Differential expression and regulation of sucrose transporters in rice (Orzya sativa L, cv Nipponbare) during environmental stress conditions

Plant productivity is greatly affected by environmental stresses such as drought, salinity and insect herbivory. Plants respond and adapt to these stresses by exhibiting physiological as well as biochemical changes at the cellular and molecular levels in order to survive. Expression of a variety of genes which encode numerous membrane transporters have been demonstrated to be induced by these stresses in a variety of plants. The nutritional status of plants is controlled by these transporters, which are regulated by the transcription of the corresponding genes. In spite of these adverse stress effects on agricultural yield, only a few studies have focused on gene transcriptional and translational regulation of membrane transporters during environmental stress situations. Rice, like other plants, contains a number of sucrose transporters encoded by a family of genes. However, detailed knowledge of their roles, localization and regulation during environmental stress conditions is lacking. Bioinformatic tools were used to identify putative cis-acting regulatory elements that may be involved in the regulation of rice and Arabidopsis thaliana sucrose transporters. The possible cis-acting regulatory elements were predicted by scanning genomic sequences 1.5 kbp upstream of the sucrose transporter genes translational start sites, using Plant CARE, PLACE and Genomatix Matinspector professional data bases. Several cis-acting regulatory elements that are associated with plant development, plant hormonal regulation and stress response were identified, and were present in varying frequencies within the 1.5 kbp of 5′ regulatory region. The putative cis-acting regulatory elements that possibly are involved in the expression and regulation of sucrose transporter gene families in rice and Arabidopsis thaliana during cellular development or environmental stress conditions were identified as: A-box, RY, CAT, Pyrimidine-box, Sucrose-box, ABRE, ARF, ERE, GARE, Me-JA, ARE, DRE, GA-motif, GATA, GT-1, MYC, MYB, W-box, and I-box. Expression analysis was used to elucidate the role of rice (Oryza sativa L. cv Nipponbare) sucrose transporter (OsSUT) genes during drought and salinity treatments of three week old rice plants ( at four leaf stage) over a 10 days. Among the five rice OsSUT genes identified, only OsSUT2 was observed to be progressively up-regulated during drought and salinity treatments, while OsSUT1, OsSUT4 and OsSUT5 were expressed at low levels, and OsSUT3 showed no detectable transcript expression. Sucrose transport will be essential to meet the cellular energy demands and also for osmoprotectant activities during drought and salinity stresses. It therefore indicates that OsSUT2 which facilitates transport of sucrose from photosynthetic cells will be III essential for rice plants to cope with drought and salinity stresses, and cultivars with a higher OsSUT2 expression should be able to tolerate these environmental stresses better. The role of OsSUT in assimilate transport during rusty plum aphids (Hysteroneura setariae; Thomas) infestation on the leaves of three week old rice (Orzya sativa L. cv Nipponbare) cultivar plants, over a time-course of 1 to 10 days of treatments, was also examined by combination of gene expression and β-glucuronidase (GUS) reporter gene analysis. Real Time PCR analysis of the five OsSUT genes revealed that the expression of OsSUT1 was progressively up-regulated during the course of aphid infestation. OsSUT2 and OsSUT4 expression were comparatively low in both the control and treated plants. OsSUT5 showed no clear difference in transcript expression in both control and treated plants, while no detectable transcript expression of OsSUT3 could be found. The up-regulation of OsSUT1 gene was verified at protein level by western blot analysis in both the control and treated plants. OsSUT1 protein expression was found to increase with time during aphid infestation. A similar trend was noticeable in the control plants, however at a lower expression level. These demonstrate that the cellular expression of OsSUT1is regulated by both developmental and environmental factors. OsSUT1-promoter:::GUS reporter gene expression was observed within the vascular parenchyma and/or companion cells associated with phloem sieve elements of the large and small bundles in the phloem tissues of the flag leaf blade regions where feeding aphids were confined, which progressively increased with time of infestation. It is suggested that OsSUT1 may primarily play an essential role in phloem transport of assimilate to wounded tissues from adjacent health tissues or may be involved in the retrieval of assimilate back into the phloem to minimize loss caused by the infestation. Some OsSUT1-promoter:::GUS expression was also found in the metaxylem at 10 days after infestation, which could signify a recovery system in which sucrose lost into the xylem as a result of aphids feeding are retrieved back into the phloem through the vascular parenchyma. This was supported by the exposure of cut ends of matured OsSUT1-promoter:::GUS rice plant leaf to 2% sucrose solution. OsSUT1-promoter:::GUS expression was observed within the protoxylem, xylem and phloem parenchyma tissues. This indicates that sucrose translocating within the xylem tissues are retrieved into the phloem via the OsSUT1 localized within the parenchyma tissues. In conclusion, the differential expression and regulation of rice (Orzya sativa L. cv Nipponbare) sucrose transporters as reported here suggest that OsSUT2 and OsSUT1 were constitutively expressed compared to other rice sucrose transporters during drought and salinity, and rusty plum aphids (Hysteroneura setariae; Thomas) infestation stresses respectively. Thus, the expression and regulation of the sucrose transporters could be related to the physiological and nutritional requirements of the cells during plant developmental or environmental stress state that allows their differential expression

