Using euhalophytes to understand salt tolerance and to develop saline agriculture: Suaeda salsa as a promising model

Background As important components in saline agriculture, halophytes can help to provide food for a growing world population. In addition to being potential crops in their own right, halophytes are also potential sources of salt-resistance genes that might help plant breeders and molecular biologists increase the salt tolerance of conventional crop plants. One especially promising halophyte is Suaeda salsa, a euhalophytic herb that occurs both on inland saline soils and in the intertidal zone. The species produces dimorphic seeds: black seeds are sensitive to salinity and remain dormant in light under high salt concentrations, while brown seeds can germinate under high salinity (e.g. 600 mm NaCl) regardless of light. Consequently, the species is useful for studying the mechanisms by which dimorphic seeds are adapted to saline environments. S. salsa has succulent leaves and is highly salt tolerant (e.g. its optimal NaCl concentration for growth is 200 mm). A series of S. salsa genes related to salt tolerance have been cloned and their functions tested: these include SsNHX1, SsHKT1, SsAPX, SsCAT1, SsP5CS and SsBADH. The species is economically important because its fresh branches have high value as a vegetable, and its seed oil is edible and rich in unsaturated fatty acids. Because it can remove salts and heavy metals from saline soils, S. salsa can also be used in the restoration of salinized or contaminated saline land.Scope Because of its economic and ecological value in saline agriculture, S. salsa is one of the most important halophytes in China. In this review, the value of S. salsa as a source of food, medicine and forage is discussed. Its uses in the restoration of salinized or contaminated land and as a source of salt-resistance genes are also considered.     

Wide variation in post-emergence desiccation tolerance of seedlings of fynbos proteoid shrubs

Fynbos Proteaceae that are killed by fire and bear their seeds in serotinous cones (proteoids), are entirely dependent on seedling recruitment for persistence. Hence, the regeneration phase represents a vulnerable stage of the plant life cycle. In laboratory-based experiments we investigated the effect of desiccation on the survival of newly emerged seedlings of 23 proteoid species (Leucadendron and Protea) occurring in a wide variety of fynbos habitats. We tested the hypothesis that species of drier habitats would be more tolerant of desiccation than those from more moist areas. Results showed that with no desiccation treatment, or with desiccation prior to radicle emergence, all species germinated to high levels. However, with desiccation treatments imposed after radicle emergence, there were significant declines in seedling emergence after subsequent re-wetting. Furthermore, other than three species that grow in waterlogged habitats, germination responses could not be reliably modeled as a function of soil moisture variables. An important finding was that the species had highly individualistic responses to desiccation. In conclusion, early seedling emergence represents a species-specific stage that is highly sensitive to a decrease in soil moisture. Since species are killed by fire (non-sprouting), vulnerability to increasing aridity associated with anthropomorphic climate change would increase the odds of local and global extinction.     

Mutants as tools to understand cellular and molecular drought tolerance mechanisms

The use of mutants: a most promising way to detect genes involved in development or in response to environmental stress. The model species Arabidopsis, particularly amenable to dissect the genetics and molecular mechanisms underlying physiological responses, also offers the advantage of a wide variety of mutants. As far as drought tolerance is concerned, hormonal mutants, impaired in hormone biosynthesis — deficient mutants — or in the signal transduction pathway—responsive mutants—provide a valuable tool to analyse the role of phytohormone interaction in the plant drought behaviour as well as to differentiate the mutant phenotypes with new criteria.These two categories of mutants (in particular the abscisic acid, ABA, mutants) were shown to be affected in developmental processes during seed maturation-in the desiccation phase- and/or in response to environmental stress (drought, ...) in vegetative tissues. The present report will focus on this last aspect: alterations in drought responses in vegetative tissues (adaptive strategies and drought tolerance mechanisms) essentially in Arabidopsis hormonal mutants (ABA-deficient and ABA-insensitive, GA-deficient, auxin and ethylene-insensitive).Some of the results are discussed with regard to the predicted functions of genes affected by the mutations.     

