Stress-associated Protein Gene Research Articles

Genomic Analysis of Stress Associated Proteins in Soybean and the Role of GmSAP16 in Abiotic Stress Responses in Arabidopsis and Soybean.

Stress associated proteins (SAPs) containing A20/AN1 zinc finger domains have emerged as novel regulators of stress responses. In this study, 27 SAP genes were identified in soybean. The phylogenetic relationships, exon–intron structure, domain structure, chromosomal localization, putative cis-acting elements, and expression patterns of SAPs in various tissues under abiotic stresses were analyzed. Among the soybean SAP genes, GmSAP16 was significantly induced by water deficit stress, salt, and abscisic acid (ABA) and selected for further analysis. GmSAP16 was located in the nucleus and cytoplasm. The overexpression of GmSAP16 in Arabidopsis improved drought and salt tolerance at different developmental stages and increased ABA sensitivity, as indicated by delayed seed germination and stomatal closure. The GmSAP16 transgenic Arabidopsis plants had a higher proline content and a lower water loss rate and malondialdehyde (MDA) content than wild type (WT) plants in response to stresses. The overexpression of GmSAP16 in soybean hairy roots enhanced drought and salt tolerance of soybean seedlings, with higher proline and chlorophyll contents and a lower MDA content than WT. RNA inference (RNAi) of GmSAP16 increased stress sensitivity. Stress-related genes, including GmDREB1B;1, GmNCED3, GmRD22, GmDREB2, GmNHX1, and GmSOS1, showed significant expression alterations in GmSAP16-overexpressing and RNAi plants under stress treatments. These results indicate that soybean SAP genes play important roles in abiotic stress responses.

A Stress-Associated Protein, PtSAP13, From Populus trichocarpa Provides Tolerance to Salt Stress.

The growth and production of poplars are usually affected by unfavorable environmental conditions such as soil salinization. Thus, enhancing salt tolerance of poplars will promote their better adaptation to environmental stresses and improve their biomass production. Stress-associated proteins (SAPs) are a novel class of A20/AN1 zinc finger proteins that have been shown to confer plants’ tolerance to multiple abiotic stresses. However, the precise functions of SAP genes in poplars are still largely unknown. Here, the expression profiles of Populus trichocarpa SAPs in response to salt stress revealed that PtSAP13 with two AN1 domains was up-regulated dramatically during salt treatment. The β-glucuronidase (GUS) staining showed that PtSAP13 was accumulated dominantly in leaf and root, and the GUS signal was increased under salt condition. The Arabidopsis transgenic plants overexpressing PtSAP13 exhibited higher seed germination and better growth than wild-type (WT) plants under salt stress, demonstrating that overexpression of PtSAP13 increased salt tolerance. Higher activities of antioxidant enzymes were found in PtSAP13-overexpressing plants than in WT plants under salt stress. Transcriptome analysis revealed that some stress-related genes, including Glutathione peroxidase 8, NADP-malic enzyme 2, Response to ABA and Salt 1, WRKYs, Glutathione S-Transferase, and MYBs, were induced by salt in transgenic plants. Moreover, the pathways of flavonoid biosynthesis and metabolic processes, regulation of response to stress, response to ethylene, dioxygenase activity, glucosyltransferase activity, monooxygenase activity, and oxidoreductase activity were specially enriched in transgenic plants under salt condition. Taken together, our results demonstrate that PtSAP13 enhances salt tolerance through up-regulating the expression of stress-related genes and mediating multiple biological pathways.

Genome-wide identification of stress-associated proteins (SAP) with A20/AN1 zinc finger domains associated with abiotic stresses responses in Brassica napus