Semi-global Analysis of the Early Cold Stress Response Transcriptome of Developing Seedlings of Rice (Oryzasativa L.,japonica)

Plants are either sensitive or insensitive to low temperatures. Cool-season species acclimate to chilling and develop tolerance to freezing. Warm-season species vary in the degree of sensitivity to chilling and are not capable of acclimation. Both freezing and chilling tolerance involve the activity of an intricately coordinated network of genes. The regulatory network that defines cold acclimation and freezing tolerance is well understood in Arabidopsis. The centerpiece of this network is a group of transcriptional activators that coordinate a battery of downstream defense-related genes. In contrast, little is known about the corresponding regulon in non-acclimating species. This study utilized the available tools of functional genomics and genome sequence resources of rice (Oryza sativa L.), a chilling-sensitive species, to examine the cold stress transcriptome of young seedlings of a relatively tolerant japonica cultivar (CT6748-8-CA-17). The goal was to use rice as a model towards understanding the dynamics of the early response regulatory network through a survey of gene expression changes during the critical first 24 hours of stress. The expression of 1,550 unique genes represented as spotted cDNA probes on a microarray was profiled by interrogation with a pair of control (28°C) and stressed (10°C) RNA isolated after 0.5, 2, 6, 12 and 24 hours. The expression data showed that early response involves two waves of induction and repression. The first wave started within the first 2 hours of stress, hence the genes were collectively designated as \u27rapidly induced/repressed early response genes\u27 (Group-I). The second wave did not start until after 2 hours of stress, hence the genes were collectively designated as \u27delayed induced/repressed early response genes\u27 (Group-II). The functional categories of genes overlap between Groups-I and II. In general, balanced upregulation and downregulation of genes involved in signal transduction, growth and development, metabolism, transport, protein synthesis, modification and degradation reflects the \u27physiologically stressed\u27 status of the plant, consistent with the initial reduction in growth rate. Early gene expression responses also suggest an active mechanism by which cellular resources are temporarily being redirected from growth related processes to overall physiological adjustments and early defenses. Potential regulators of early responses were also identified in the survey. The first is a novel cold stress-related bZIP transcription factor (OsbZIP) induced rapidly but transiently within the first 2 to 6 hours. Other potential early regulators are transiently expressed after 6 hours. These include a bHLH transcription (OslCE)factor similar to the ICE1 of Arabidopsis, a Myb protein similar to the previously identified cold stress response regulator Osmyb4, and a C3HC4 zinc-finger protein related to HOSl of Arabidopsis. The layered fashions by which these transcription factors are expressed suggest a mechanism for fine-tuned regulation of the early response genetic network. The transcriptome survey also suggests that ABA is not essential in the early responses and other molecules such as H2O2 are probably involved. The results of this study set the stage for future investigation of the entire pathway by a combinatorial approach that includes genome-wide transcript profiling, promoter-reporter assay, protein-DNA interaction analysis and reverse genetics