Zebrafish as a model to understand autophagy and its role in neurological disease

In the past decade, the zebrafish (Danio rerio) has become a popular model system for the study of vertebrate development, since the embryos and larvae of this species are small, transparent and undergo rapid development ex utero, allowing in vivo analysis of embryogenesis and organogenesis. These characteristics can also be exploited by researchers interested in signaling pathways and disease processes and, accordingly, there is a growing literature on the use of zebrafish to model human disease. This model holds great potential for exploring how autophagy, an evolutionarily conserved mechanism for protein degradation, influences the pathogeneses of a range of different human diseases and for the evaluation of this pathway as a potential therapeutic strategy. Here we summarize what is known about the regulation of autophagy in eukaryotic cells and its role in neurodegenerative disease and highlight how research using zebrafish has helped further our understanding of these processes.     

Sugars and desiccation tolerance in seeds

Soluble sugars have been shown to protect liposomes and lobster microsomes from desiccation damage, and a protective role has been proposed for them in several anhydrous systems. We have studied the relationship between soluble sugar content and the loss of desiccation tolerance in the axes of germinating soybean (Glycine max L. Merr. cv Williams), pea (Pisum sativum L. cv Alaska), and corn (Zea mays L. cv Merit) axes. The loss of desiccation tolerance during imbibition was monitored by following the ability of seeds to germinate after desiccation following various periods of preimbibition and by following the rates of electrolyte leakage from dried, then rehydrated axes. Finally, we analyzed the soluble sugar contents of the axes throughout the transition from desiccation tolerance to intolerance. These analyses show that sucrose and larger oligosaccharides were consistently present during the tolerant stage, and that desiccation tolerance disappeared as the oligosaccharides were lost. The results support the idea that sucrose may serve as the principal agent of desiccation tolerance in these seeds, with the larger oligosaccharides serving to keep the sucrose from crystallizing.     

Yeast as a model to understand the interaction between genotype and the response to calorie restriction

Calorie restriction is reported to enhance survival and delay the onset of age-related decline in many different species. Several proteins have been proposed to play a role in mediating the response to calorie restriction, including the target of rapamycin kinase, sirtuins, and AMP kinase. An enhanced mechanistic understanding of calorie restriction has popularized the concept of "calorie restriction mimetics", drugs that mimic the beneficial effects of caloire restriction without requiring a reduction in nutrient intake. In theory, such drugs should delay the onset and progression of multiple age-related diseases, similar to calorie restriction in mammals. Despite the potential benefits of such calorie restriction mimetics, however, relatively little is known about the interaction between genetic variation and individual response to calorie restriction. Limited evidence from model systems indicates that genotype plays a large role in determining both the magnitude and direction of effect that calorie restriction has on longevity. Here we present an overview of these data from the perspective of using yeast as a model to study aging and describe an approach we are taking to further characterize the molecular mechanisms underlying genotype-dependent responses to calorie restriction.     

Using zebrafish as the model organism to understand organ regeneration

The limited regenerative capacity of several organs, such as central nervous system(CNS), heart and limb in mammals makes related major diseases quite difficult to recover. Therefore, dissection of the cellular and molecular mechanisms underlying organ regeneration is of great scientific and clinical interests. Tremendous progression has already been made after extensive investigations using several model organisms for decades. Unfortunately, distance to the final achievement of the goal still remains. Recently, zebrafish became a popular model organism for the deep understanding of regeneration based on its powerful regenerative capacity, in particular the organs that are limitedly regenerated in mammals. Additionally, zebrafish are endowed with other advantages good for the study of organ regeneration. This review summarizes the recent progress in the study of zebrafish organ regeneration, in particular regeneration of fin, heart, CNS, and liver as the representatives. We also discuss reasons of the reduced regenerative capacity in higher vertebrate, the roles of inflammation during regeneration, and the difference between organogenesis and regeneration.     