Stress-associated proteins (SAP) that contain the A20/AN1 zinc-finger domain play important roles in plants abiotic stress responses. However, little is known about the SAP gene family in Brassica napus (genome AnAnCnCn), was formed by recent allopolyploidy between ancestors of Brassica rapa (genome ArAr) and Brassica oleracea (genome CoCo). To explore the distribution and expression patterns of these genes, a genome-wide identification and a systematic analysis of genes in the SAP family in B. napus were performed. A total of 57 genes encoding SAP were identified, and subsequently classified into 5 groups. In contrast to results seen in other plant species, further analyses of gene structures and conserved protein motifs revealed that a large percentage of the BnaSAP (16/57) genes were A20-type or AN1-type SAP genes, most of which were not found to have homologous genes in B. rapa or B. oleracea. The cis-elements of the BnaSAP promoters, and BnaSAP gene expression levels in different tissues, and under different stress treatments, were also investigated based on RNA-seq data. All the typical A20-AN1-type and AN1-AN1(-C2H2)-type BnaSAP genes were constitutively expressed among all tissues, except 4 members in Group II. In contrast, almost all A20 type and AN1 type members showed very low detection levels in all the tested rapeseed tissues from which the transcriptome data was derived. Additionally, most of the typical A20-AN1-type and AN1-AN1-type BnaSAP genes were found to be induced by multiple stresses and phytohormones. Among the identified BnaSAP genes, four members of Group III (BnaA07g15770D, BnaC06g14100D, BnaC04g27800D, and BnaC08g24010D) were induced following various low-temperature stresses in leaves, and were induced by NaCl and PEG stress in roots. BnaA06g02460D and BnaC06g04440D from Group IV were induced by heat stress, while BnaA06g31640D and BnaC07g25010D from Group V were induced by heat stress and salicylic acid (SA) treatment. This study provides a comprehensive analysis of SAP genes in rapeseed, and may aid in future efforts to identify the functions of A20/AN1-type proteins and the responses of rapeseed under a variety of abiotic stresses.

IDENTIFICATION OF GENES ENCODING STRESS-ASSOCIATED PROTEINS CONTAINING THE DOMAINS A20/AN1 IN THE APPLE GENOME IN SILICO AND ANALYSIS OF THEIR PHYLOGENETIC RELATIONSHIP

Stress-associated proteins (SAPs) are the transcription factors containing the zinc-finger domains A20 and/or AN1. Genome-wide identification and characterization of an apple variety of Golden Delicious stress-associated proteins was performed, potential SAPs were found. Each of them contains, at least, one zinc-finger domain AN1. A phylogenetic tree was constructed on the basis of the amino acid sequences of apple SAP and homologues from other species (dicotyledonous cotton and monocotyledonous rice) to assess their phylogenetic relationships. The sequences ahead of the translation initiation site of SAPs genes were analyzed to discover cis-elements involved in the gene expression regulation. The analysis shows that apple SAPs are involved in a complex network of regulatory elements that controls the vitality of plant cells. It can be assumed that the expression of SAPs in apple trees occurs constantly, but if conditions of a plant growth are changed, their level increases. Regulation of the SAP expression may depend on a plant ontogenesis stage and it may vary in different tissues.

Genome-Wide Analysis and Cloning of the Apple Stress-Associated Protein Gene Family Reveals MdSAP15, Which Confers Tolerance to Drought and Osmotic Stresses in Transgenic Arabidopsis

Stress-associated proteins (SAPs) are novel A20/AN1 zinc finger domain-containing proteins that are now favorable targets to improve abiotic stress tolerance in plants. However, the SAP gene family and their biological functions have not been identified in the important fruit crop apple (Malus × domestica Borkh.). We conducted a genome-wide analysis and cloning of this gene family in apple and determined that the overexpression of MdSAP15 enhances drought tolerance in Arabidopsis plants. We identified 30 SAP genes in the apple genome. Phylogenetic analysis revealed two major groups within that family. Results from sequence alignments and analyses of 3D structures, phylogenetics, genomics structure, and conserved domains indicated that apple SAPs are highly and structurally conserved. Comprehensive qRT-PCR analysis found various expression patterns for MdSAPs in different tissues and in response to a water deficit. A transgenic analysis showed that the overexpression of MdSAP15 in transgenic Arabidopsis plants markedly enhanced their tolerance to osmotic and drought stresses. Our results demonstrate that the SAP genes are highly conserved in plant species, and that MdSAP15 can be used as a target gene in genetic engineering approaches to improve drought tolerance.