The Medicago truncatula sucrose transporter family

In plants, long distance transport of sugars from photosynthetic source leaves to sink organs comprises different crucial steps depending on the species and organ types. Sucrose, the main carbohydrate for long distance transport is synthesized in the mesophyll and then loaded into the phloem. After long distance transport through the phloem vessels, sucrose is finally unloaded towards sink organs. Alternatively, sugar can also be transferred to non-plant sinks and plant colonization by heterotrophic organisms increases the sink strength and creates an additional sugar demand for the host plant. These sugar fluxes are coordinated by transport systems. Main sugar transporters in plants comprise sucrose (SUTs) and monosaccharide (MSTs) transporters which constitute key components for carbon partitioning at the whole plant level and in interactions with fungi. Although complete SUTs and MSTs gene families have been identified from the reference Dicot Arabidopsis thaliana and Monocot rice (Oriza sativa), sugar transporter families of the leguminous plant Medicago truncatula, which represents a widely used model for studying plant-fungal interactions in arbuscular mycorrhiza (AM), have not yet been investigated. With the recent completion of the M. truncatula genome sequencing as well as the release of transcriptomic databases, monosaccharide and sucrose transporter families of M. truncatula were identified and now comprise 62 MtMSTs and 6 MtSUTs. I focused on the study of the newly identified MtSUTs at a full family scale; phylogenetic analyses showed that the 6 members of the MtSUT family distributed in all three Dicotyledonous SUT clades; they were named upon phylogenetic grouping into particular clades: MtSUT1-1, MtSUT1-2, MtSUT1-3, MtSUT2, MtSUT4-1 and MtSUT4-2. Functional analyses by yeast complementation and expression profiles obtained by quantitative RT-PCR revealed that MtSUT1-1 and MtSUT4-1 are H+/sucrose symporters and represent key members of the MtSUT family. Conservation of transport capacity between orthologous leguminous proteins, expression profiles and subcellular localization compared to previously characterized plant SUTs indicate that MtSUT1-1 is the main protein involved in phloem loading in source leaves whilst MtSUT4-1 mediates vacuolar sucrose export for remobilization of intracellular reserve. The AM symbiosis between plants and fungi from the phylum Glomeromycota is characterized by trophic exchanges between the two partners. The fungus supplies the autotrophic host with nutrients and thereby promotes plant growth. In return, the host plant provides photosynthate (sugars) to the heterotrophic symbiont. Here, sugar fluxes from plant source leaves towards colonized sink roots in the association between the model leguminous plant M. truncatula and the arbuscular mycorrhizal fungus (AMF) Glomus intraradices were investigated. Sugar transporter candidates from both the plant and fungal partners presenting differential expression profiles using available transcriptomic tools were pinpointed. Gene expression profiles of MtSUTs and sugar quantification analyses upon high and low phosphorus nutrient supply and inoculation by the AMF suggest a mycorrhiza-driven stronger sink in AM roots with a finetuning regulation of MtSUT gene expression. Conserved regulation patterns were observed for orthologous SUTs in response to colonization by glomeromycotan fungi. In parallel, a non-targeted strategy consisting in the development of a M. truncatula - G. intraradices expression library suitable for yeast functional complementation and screening of symbiotic marker genes, similar to the approach that led to the identification of the first glomeromycotan hexose transporter (GpMST1), has been developed in this study. Taken together, with the identification, functional characterization and gene expression pattern of sugar transporter families, a more complete picture of sugar fluxes in the AM symbiosis has begun to emerge. This study opens new perspectives by identifying interesting candidate genes involved in sugar partitioning at both the plant and fungal levels and at the symbiotic interface in the widely used AM symbiosis model between M. truncatula and G. intraradices

In silico study of cis-acting elements revealing the plastid gene involved in oxidative phosphorylation are responsive to abiotic stresses

In order to study plastid gene response to abiotic stresses, the chloroplast genome of Brassica nigra and studied cis-acting elements were downloaded. All upstream regions of genes were determined and searched for the presence of known cis-acting elements. In these regions, 83 types of cis-acting elements were recognized. Unnamed elements (139 times), CAAT-box (96 times), and TATA-box (92 times) were in high frequency, whereas ATCC-motif, Box III, CE1, CE3, C-repeat/DRE, E2Fb, Gap-box, L-box, RY-element, and TGA-box occurred only one time. All of the cis-acting elements were grouped into seven categories, which 17% of cis-acting elements placed into abiotic and biotic-related elements. ARE (31 times) and LTR (21 times) elements were in high frequency. Among 42 genes with abiotic stress-related elements, 29 genes showed co-expression. Our results show that in response to anaerobic conditions and cold stress, chloroplast alters the genes-encoding proteins involved in complex I and V in oxidative phosphorylation pathway. This process, probably, is to reduce electron flow and convert NADPH and FADH forms to ATP form. These actions could decrease generating reactive oxygen species under stressful conditions. These findings could offer new insights on the strategies which chloroplasts take into account for preventing oxidative damage

Stress-Mediated cis-Element Transcription Factor Interactions Interconnecting Primary and Specialized Metabolism in planta

Plant specialized metabolites are being used worldwide as therapeutic agents against several diseases. Since the precursors for specialized metabolites come through primary metabolism, extensive investigations have been carried out to understand the detailed connection between primary and specialized metabolism at various levels. Stress regulates the expression of primary and specialized metabolism genes at the transcriptional level via transcription factors binding to specific cis-elements. The presence of varied cis-element signatures upstream to different stress-responsive genes and their transcription factor binding patterns provide a prospective molecular link among diverse metabolic pathways. The pattern of occurrence of these cis-elements (overrepresentation/common) decipher the mechanism of stress-responsive upregulation of downstream genes, simultaneously forming a molecular bridge between primary and specialized metabolisms. Though many studies have been conducted on the transcriptional regulation of stress-mediated primary or specialized metabolism genes, but not much data is available with regard to cis-element signatures and transcription factors that simultaneously modulate both pathway genes. Hence, our major focus would be to present a comprehensive analysis of the stress-mediated interconnection between primary and specialized metabolism genes via the interaction between different transcription factors and their corresponding cis-elements. In future, this study could be further utilized for the overexpression of the specific transcription factors that upregulate both primary and specialized metabolism, thereby simultaneously improving the yield and therapeutic content of plants

