Plant tolerance of flooding stress – recent advances

Soil waterlogging damages most crops, with the exception of rice, which like other wetland species thrives. Floods causing complete submergence can be catastrophic for crops, even for rice, but sustained interdisciplinary research efforts have resulted in breakthroughs in understanding submergence tolerance of rice and importantly the breeding of new submergence tolerant varieties (Bailey-Serres et al. 2010). Nevertheless, flooding is one of several abiotic stress factors impacting each year on food supplies and the economic situation of some communities. Regimes of excess water are a defining feature of several ecosystems and influence plant communities (e.g. floodplains, wetlands, riparian zones, tidal zones and salt marshes). Heavy rainfall, poor soil drainage, some irrigation practices and floods can impact negatively on crop production in many regions of the world. Plant species show an incredible diversity in tolerance to soil waterlogging, and also to shoot submergence; ‘dryland’ species (including many crops) are sensitive, wetland species (including rice) are tolerant and aquatic species even live submerged. Papers in this Special Issue address the important questions of how plants sense and acclimate to waterlogging/soil flooding (excess water in the root zone) or submergence when floods are of a depth that covers the shoots, and the adaptive mechanisms that confer tolerance. Excess water results in complex changes in several environmental parameters, caused by impeded gas exchange and subsequent responses of the soil microflora and micro-biome of the plant. Soil waterlogging leads to hypoxia and progressively to anoxia and high CO2 in the root zone, which is often accompanied by increased mobilization of ‘phytotoxins’ in reduced soils, all with consequences for root metabolism, nutrient acquisition and thus growth (and survival) of roots and shoots. As a lack of O2 inhibits respiration, metabolic plant adaptations to cope with the hypoxic and anoxic conditions and resulting energy deficits, as well as anatomical and morphological adaptations to improve internal O2 supply, have been topics of flooding stress research. Insight into these central themes, as well as improved understanding of plant responses to the multiple stress factors comprising flooding stress, has seen recent advances as highlighted by the contributions in this Special Issue. Studies reported over 75 years ago first gleaned that cells of plant tissues (seeds, roots and aerial organs) surrounded by stagnant water undergo a shift from carbon-efficient aerobic respiration to carbon-inefficient anaerobic metabolism to produce sufficient ATP to maintain viability (reviewed by Turner 1951; Drew 1997; Bailey-Serres & Voesenek 2008). The rate of anaerobic metabolism promoted by low O2 or flooding stress can vary considerably, with some cells vigorously increasing flux through glycolysis (Pasteur effect) and others with more conservative carbohydrate consumption. A hallmark of the onset of anaerobic metabolism in plant cells is increased transcription of genes and production of enzymes for ethanolic fermentation, namely pyruvate decarboxylase (PDC) and alcohol dehydrogenase (ADH), as reported in the first molecular characterization of the response to low O2 in a plant by Sachs et al. (1980). This can be accompanied with enhanced catabolism of carbohydrates such as starch to fuel fermentative metabolism and in some highly tolerant species even growth. Several papers in this Special Issue consider the reconfiguration of gene expression and metabolism that occur in response to low O2 and various flooding regimes of model and crop plants, including tree species (Kreuzwieser & Rennenberg 2014; Lee et al. 2014; Mustroph et al. 2014; Santaniello et al. 2014; Shingaki-Wells et al. 2014). These highlight a wealth of insight into the complex ramifications of low O2 stress at the cell, organ and whole-plant levels. Mustroph et al. (2014), for example, perform a detailed analysis of the cell-type specific expression of genes encoding enzymes associated with central carbon and nitrogen metabolism, which was paired with metabolite profiles of the roots and shoots of Arabidopsis thaliana seedlings. The study illustrates the invariably pronounced induction of genes encoding the enzymes sucrose synthase (SUS), PDC, ADH, alanine aminotransferase and aspartate aminotransferase. The study shows that increased levels of the mRNA transcripts of genes encoding these enzymes are more pronounced in roots in response to hypoxia, submergence or waterlogging. Variations in transcript induction were evident at the cell- and organ-specific levels, with nitrate reductase preferentially up in roots and gamma-aminobutyric acid (GABA) transaminase markedly up in shoots of submerged and waterlogged plants. These and other organ-specific dynamics in gene transcripts and metabolites were recognized and discussed. Moreover, the hypoxia- and light-dependent up-regulation of genes associated with galactolipid or sulfolipid metabolism in shoots was