A wheat gene TaSAP17-D encoding an AN1/AN1 zinc finger protein improves salt stress tolerance in transgenic Arabidopsis

The stress-associated protein (SAP) multigene family is conserved in both animals and plants. Its function in some animals and plants are known, but it is yet to be deciphered in wheat (Triticum aestivum L.). We identified the wheat gene TaSAP17-D, a member of the SAP gene family with an AN1/AN1 conserved domain. Subcellular localization indicated that TaSAP17-D localized to the nucleus, cytoplasm, and cell membrane. Expression pattern analyses revealed that TaSAP17-D was highly expressed in seedlings and was involved in NaCl response, polyethylene glycol (PEG), cold, and exogenous abscisic acid (ABA). Constitutive expression of TaSAP17-D in transgenic Arabidopsis resulted in enhanced tolerance to salt stress, confirmed by improved multiple physiological indices and significantly upregulated marker genes related to salt stress response. Our results suggest that TaSAP17-D is a candidate gene that can be used to protect crop plants from salt stress.

A dCAPS marker developed from a stress associated protein gene TaSAP7-B governing grain size and plant height in wheat

Stress associated proteins (SAPs) are the A20/AN1 zinc-finger proteins which confer to abiotic stresses in plants. In this study, TaSAP7-B, including two AN1 domains, was isolated from B genome of wheat (Triticum aestivum L.). Sequencing analysis on TaSAP7-B illustrated one InDel (insertion-deletion) and one SNP (single nucleotide polymorphism) in the promoter region while no diversity was observed in the coding region. On the basis of SNP in the promoter region (–260 bp), a dCAPS (derived cleaved amplified polymorphic sequences) marker SNP-260 was developed for TaSAP7-B. Using a natural population consisting of 262 wheat accessions, significant associations were detected between the marker SNP-260 and agronomic traits, such as plant height (PH), peduncle length (PL), length of penultimate internode (LPI), number of spike per plant (NSP), and 1 000-grain weight (TGW). Two genotypes were identified using marker SNP-260 in the natural population. Among them, the genotypes possessing C allele exhibited a higher TGW and shorter PH than the T genotypes. Hence, base C was considered as the superior allele. The dCAPS marker of TaSAP7-B can be instrumental for marker-assisted selection for high grain size and short plant height.

A stress-associated protein, AtSAP13, from Arabidopsis thaliana provides tolerance to multiple abiotic stresses.

Members of Stress-Associated Protein (SAP) family in plants have been shown to impart tolerance to multiple abiotic stresses, however, their mode of action in providing tolerance to multiple abiotic stresses is largely unknown. There are 14 SAP genes in Arabidopsis thaliana containing A20, AN1, and Cys2-His2 zinc finger domains. AtSAP13, a member of the SAP family, carries two AN1 zinc finger domains and an additional Cys2-His2 domain. AtSAP13 transcripts showed upregulation in response to Cd, ABA, and salt stresses. AtSAP13 overexpression lines showed strong tolerance to toxic metals (AsIII, Cd, and Zn), drought, and salt stress. Further, transgenic lines accumulated significantly higher amounts of Zn, but less As and Cd accumulation in shoots and roots. AtSAP13 promoter-GUS fusion studies showed GUS expression predominantly in the vascular tissue, hydathodes, and the apical meristem and region of root maturation and elongation as well as the root hairs. At the subcellular level, the AtSAP13-eGFP fusion protein was found to localize in both nucleus and cytoplasm. Through yeast one-hybrid assay, we identified several AP2/EREBP family transcription factors that interacted with the AtSAP13 promoter. AtSAP13 and its homologues will be highly useful for developing climate resilient crops.

Genome-wide identification and characterization of stress-associated protein (SAP) gene family encoding A20/AN1 zinc-finger proteins in Medicago truncatula