Influence of the ethylene on the grape berry development and related-gene expression

Le raisin est un fruit non climactérique dont la maturation semble ne pas nécessiter l'éthylène. Ici, il est confirmé que l’accumulation d’anthocyanes est liée à l’accumulation d’une glucoslyltransferase (UFGT), dont le promoteur a été cloné. Il a été trouvé 7 cis -éléments éthylène-dépendants. Cette étude a montré la stimulation d'expression de l'ufgt par l’éthylène n'est pas dépendant de MybA, un des régulateurs de la transcription de l'ufgt. Des expériences ont été conçues pour étudier l’ensemble des gènes affecté par éthylène au début de la maturation du raisin. Parmi eux certains sont en relation avec les variations de diamètre de la baie. Ces gènes sont impliqués dans la circulation de l'eau : plusieurs aquaporines, et dans la structure de la paroi cellulaire : polygalactoronases, xyloglucan endotransglucosylases, pectine méthylesterase, cellulose synthase et expansines. L’éthylène stimule l'accumulation de la plupart de leurs transcrits entre 1 heure et 24 heures d’incubation. ABSTRACT : The grape is a non-climacteric fruit which maturation apparently does not require ethylene. Here, it is confirmed that the accumulation of anthocyanins is linked to the accumulation of a glucoslyltransferase (UFGT), whose promoter was cloned. We found 7 cis-elements ethylene-dependent. This study showed the stimulation of ufgt expression by ethylene is not dependent to MybA, transcription regulators of the ufgt. Experiments were designed to investigate all genes affected by ethylene in early ripening grapes. Among them some are in relation to variations in berry diameter. These genes are involved in the movement of water : several aquaporins, and the structure of the cell wall : polygalactoronases, xyloglucan endotransglucosylases, méthylesterase pectin, cellulose synthase and expansines. Ethylene stimulates the accumulation of most of their transcripts between 1 hour and 24 hours of incubation

Sistema de informação cis-elementos.

o objetivo deste estudo foi desenvolver um sistema computacional capaz de identificar a presença de cis-elementos conhecidos, utilizando como modelo a família de genes ERDs (Early Responsive to Dehydration) expressos por Arabidopsis thaliana durante estresse hídrico (TAJI et al., 1999). Este sistema computacional também pode ser usado para outras culturas, como o feijão, arroz, trigo, etc.CONAFE

Plant Responses and Tolerance to Salt Stress: Physiological and Molecular Interventions

Overall, the 19 contributions in this Special Issue “Plant Responses and Tolerance to Salt Stress: Physiological and Molecular Interventions” discuss the various aspects of salt stress responses in plants. It also discusses various mechanisms and approaches to conferring salt tolerance on plants. These types of research studies provide further directions in the development of crop plants for the saline environment in the era of climate change

Roles of aquaporins in Setaria viridis stem development and sugar storage

Setaria viridis is a C4 grass used as a model for bioenergy feedstocks. The elongating internodes in developing S. viridis stems grow from an intercalary meristem at the base, and progress acropetally toward fully expanded cells that store sugar. During stem development and maturation, water flow is a driver of cell expansion and sugar delivery. As aquaporin proteins are implicated in regulating water flow, we analyzed elongating and mature internode transcriptomes to identify putative aquaporin encoding genes that had particularly high transcript levels during the distinct stages of internode cell expansion and maturation. We observed that SvPIP2;1 was highly expressed in internode regions undergoing cell expansion, and SvNIP2;2 was highly expressed in mature sugar accumulating regions. Gene co-expression analysis revealed SvNIP2;2 expression was highly correlated with the expression of five putative sugar transporters expressed in the S. viridis internode. To explore the function of the proteins encoded by SvPIP2;1 and SvNIP2;2, we expressed them in Xenopus laevis oocytes and tested their permeability to water. SvPIP2;1 and SvNIP2;2 functioned as water channels in X. laevis oocytes and their permeability was gated by pH. Our results indicate that SvPIP2;1 may function as a water channel in developing stems undergoing cell expansion and SvNIP2;2 is a candidate for retrieving water and possibly a yet to be determined solute from mature internodes. Future research will investigate whether changing the function of these proteins influences stem growth and sugar yield in S. viridis.Samantha A. McGaughey, Hannah L. Osborn, Lily Chen, Joseph L. Pegler, Stephen D. Tyerman, Robert T. Furbank, Caitlin S. Byrt and Christopher P. L. Gro