more fully examined by the Mustroph group (Klecker et al. 2014). Nicely juxtaposed to the Arabidopsis study is an insightful ‘omic’ survey of responses of trees to waterlogging (Kreuzwieser & Rennenberg 2014). This paper emphasizes the underappreciated integrated molecular and physiological responses of the whole plant. For example, root waterlogging has ramifications on hydraulic conductivity and metabolite transport. By comparison of trees with different flooding sensitivity, the importance of root-to-shoot transport of metabolites to ‘use rather than lose’ precious carbon emerges as a strategy of the more waterlogging tolerant species. Arabidopsis provides the opportunity to exploit gene mutant and protein overproduction lines to manipulate responses to low O2 stress. This approach was taken by Santaniello et al. (2014) to better understand the importance of flux via SUS versus cytosolic invertase (INV) in sucrose catabolism and ethanol production during hypoxia. Despite the consensus that flux via SUS is energetically advantageous over that of ATP-consuming INV/hexokinase route, the use of genotypes compromised in SUS or INV activity revealed that flux through SUS was beneficial for survival of waterlogging or submergence of older plants but not of O2-deprived seedlings grown on carbon-containing medium. These researchers suggest that the pronounced up-regulation of SUS genes observed across plant species in response to low O2 conditions may be less important in anaerobic metabolism than previously thought. Mutants of transcription factor and metabolic enzyme genes as well as application of the hormones ethylene, gibberellin and abscisic acid (ABA) have been used to unravel the signalling and response pathways that regulate carbon catabolism needed for germination of seeds in stagnant water and elongation under water of shoot tissues in rice (Oryza sativa). Studies along these lines are refreshingly presented in two reviews. The first by Lee et al. (2014) highlights the conserved SNF1/SnRK1/AMPK1 kinase family that regulates energy metabolism across eukaryotes and is an important regulator of starch catabolism and ADH expression during seed germination under O2 deficiency in rice. There is also a research report on the Arabidopsis SnRK1 called KIN10 and its interplay with ABA-regulated transcription and salt stress, which raises questions about crosstalk between saline and submergence stress responses (Im et al. 2014). In the second review, Shingaki-Wells et al. (2014) evaluate more than 125 papers in a discussion of the constraints of O2 deprivation and reoxygenation on mitochondrial metabolism. The topics addressed range from the reversible change in mitochondrial ultrastructure reported by Vartapetian in the 1970s to the recent description of the O2-sensing turnover of ethylene responsive transcription factors (ERF-VIIs) that regulate PDC, ADH and other anaerobic response genes (Gibbs et al. 2011; Licausi et al. 2011). Questions are raised about the source, dynamics and roles of reactive nitrogen species (i.e. nitric oxide) and reactive oxygen species (ROS, i.e. superoxide and hydrogen peroxide) in response to severe O2 deprivation and subsequent reoxygenation. The topic of reoxygenation is also considered in an integrative physiological and genome-scale ‘omic’ analysis of the response to submergence of soybean (Glycine max) seedlings (Tamang et al. 2014). This nicely executed study can be easily compared with knowledge from rice and Arabidopsis and will serve as a foundation for surveys of genetic determinants of submergence tolerance in soybean germplasm. Soybeans, like other plants, induce ERF-VIIs in response to submergence as well as ethylene, a key regulator of diverse responses to flooding. The Tsai et al. (2014) study on Arabidopsis in this Special Issue demonstrates that ethylene and probably also jasmonic acid are important in the return to homeostasis following O2 deprivation. Waterlogging has adverse impacts on productivity of crops (e.g. wheat and barley; Setter & Waters 2003) and forages (e.g. Bennett et al. 2009). Striker et al. (2014) evaluated growth responses and various traits of a Lotus japonicus recombinant inbred line population subjected to 21 d of soil waterlogging. The diversity in growth and physiological responses in waterlogged conditions enabled identification of quantitative trait loci (QTL) for various characters. Root aerenchyma correlated with root dry mass in waterlogged conditions, but somewhat surprisingly it was not correlated with shoot dry mass, which was instead related to higher stomatal conductance and Fv/Fm under waterlogging. The authors emphasized that further work is needed to understand the contributions of root- and shoot-related traits to waterlogging tolerance, and that the responses of these organs might be more independent than previously recognized. Aerenchyma provides a low-resistance pathway for diffusion of O2 from the shoot base to the root tip, enabling growth into anoxic soils (Armstrong 1979; Justin & Armstrong 1987). Roots of many wetland species contain aerenchyma