Glass formation and desiccation tolerance in seeds

The formation of intracellular glass may help protect embryos from damage due to desiccation. Soluble sugars similar to those found in desiccation tolerant embryos were studied with differential scanning calorimetry. Those sugars from desiccation tolerant embryos can form glasses at ambient temperatures, whereas those from embryos that do not tolerate desiccation only form glasses at subzero temperatures. It is concluded that tolerant embryo cells probably contain sugar glasses at storage temperatures and water contents, but intolerant embryo cells probably do not.     

Rate of dehydration and cumulative desiccation stress interacted to modulate desiccation tolerance of recalcitrant cocoa and ginkgo embryonic tissues

Rate of dehydration greatly affects desiccation tolerance of recalcitrant seeds. This effect is presumably related to two different stress vectors: direct mechanical or physical stress because of the loss of water and physicochemical damage of tissues as a result of metabolic alterations during drying. The present study proposed a new theoretic approach to represent these two types of stresses and investigated how seed tissues responded differently to two stress vectors, using the models of isolated cocoa (Theobroma cacao) and ginkgo (Ginkgo biloba) embryonic tissues dehydrated under various drying conditions. This approach used the differential change in axis water potential (DeltaPsi/Deltat) to quantify rate of dehydration and the intensity of direct physical stress experienced by embryonic tissues during desiccation. Physicochemical effect of drying was expressed by cumulative desiccation stress [integralf(psi,t)], a function of both the rate and time of dehydration. Rapid dehydration increased the sensitivity of embryonic tissues to desiccation as indicated by high critical water contents, below which desiccation damage occurred. Cumulative desiccation stress increased sharply under slow drying conditions, which was also detrimental to embryonic tissues. This quantitative analysis of the stress-time-response relationship helps to understand the physiological basis for the existence of an optimal dehydration rate, with which maximum desiccation tolerance could be achieved. The established numerical analysis model will prove valuable for the design of experiments that aim to elucidate biochemical and physiological mechanisms of desiccation tolerance.     

Freezing and desiccation tolerance of imbibed canola seed

Depending on the environmental conditions, imbibed seeds survive subzero temperatures either by supercooling or by tolerating freezing-induced desiccation. We investigated what the predominant survival mechanism is in freezing canola ( Brassica napus cv. Quest) and concluded that it depends on the cooling rate. Seeds cooled at 3°C h⁻¹ or faster supercooled, whereas seeds cooled over a 4-day period to −12°C and then cooled at 3°C h⁻¹ to−40°C did not display low temperature exotherms. Both differential thermal analysis and nuclear magnetic resonance (NMR) spectroscopy confirmed that imbibed canola seeds undergo freezing-induced desiccation at slow cooling rates. The freezing tolerance of imbibed canola seed (LT₅₀) was determined by slowly cooling to −12°C for 48 h, followed with cooling at 3°C h⁻¹ to −40°C, or by holding at a constant −6°C (LD₅₀). For both tests, the loss in freezing tolerance of imbibed seeds was a function of time and temperature of imbibition. Freezing tolerance was rapidly lost after radicle emergence. Seeds imbibed in 100 μ M abscisic acid (ABA), particularly at 2°C, lost freezing tolerance at a slower rate compared with water-imbibed seeds. Seeds imbibed in water either at 23°C for 16 h, or 8°C for 6 days, or 2°C for 6 days were not germinable after storage at −6°C for 10 days. Seeds imbibed in ABA at 23°C for 24 h, or 8°C for 8 days, or 2°C for 15 days were highly germinable after 40 days at a constant −6°C. Desiccation injury induced at a high temperature (60°C), as with injury induced by freezing, was found to be a function of imbibition temperature and time.     

