a novel salt-inducible gene SbSLSP (Salicornia brachiata SNARE-like superfamily protein), showed up-regulation upon salinity and dehydration stress. The presence of cis-regulatory motifs related to abiotic stress in the putative promoter region supports our finding that SbSLSP gene is inducible by abiotic stress. The SbSLSP protein showed a high sequence identity to hypothetical/uncharacterized proteins from Beta vulgaris, Spinacia oleracea, Eucalyptus grandis, and Prunus persica and with SNARE-like superfamily proteins from Zostera marina and Arabidopsis thaliana. Bioinformatics analysis predicted a clathrin adaptor complex small-chain domain and N-myristoylation site in the SbSLSP protein. Subcellular localization studies indicated that the SbSLSP protein is mainly localized in the plasma membrane. Using transgenic tobacco lines, we establish that overexpression of SbSLSP resulted in elevated tolerance to salt and drought stress. The improved tolerance was confirmed by alterations in a range of physiological parameters, including high germination and survival rate, higher leaf chlorophyll contents, and reduced accumulation of Na(+) ion and reactive oxygen species (ROS). Furthermore, overexpressing lines also showed lower water loss, higher cell membrane stability, and increased accumulation of proline and ROS-scavenging enzymes. Overexpression of SbSLSP also enhanced the transcript levels of ROS-scavenging and signaling enzyme genes. This study is the first investigation of the function of the SbSLSP gene as a novel determinant of salinity/drought tolerance. The results suggest that SbSLSP could be a potential candidate to increase salinity and drought tolerance in crop plants for sustainable agriculture in semi-arid saline soil.