Stress associated proteins (SAPs) play important roles in developmental processes, responses to various stresses and hormone stimulation in plants. However, little is known about the SAP gene family in Medicago truncatula . In this study, a total of 17 MtSAP genes encoding A20/AN1 zinc-finger proteins were characterized. Out of these 17 genes, 15 were distributed over all 8 chromosomes at different densities, and two segmental duplication events were detected. The phylogenetic analysis of these proteins and their orthologs from Arabidopsis and rice suggested that they could be classified into five out of the seven groups of SAP family genes, with genes in the same group showing similar structures and conserved domains. The cis -elements of the Mt SAP promoters were studied, and many cis -elements related to stress and plant hormone responses were identified. We also investigated the stress-responsive expression patterns of the MtSAP genes under various stresses, including drought, exposure to NaCl and cold. The qRT-PCR results showed that numerous MtSAP genes exhibited transcriptional responses to multiple abiotic stresses. These results lay the foundation for further functional characterization of SAP genes. To the best of our knowledge, this is the first report of a genome-wide analysis of the SAP gene family in Medicago truncatula . https://doi.org/10.2298/ABS170529028Z Received: May 29, 2017; Revised: July 28, 2017; Accepted: July 31, 2017; Published online: August 24, 2017 How to cite this article: Zhou Y, Zeng L, Chen R, Wang Y, Song J. Genome-wide identification and characterization of stress-associated protein (SAP) gene family encoding A20/AN1 zinc-finger proteins in Medicago truncatula . Arch Biol Sci. 2018;70(1):87-98.

The LmSAP gene isolated from the halotolerant Lobularia maritima improves salt and ionic tolerance in transgenic tobacco lines.

The A20/AN1 zinc-finger domain-containing proteins of the stress-associated proteins (SAPs) family are fast emerging as potential candidates for biotechnological approaches to improve abiotic stress tolerance in plants. We identified LmSAP, one of the SAPs genes in Lobularia maritima (L.) Desv., a halophyte brassicaceae, through its transcript accumulation in response to salinity and ionic stresses. Sequence homology analysis revealed that LmSAP contains two conserved zinc-finger domains A20 and AN1. Phylogeny analyses showed that LmSAP exhibited high amino acid sequence identity to other plant SAPs. Heterologous expression of LmSAP in yeast increased cell tolerance to salt and osmotic stress. In addition, the overexpression of LmSAP conferred high salt and ionic tolerance to transgenic tobacco plants. Transgenic tobacco seedlings showed higher survival rates and antioxidant activities under salt and ionic stresses. Enhanced antioxidant activities paralleled lower malondialdehyde and superoxide anion O2- levels in the LmSAP transgenic seedlings. Overall, our results suggest that overexpression of LmSAP enhanced salt tolerance by maintaining ionic balance and limiting oxidative and osmotic stresses.

Genome-wide identification and expression analysis of stress-associated proteins (SAPs) containing A20/AN1 zinc finger in cotton.

Stress-associated proteins (SAPs) containing the A20/AN1 zinc-finger domain play important roles in response to both biotic and abiotic stresses in plants. Nevertheless, few studies have focused on the SAP gene family in cotton. To explore the distributions and expression patterns of these genes, we performed genome-wide identification and characterization of SAPs in tetraploid Gossypium hirsutum L. TM-1 (AD1). A total of 37 genes encoding SAPs were identified, 36 of which were duplicated in the A and D sub-genomes. The analysis of gene architectures and conserved protein motifs revealed that nearly all A20-AN1-type SAPs were intron-free, whereas AN1-AN1-type SAPs contained one intron. The cis-elements of the SAP promoters were studied, as were the expression levels of cotton SAP genes under different stresses based on RNA-seq data and validated by qRT-PCR. Most cotton SAP genes were induced by multiple stresses and phytohormones, particularly salt stress, indicating that SAP genes may play important roles in cotton's response to unfavorable environmental changes. Among these identified SAPs, the expression of GhSAP17A/D is suppressed in cotton response to Vertillium dahliae, and the GhSAP17A/D-silenced cotton exhibits more resistance to V. dahliae. This study provides insight into the evolution of SAP genes in upland cotton and may aid in efforts at further functional identification of A20/AN1-type proteins and cotton's response to different stresses.