Patterns of desiccation tolerance and recovery in bryophytes

The study of desiccation tolerance in bryophytes avoids thecomplications of higher-plant vascular systems and complex leaf structures, butremains a multifaceted problem. Some of the pertinent questions have at leastpartial analogues in seed biology – events during a drying-rewettingcyclewith processes in seed maturation and germination, and the gradual loss ofviability on prolonged desiccation, and the relation of this to intensity ofdesiccation and temperature, with parallel questions in seed storage. Pastresearch on bryophyte desiccation tolerance is briefly reviewed. Evidence ispresented from chlorophyll-fluorescence measurements and experiments withmetabolic inhibitors that recovery of photosynthesis in bryophytes followingdesiccation depends mainly on rapid reactivation of pre-existing structures andinvolves only limited de novo protein synthesis. Followinginitial recovery, protein synthesis is demonstrably essential to themaintenanceof photosynthetic function in the light, but the rate of maintenance turnoverinthe dark appears to be slow. Factors leading to long-term desiccation damagearediverse; indications are that desiccation tolerant species often survive bestinthe range –100 to –200 MPa.     

ROS removal by DJ-1: Arabidopsis as a new model to understand Parkinson disease

Reactive oxygen species represent one of the principal factors that cause cell death and scavenging of reactive oxygen species by superoxide dismutase-related pathway is essential for cell survival. The Parkinson disease-related DJ-1 protein (also known as PARK7) has been implicated in resistance against oxidative stress in dopaminergic neurons however, its molecular mechanism has to date been unknown. We have used Arabidopsis thaliana as a model system to demonstrate that DJ-1, in both plant and mammalian cells, directly influence SOD activity in a highly conserved manner thereby preventing cell death. These data not only provides evidence for the molecular mechanisms associated with DJ-1-induced Parkinson disease but also highlight the unprecedented value of plants as a tool in understanding human disease mechanisms.Key words: DJ-1, stress, cell death, Parkinson disease, ArabidopsisReactive oxygen species (ROS) are involved in a myriad of fundamental biological processes including cell signaling and cellular defense pathways in plants and animals.^1–3 Despite its role as a signaling molecule, inappropriate and elevated levels of ROS have a major impact on the etiology of neurodegenerative diseases, such as for example Parkinson disease (PD), and in oxidative stress responses in plants. In general ROS can cause damage to DNA, lipids, proteins and various cofactors. During normal physiological conditions, when ROS are continuously generated, antioxidant defense systems are adequately equipped to prevent ROS-induced tissue dysfunction.^4,5 However upon elevated ROS generation the cellular antioxidant systems either recruit additional factors to minimize ROS-induced damage or cells suffer the consequences of cell death. Because of this dichotomy, where ROS plays a vital role during growth and development but can also have overwhelming damaging effects, it is clear that strict regulatory mechanisms need to be in place to effectively control ROS levels. In both plant and mammalian cells elevated ROS levels lead to cell death and in various human disease such as PD, Alzheimer disease, amyotrophic lateral sclerosis and Huntington disease proteins involved in stress-related pathways are often mutated.⁶In both plants and mammals mitochondria act as an important source of ROS however, plants also produce ROS in chloroplasts as part of photosynthetic activity. Combined with the fact that plants are sessile organisms it suggests that, although similar in nature, plants most probably have more complex antioxidant systems than other organisms.Strategies for removing excess ROS are similar in plants and humans. The principle ROS removal pathway involves superoxide dismutases (SOD) (or copper/zinc superoxide dismutase-CSD in plants), glutathione peroxidases (GPX) and catalases (CAT) localized in the cytosol, mitochondria and chloroplasts (Fig. 1). SOD converts superoxide anion to H₂O₂, which is then detoxified to H₂O by GPX and CAT. In Arabidopsis, besides SOD, GPX and CAT, there are five ascorbate peroxidases (APX) located in the cytosol and chloroplasts, involved in scavenging ROS generated during photosynthesis.^7,8 Therefore, SOD, GPX, CAT and APX, together with other auxiliary proteins, form the main line of defense against ROS.Open in a separate windowFigure 1Involvement of DJ-1-like proteins in ROS scavenging pathways. Produced O₂^- is converted to H₂O₂ by SOD in human or CSD (Arabidopsis SOD) in Arabidopsis. H₂O₂ is then converted to H₂O by GPX or CAT (catalase) in human or APX, GPX or CAT in Arabidopsis. DJ-1-like proteins interact with SOD or GPX in humans and CSD, GPX and APX in Arabidopsis. It is assumed here that DJ-1-like proteins may also interact with catalases (broken arrows).DJ-1 was originally identified as an oncogene and represents a ubiquitous redox-responsive cytoprotective protein with diverse functions where one of its main roles have been attributed to oxidative stress protection.