and also a barrier against radial O2 loss (ROL); the ROL barrier further enhances O2 movement to the apex and impedes entry of soil toxins (Armstrong 1979; Armstrong & Armstrong 2005). The ROL barrier is induced by growth conditions in roots of some species, including rice, for which the molecular regulation (Shiono et al. 2014) and biochemical processes (Kulichikhin et al. 2014) are only now being elucidated. Use of laser microdissection to sample the outer cell layers of rice roots exposed to stagnant (ROL barrier-inducing conditions) versus aerated conditions, and metabolite-profiling and transcriptome analyses, has highlighted the increased abundances of transcripts encoding enzymes in suberin synthesis and associated metabolic precursors (Kulichikhin et al. 2014). This progress in understanding ROL barrier induction is further revealing the sophisticated adaptation of roots of rice to soil waterlogging and since some wild relatives of dryland crops also possess this trait (e.g. Malik et al. 2011; Abiko et al. 2012) could help in future design strategies aimed at improvement of root aeration (and potentially phytotoxin exclusion) in some dryland crops. Waterlogged soils are hostile to non-adapted species as not only is O2 lacking (anoxia) but the activity of anaerobic microorganisms results in elevated levels of phytotoxins such as Mn2+, Fe2+, H2S and various short-chain fatty acids (e.g. acetic acid, butyric acid, propionic acid), depending on soil type and conditions. Phytotoxins can impede root growth or even result in death, but less appreciated has been that sub-lethal levels can also directly influence nutrient acquisition via effects on the activity of various membrane transporters. Shabala et al. (2014) review the impacts of various phytotoxins on root cell plasma membrane transporters, as well as consider the effects of low O2, and argue that the rapid responses of transporters to these soil conditions can initiate signalling cascades (including those involving ROS) and likely determine responses in roots, with consequences also for shoots. Further comparisons of root membrane transporters of wetland species compared with waterlogging-sensitive crops are needed. Shabala et al. (2014) conclude that breeding strategies aimed to improve waterlogging tolerance in crops will require pyramiding of numerous traits, including the role of membrane transporters in control of cytosolic ion homeostasis (especially K+) and resistance of roots to soil phytotoxins. Experiments by Zeng et al. (2014) further evaluated K+ homeostasis in hypoxic and anoxic barley roots, using the high spatial and temporal resolution afforded by micro-electrodes. These authors recorded membrane potential and net K+ flux responses to O2 availability of root cells within the elongation and mature zones of two varieties of contrasting waterlogging tolerance. The extent of membrane depolarisation was less pronounced in the roots of the tolerant variety and prevention of internal O2 movement diminished H+-pumping activity. This was associated with more pronounced membrane depolarisation, large net K+ efflux and a rapid loss of cell viability. Tissue-specific responses were evident and likely result from differences in O2 available along and within roots, owing to diffusion gradients and/or differences between tissues in anoxia sensitivity. The authors discuss the role of membrane processes in altered K+ status and the possible link to programmed cell death in severely hypoxic and anoxic roots. This study and that of Kulichikhin et al. (2014) emphasize the significance of localized responses to O2 deficiency on metabolism in roots. When shoots are completely submerged, O2 supply for respiration and CO2 uptake for photosynthesis are reduced because diffusion is 10 000 slower in water than in air. Plant adaptation to complete submergence can involve contrasting strategies depending on the depth and duration of flooding: either a shoot elongation response to regain contact with air or a lack of elongation and thus conservation of carbohydrates to outlast the submergence period (cf. Bailey-Serres & Voesenek 2008). Deepwater rice grows in areas with prolonged floods and maintains a portion of its shoot above water – this provides a ‘snorkel’ and is achieved via elongation of internodes, triggered by ethylene that acts on two SNORKEL genes, also ERF-VII transcriptional regulators (Hattori et al. 2009). By contrast, for lowland rice completely submerged during transient floods (a few days up to 3 weeks), shoot elongation can be detrimental so that tolerant genotypes suppress shoot elongation and then show better recovery growth after the water recedes (Mackill et al. 2012). This response is regulated by SUB1A-1, an ERF-VII transcriptional regulator that reduces gibberellin responsiveness and diminishes shoot elongation and enhances other aspects of the flooding stress response (Xu et al. 2006; Fukao et al. 2006; Fukao & Bailey-Serres 2008). The shoot elongation growth response of deepwater rice to escape submergence was further studied by Ayano et al. (2014), and