Genome-wide survey and expression analysis of the stress-associated protein gene family in desert poplar, Populus euphratica

Stress-associated proteins (SAPs) are a novel class of zinc finger proteins that extensively participate in abiotic stress responses. To date, no overall analysis and expression profiling of SAP genes in woody plants have been reported. Populus euphratica is distributed in desert regions and is extraordinarily adaptable to abiotic stresses. Thus, it is regarded as a promising candidate for studying abiotic stress resistance mechanisms of woody plants. In this study, 18 non-redundant SAP genes were identified from the genome of P. euphratica using basic local alignment search tool algorithms and functional domain verification. Among these 18 PeuSAP genes, 15 were intronless. To investigate the evolutionary relationships of SAP genes in P. euphratica and other Salicaceae plants, phylogenetic analyses were performed. Subsequently, the expression profiles of the 18 PeuSAP genes were analyzed in different tissues and under various stresses (drought, salt, heat, cold, and abscisic acid (ABA) treatment) using quantitative real-time PCR. Tissue expression analysis indicated that PeuSAPs showed no tissue specificity. PeuSAPs were induced by multiple abiotic stresses, especially drought, salt, and heat stresses, perhaps because of abundant cis-acting heat shock elements and drought-inducible elements in the promoter regions of the PeuSAPs. Moreover, single nucleotide polymorphisms (SNPs) variant analysis revealed many synonymous and non-synonymous SNPs in PeuSAP genes, but the zinc finger structure was conserved during evolution. These results provide an overview of the SAP gene family in P. euphratica and a reference for further functional research on PeuSAP genes.

The promoter of the AlSAP gene from the halophyte grass Aeluropus littoralis directs a stress-inducible expression pattern in transgenic rice plants.

When fused to " Pr AlSAP " promoter, transcripts of gusA exhibited similar accumulation patterns in transgenic rice as AlSAP transcripts in A. littoralis. Pr AlSAP can be used for engineering abiotic stress tolerance. We previously showed that ectopic expression of a stress-associated protein gene from Aeluropus littoralis (AlSAP) enhances tolerance to multiple abiotic stresses in tobacco, wheat and rice. The ortholog of AlSAP in rice is OsSAP9. Here, we demonstrate that AlSAP transcripts accumulate in Aeleuropus in response to multiple abiotic stresses and at a higher level in roots, while those of OsSAP9 are preferentially induced by cold and heat treatments and accumulate preferentially in leaves of rice. In silico analysis of the AlSAP promoter "Pr AlSAP " predicted several cis-acting elements responsible for gene regulation by dehydration, salt, heat, ABA, SA, wounding and tissue-specific expression. The Pr AlSAP promoter was fused to the gusA gene and used to produce transgenic rice plants. Transcripts of gusA exhibited similar accumulation patterns in transgenic rice as AlSAP transcripts in A. littoralis. Indeed, accumulation of gusA transcripts was higher in roots than in leaves and induced by salt, drought, cold and heat treatments. GUS activity was confirmed in roots, coleoptiles, leaves and glumes, but absent in the root cell elongation zone and in dry seeds. A wound treatment strongly induced GUS accumulation in leaves and imbibed seeds. Altogether, these results indicate that the regulatory regions of two ortholog genes "AlSAP" and "OsSAP9" have diverged in the specificity of the signals promoting their induction, but that the trans-acting elements allowing the correct spatiotemporal regulation and stress induction of Pr AlSAP exist in rice. Therefore, the AlSAP promoter appears to be an interesting candidate for engineering abiotic stress tolerance in cereals.

Rice OsiSAP7 negatively regulates ABA stress signalling and imparts sensitivity to water-deficit stress in Arabidopsis

Stress associated protein (SAP) genes in plants regulate abiotic stress responses. SAP gene family consists of 18 members in rice. Although their abiotic stress responsiveness is well established, the mechanism of their action is poorly understood. OsiSAP7 was chosen to investigate the mechanism of its action based on the dual nature of its sub-cellular localization preferentially in the nucleus or sub-nuclear speckles upon transient expression in onion epidermal cells. Its expression was down-regulated in rice seedlings under abiotic stresses. OsiSAP7 was localized evenly in the nucleus under unstressed conditions and in sub-nuclear speckles on MG132 treatment. OsiSAP7 exhibits E3 ubiquitin ligase activity in vitro. Abiotic stress responses of OsiSAP7 were assessed by its overexpression in Arabidopsis under the control of a stress inducible promoter rd29A. Stress response assessment was done at seed germination and advanced stages of development. Transgenics were ABA insensitive at seed germination stage and sensitive to water-deficit stress at advanced stage as compared to wild type (WT). They were also impaired in ABA and stress-responsive gene expression. Our study suggests that OsiSAP7 acts as a negative regulator of ABA and water-deficit stress signalling by acting as an E3 ubiquitin ligase.