⁹ Numerous studies have shown that several DJ-1 mutations in humans cause autosomal recessive, early onset PD however, its mode of action has been elusive in terms of having a direct influence on neuronal cell death.¹⁰ In an attempt to clarify the mechanism of DJ-1 we established Arabidopsis thaliana as a new and novel model system.¹¹ The Arabidopsis genome contains three DJ-1 homologs compared to the single DJ-1 locus found in humans and we showed that two of these (AtDJ-1b and AtDJ-1c) localizes to chloroplasts whilst one, AtDJ-1a, localizes to the cytosol and nucleus as observed for human DJ-1.¹¹ As mutated DJ-1 in mammals leads to cell death we identified and characterized a DJ-1 loss-of-function mutant which showed increased cell death in aging plants. Using Bimolecular Fluorescence Complementation (BiFC) and isothermal titration calorimetry (ITC) assays we showed that AtDJ-1a interacts with CSD1, the cytosolic SOD in Arabidopsis, and with human SOD1 in plant cells. Further we demonstrated that the human DJ-1 protein interacts with SOD1 in mammalian CHO cells.¹¹ Similar approaches were also employed to show that AtDJ-1a and human DJ-1 had an interaction with GPX2 in plant and mammalian cells.¹¹Enzyme assays revealed that AtDJ-1a and DJ-1 stimulated SOD/CSD1 activity and that only the copper-loaded forms of AtDJ-1a and DJ-1 had this effect suggesting that AtDJ-1a/DJ-1 may provide copper for SOD/CSD1.¹¹ Although the observed SOD activation provides clues towards the role of DJ-1 in detoxification of ROS, SOD only converts superoxide anion to H₂O₂ which must further be detoxified to H₂O by GPX and CAT. Although we showed that AtDJ-1a and human DJ-1 can interact with AtGPX2 and GPX2, respectively, we observed no changes in GPX2 activity upon DJ-1 interaction. The reason for this may be several-fold. First, cellular GPX2 activity levels may be sufficient to convert SOD-generated H₂O₂ to H₂O. Second, DJ-1 may indeed have no effect on GPX2 activity but simply act as an anchor to dock GPX2 in the vicinity of SOD. To test whether the DJ-1/SOD/GPX2 complex recruits other auxiliary proteins we have also shown that AtDJ-1a interacts with the Arabidopsis cytosolic APX1 protein (Fig. 2, unpublished data). It is also highly possible that DJ-1 interacts with catalase or at least influences its activity (Fig. 1). Although we have no data to date indicating a functional significance of the DJ-1/APX1 interaction we speculate that DJ-1 indeed acts as a scaffold protein bringing together SOD, GPX and possibly APX1 to mediate and control ROS scavenging, ultimately preventing oxidative stress-induced cell death (Fig. 3).Open in a separate windowFigure 2Interaction of AtDJ-1a with APX1. AtDJ-1a tagged with the N-terminal region of GFP and APX1 tagged with the C-terminal region of GFP gene were co-transformed into tobacco cells. The observed GFP signal in (B) demonstrates an AtDJ-1a/APX1 interaction through reconstitution of functional GFP molecules. (A) Negative control.Open in a separate windowFigure 3Working model of AtDJ-1a and DJ-1 mode of action. AtDJ-1a and DJ-1 interacts with SOD and GPX2 leading to SOD activation in a copper-dependent fashion. It is proposed that AtDJ-1a and DJ-1 delivers copper to SOD enhancing its activity whilst GPX2 is anchored by AtDJ-1 and DJ-1 to the protein complex to ensure conversion of the SOD-generated H₂O₂ to H₂O.The fact that Arabidopsis has three DJ-1 homologs where two of these, AtDJ-1b and AtDJ-1c, are localized to chloroplasts¹¹ underlines the protective role of DJ-1-like proteins during oxidative stress in plants. From our localization studies it appears that AtDJ-1b is localized to the chloroplast stroma whilst AtDJ-1c is localized to both the stroma and the thylakoid membranes (unpublished data). Whether AtDJ-1b and AtDJ-1c act in isolation or in concert and how these two proteins are involved in photosynthesis-induced ROS regulation is unclear but represent exciting future challenges.The notion that plants can be used as tools to increase our understanding of human disease mechanisms is somewhat obscure to the general scientific community. The fact remains that many discoveries with direct relevance to human health and disease have been elaborated using Arabidopsis, and several processes important to human biology are more easily studied in this versatile model plant.¹² The use of Arabidopsis to understand human disease states has several advantages: (1) Arabidopsis represents a well established model organism with a fully annotated genome, (2) The Arabidopsis genome contains homologs of numerous genes involved in human disease, (3) The identification and generation of Arabidopsis mutants is simple and requires little effort, (4) Arabidopsis growth and maintenance requires little infrastructure and running costs and (5) Arabidopsis research has few ethical constraints.Despite the advantages of Arabidopsis as a model system for elucidating human disease mechanisms it is important to appreciate that Arabidopsis and plant research in general can only reach its full potential in the field of medical research if combined with complementary, and perhaps more conventional, model systems.     