they found that the elongation capacity depends on developmental stage; plants must first reach the six-leaf stage. Elongation at this stage occurred for internodes 6 and 7 and was dependant on increased gibberellins (i.e. GA1 and GA4) in these internodes. Other wetland species also show a shoot elongation response during submergence, such as the wild wetland species Rumex palustris which has been studied in detail by Voesenek & co-workers (summarized in Bailey-Serres & Voesenek 2008) and further elucidated at the level of gene regulation by use of mRNA sequencing (van Veen et al. 2013). Herzog & Pedersen (2014) evaluated the benefit to internal aeration of restoration of atmospheric contact by leaf tips of the ‘flood escaper’ R. palustris and flood-intolerant R. acetosa, which is without a shoot elongation response. Leaf emergence into air above the water provided substantial enhancement of internal O2 supply in R. palustris as the petioles elongate and contain aerenchyma. By contrast, R. acetosa with petioles of low porosity (and which do not elongate) did not benefit when leaf tips were artificially brought above the water level. Moreover, experiments conducted in darkness or with light available to the submerged shoots highlighted the importance of underwater photosynthesis to internal O2 status of submerged plants (Herzog & Pedersen 2014), which has also recently been demonstrated in field conditions for submerged rice (Winkel et al. 2013). Complementing this is the van Veen et al. (2014) systematic comparison of these two Rumex species (R. acetosa and R. palustris) as well as two Rorippa species (Ro. sylvestris and Ro. amphibia) with their opposing survival strategies. Consistent with the soybean study (Tamang et al. 2014) and earlier rice and Arabidopsis work, the ERF-VIIs of these species are identified and monitored. In a tour de force, the authors compare gene, metabolite, growth and survival characteristics of the four species considering root and shoot tissue as well as the leaf petiole region, that is, what drives submergence escape in these dicots. Photosynthesis by leaves of submerged plants depends on environmental conditions (CO2, light, turbulence) and leaf traits (Colmer et al. 2011). Leaves of some plants, including rice and several other wetland species, are super-hydrophobic and retain a gas film when submerged. Leaf gas films enhance underwater gas exchange between the tissues and floodwater. Apparent resistance to CO2 entry during light periods was ∼5 times lower when gas films were present on submerged rice leaves (Pedersen et al. 2009) and O2 entry during darkness was also enhanced by gas films (Pedersen et al. 2009; Winkel et al. 2013). Using a mathematical modelling approach, Verboven et al. (2014) explored the mechanism by which surface gas films can enhance O2 movement into leaves when submerged. The modelling, which used the scenario of a respiring leaf in darkness to understand O2 entry, supported the conclusion that gas films significantly reduce the resistance to O2 entry into submerged leaves when stomata are at least partly open; O2 enters from the floodwater and can move rapidly within the gas film to stomata. Gas films were also the subject of experiments of Teakle et al. (2014) on Melilotus siculus, an annual pasture legume, when in mildly saline submergence water. These envelopes of gas protected against ion (Na+ and Cl−) entry into the leaves in saline water, with enhanced tissue health (e.g. K+ retention) and augmented of underwater photosynthesis after 3 d of submergence in floodwater The authors that the gas provided a between the tissue and saline water which in reduced and entry into the submerged gas films to be beneficial to plant tolerance of submergence when in or saline is a severe abiotic most plants are to soil waterlogging and even more are of complete stress research has from use of rice as a a crop species with flooding tolerance and Other and use of wild wetland species have also knowledge of responses to flooding and O2 including the key role of ethylene in these interdisciplinary research has resulted in breeding of submergence tolerant varieties (Bailey-Serres et al. but knowledge and for further improvement of rice for various ecosystems and improvement of other crop and pasture species (e.g. and Soil waterlogging leads to high CO2 and ethylene, and phytotoxins in reduced soils, with impacts also when shoots are also important is the to from with water to a stagnant waterlogged or even is the and effects of these of the caused by flooding. The in roles of ethylene and the SnRK1 signalling as well as further of wild species, is likely to As flooding can with other stress factors (e.g. or the of plant responses to of stress is also are likely to to the most rapid in knowledge of plant stress tolerance. Several papers in this Special Issue were on topics of presented at the of the for Plant at the the and the at for their during the were from various research independent of at the all for their and on the to that constraints the of papers that could be in this Special Issue.