OsiSAP1 overexpression improves water-deficit stress tolerance in transgenic rice by affecting expression of endogenous stress-related genes.

OsiSAP1, an A20/AN1 zinc-finger protein, confers water-deficit stress tolerance at different stages of growth by affecting expression of several endogenous genes in transgenic rice. Transgenic lines have been generated from rice constitutively expressing OsiSAP1, an A20/AN1 zinc-finger containing stress-associated protein gene from rice, driven by maize UBIQUITIN gene promoter and evaluated for water-deficit stress tolerance at different stages of growth. Their seeds show early germination and seedlings grow better under water-deficit stress compared to non-transgenic (NT) rice. Leaves from transgenic seedlings showed lesser membrane damage and lipid peroxidation under water-deficit stress. Relatively lower rate of leaf water loss has been observed in detached intact leaves from transgenic plants during late vegetative stage. Delayed leaf rolling and higher relative water content were also observed in transgenic plants under progressive water-deficit stress during reproductive developmental stage. Although reduction in grain yield is observed under unstressed condition, the relative water-deficit stress-induced yield losses are lower in transgenic rice vis-à-vis NT plants thereby resulting in yield loss protection. Transcriptome analysis suggests that overexpression of OsiSAP1 in transgenic rice results in altered expression of several endogenous genes including those coding for transcription factors, membrane transporters, signaling components and genes involved in metabolism, growth and development. A total of 150 genes were found to be more than twofold up-regulated in transgenic rice of which 43 genes are known to be involved in stress response. Our results suggest that OsiSAP1 is a positive regulator of water-deficit stress tolerance in rice.

A novel stress-associated protein SbSAP14 from Sorghum bicolor confers tolerance to salt stress in transgenic rice

We have characterized a member of the stress-associated protein (SAP) gene family from Sorghum bicolor (SbSAP14) with A20 and AN1 zinc-finger domains. Expression profiling revealed that SbSAP14 is specifically induced in response to dehydration, salt, and oxidative stress as well as abscisic acid treatment. During the early stage of salt stress, overexpression of SbSAP14 was able to prevent yellowing and withering of the leaf tip of rice plants. Measurements of malondialdehyde, ion leakage, and chlorophyll content demonstrated that transgenic rice had an enhanced tolerance to oxidative damage caused by salt stress. Under prolonged salt stress, transgenic rice plants had a higher seed germination rate and higher percentage seedling survival than wild-type (WT) plants. Importantly, in vivo and in situ assays revealed that the accumulation of reactive oxygen species in transgenic rice plants was significantly lower than that in WT plants. Among the six antioxidant genes tested, APX2, CatB, CatC, and SodA1 showed a higher expression level in transgenic rice than in WT rice. Based on these results, we propose that SbSAP14 may play a key role in antioxidant defense systems and possibly be involved in the induction of antioxidant genes in plants, suggesting a possible mechanism of the SAP gene family in stress defense response.

MusaSAP1, a A20/AN1 zinc finger gene from banana functions as a positive regulator in different stress responses

A20/AN1 zinc finger domain containing Stress Associated Proteins (SAP) are involved in diverse stress response pathways in plants. In the present study, a novel banana SAP gene, MusaSAP1, was identified from banana EST database and was subsequently characterized by overexpression in transgenic banana plants. Expression profiling in native banana plants showed that MusaSAP1 was up-regulated by drought, salt, cold, heat and oxidative stress as well as by treatment with abscisic acid. Cellular localization assay carried out by making a MusaSAP1::GFP fusion protein indicated that MusaSAP1 is incompletely translocated to nucleus. Copy number analysis performed using real time PCR and Southern blotting indicated that MusaSAP1 occurs in the banana genome in a single copy per 11 chromosome set. Transgenic banana plants constitutively overexpressing MusaSAP1 displayed better stress endurance characteristics as compared to controls in both in vitro and ex vivo assays. Lesser membrane damage as indicated by reduced malondialdehyde levels in transgenic leaves subjected to drought, salt or oxidative stress pointed towards significant role for MusaSAP1 in stress amelioration pathways of banana. Strong up-regulation of a polyphenol oxidase (PPO) coding transcript in MusaSAP1 overexpressing plants together with induction of MusaSAP1 by wounding and methyl jasmonate treatment indicated possible involvement of MusaSAP1 in biotic stress responses where PPOs perform major functions in multiple defense pathways.