Xenopus leads the way: Frogs as a pioneering model to understand the human brain

From its long history in the field of embryology to its recent advances in genetics, Xenopus has been an indispensable model for understanding the human brain. Foundational studies that gave us our first insights into major embryonic patterning events serve as a crucial backdrop for newer avenues of investigation into organogenesis and organ function. The vast array of tools available in Xenopus laevis and Xenopus tropicalis allows interrogation of developmental phenomena at all levels, from the molecular to the behavioral, and the application of CRISPR technology has enabled the investigation of human disorder risk genes in a higher‐throughput manner. As the only major tetrapod model in which all developmental stages are easily manipulated and observed, frogs provide the unique opportunity to study organ development from the earliest stages. All of these features make Xenopus a premier model for studying the development of the brain, a notoriously complex process that demands an understanding of all stages from fertilization to organogenesis and beyond. Importantly, core processes of brain development are conserved between Xenopus and human, underlining the advantages of this model. This review begins by summarizing discoveries made in amphibians that form the cornerstones of vertebrate neurodevelopmental biology and goes on to discuss recent advances that have catapulted our understanding of brain development in Xenopus and in relation to human development and disease. As we engage in a new era of patient‐driven gene discovery, Xenopus offers exceptional potential to uncover conserved biology underlying human brain disorders and move towards rational drug design.     

Improving tolerance of somatic emrbyos of Picea glauca to flash desiccation with a cold treatment (desiccation after cold acclimation)

Summary Controlled mild desiccation of mature white spruce somatic embryos prior to germination improves the quality of the germinated embryos. More severe desiccation results in increased injury and death but is desirable for long-term storage of embryos and production of desiccated artificial seed. A method was developed to improve desiccation tolerance in somatic embryos using a temperature treatment. Culture plates with embryos at four stages of development were subjected to temperatures of 1, 5, 10, or 20°C for periods of 0, 1, 2, 4, or 8 wk duration. After the temperature treatment, the embryos were harvested and air-dried for 2h under a laminar flow hood. Dried embryos were placed directly on germination medium and the quality of the germinants was assessed after 4 wk. The initial maturation stage of the embryo and the temperature and duration of the treatment had a significant effect on the quality of the germinants. Most treatments caused marked differential survival of organs. The optimal response was obtained with embryos that had been grown for 51 d (cotyledonary stage) on maturation medium and that were subsequently exposed to a temperature of 5°C for 8 wk prior to air drying. This treatment produced 58% undamaged germinants with normal cotyledons, hypocotyls, and roots. Only 1% of the untreated air-dried embryos germinated normally.