Overexpression of a Medicago truncatula stress-associated protein gene (MtSAP1) leads to nitric oxide accumulation and confers osmotic and salt stress tolerance in transgenic tobacco

The impact of Medicago truncatula stress-associated protein gene (MtSAP1) overexpression has been investigated in Nicotiana tabacum transgenic seedlings. Under optimal conditions, transgenic lines overexpressing MtSAP1 revealed better plant development and higher chlorophyll content as compared to wild type seedlings. Interestingly, transgenic lines showed a stronger accumulation of nitric oxide (NO), a signaling molecule involved in growth and development processes. This NO production seemed to be partially nitrate reductase dependent. Due to the fact that NO has been also reported to play a role in tolerance acquisition of plants to abiotic stresses, the responses of MtSAP1 overexpressors to osmotic and salt stress have been studied. Compared to the wild type, transgenic lines were less affected in their growth and development. Moreover, NO content in MtSAP1 overexpressors was always higher than that detected in wild seedlings under stress conditions. It seems that this better tolerance induced by MtSAP1 overexpression could be associated with this higher NO production that would enable seedlings to reach a high protection level to prepare them to cope with abiotic stresses.

Arabidopsis SAP5 functions as a positive regulator of stress responses and exhibits E3 ubiquitin ligase activity

AtSAP5, one of approximately 14 members of the Stress Associated Protein gene family in Arabidopsis, was identified by its expression in response to salinity, osmotic, drought and cold stress. AtSAP5 shows strong homology to OSISAP1, an A20/AN1-type zinc finger protein implicated in stress tolerance in rice. To evaluate the function of AtSAP5 in the regulation of abiotic stress responses, transgenic Arabidopsis plants that over-express AtSAP5 (35S::AtSAP5) were characterized, along with wild-type and T-DNA knock-down plants. Plants that over-express AtSAP5 showed increased tolerance to environmental challenges including salt stress, osmotic stress and water deficit. Comparison of gene expression patterns between 35S::AtSAP5 transgenic plants and wild-type plants under normal conditions and water deficit stress indicated that over-expression of AtSAP5 correlates with up-regulation of drought stress responsive gene expression. Analysis of transgenic plants that express GFP-AtSAP5 showed that it is localized primarily in nuclei of root cells and recombinant AtSAP5 has E3 ubiquitin ligase activity in vitro. These results indicate that AtSAP5 has E3 ligase activity and acts as a positive regulator of stress responses in Arabidopsis.

Characterization and phylogenetic analysis of environmental stress-responsive SAP gene family encoding A20/AN1 zinc finger proteins in tomato

Characterization of genes responsive to stress is important for efforts on improving stress tolerance of plants. To address components involved in stress tolerance of tomato (Solanum lycopersicum), a stress-responsive gene family encoding A20/AN1 zinc finger proteins was characterized. In the present study, 13 members of this gene family were cloned from tomato cultivar Pusa Ruby and named as Stress Associated Protein (SAP) genes. Out of 13 genes, 12 have been mapped on their respective chromosomes. Expression of these genes in response to cold, heat, salt, desiccation, wounding, abscisic acid, oxidative and submergence stresses was analysed. All tomato SAP genes were found to be responsive to one or other type of environmental stress. The phylogenetic analysis of these genes, along with their orthologs from Solanaceae species suggests the presence of a common set of SAP genes in the studied Solanaceae species. The present study characterizes a SAP gene family, which encodes A20/AN1 zinc finger containing proteins from tomato for the first time. Genes showing high expression in response to a particular stress can be exploited for improving stress tolerance of tomato and other Solanaceae members.