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The development of male gametes, vital to sexual reproduction in crops, requires meiosis followed by successive mitotic cell divisions of haploid cells. The formation of viable pollen is especially vulnerable to abiotic stress, with consequences both for yield and for grain quality. An understanding of key molecular responses when specific stages during pollen development are subjected to stress (e.g. heat) is possible only when sampling is carefully informed by developmental biology. Traditionally, morphological characteristics have been commonly used in cereals as 'indicators' of male reproductive stages. We argue that these morphological attributes are strongly influenced by genotype and genotype-environment interactions and cannot be used reliably to define developmental events during microsporogenesis and microgametogenesis. Furthermore, asynchronous development along the axis of a single inflorescence calls for selective sampling of individual florets to define specific reproductive stages accurately. We therefore propose guidelines to standardise the sampling of cells during male reproductive development, particularly when interrogating the impact of stress on susceptible meiosis. Improved knowledge of development will largely negate the variability imposed by genotype, environment and asynchronous development of florets. Highlighting the subtleties required for sampling and investigation of male reproductive stages will make the selection of abiotic stress-tolerant cereal genotypes more reliable., (© 2024 The Author(s). New Phytologist © 2024 New Phytologist Foundation.)

Background and Aims: Five species of cotton (Gossypium) were exposed to 38°C days during early vegetative development. Commercial cotton (Gossypium hirsutum) was contrasted with four wild cotton species (G. australe, G. bickii, G. robinsonii and G. sturtianum) that are endemic to central and northern Australia., Methods: Plants were grown at daytime maxima of 30°C or 38°C for 25 d, commencing at the four-leaf stage. Leaf areas and shoot biomass were used to calculate relative rates of growth and specific leaf areas. Leaf gas exchange measurements revealed assimilation and transpiration rates, as well as electron transport rates (ETR) and carboxylation efficiency (CE) in steady-state conditions. Finally, leaf morphological traits (mean leaf area and leaf shape were quantified), along with leaf surface decorations, imaged using scanning electron microscopy., Key Results: Shoot morphology was differentially affected by heat, with three of the four wild species growing faster at 38°C than at 30°C, whereas early growth in G. hirsutum was severely inhibited by heat. Areas of individual leaves and leaf numbers both contributed to these contrasting growth responses, with fewer, smaller leaves at 38°C in G. hirsutum. CO2 assimilation and transpiration rates of G. hirsutum were also dramatically reduced by heat. Cultivated cotton failed to achieve evaporative cooling, contrasting with the transpiration-driven cooling in the wild species. Heat substantially reduced ETR and CE in G. hirsutum, with much smaller effects in the wild species. We speculate that leaf shape, as assessed by invaginations of leaf margins, and leaf size contributed to heat dispersal differentially among the five species. Similarly, reflectance of light radiation was also highly distinctive for each species., Conclusions: These four wild Australian relatives of cotton have adapted to hot days that are inhibitory to commercial cotton, deploying a range of physiological and structural adaptations to achieve accelerated growth at 38°C., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Annals of Botany Company.)

Adaptations of plants to phosphorus (P) deficiency include reduced investment of leaf P in storage (orthophosphates in vacuoles), nucleic acids and membrane lipids. Yet, it is unclear how these adaptations are associated with plant ecological strategies. Five leaf P fractions (orthophosphate P, P i ; metabolite P, P M ; nucleic acid P, P N ; lipid P, P L ; and residual P, P R ) were analysed alongside leaf economic traits among 35 Australian woody species from three habitats: one a high-P basalt-derived soil and two low-P sandstone-derived soils, one undisturbed and one disturbed by human activities with artificial P inputs. Species at the undisturbed low-P site generally exhibited lower concentrations of total leaf P ([P total ]), primarily associated with lower concentrations of P i , and P N . The relative allocation of P to each fraction varied little among sites, except that higher P L per [P total ] (rP L ) was recorded at the undisturbed low-P site than at the high-P site. This higher rP L , reflecting relative allocation to membranes, was primarily associated with lower concentrations of leaf nitrogen at the undisturbed low-P site than at the high-P site. Associations between leaf P fractions and leaf nitrogen may provide a basis for understanding the variation in plant ecological strategies dependent on soil P availability., (© 2024 The Authors. New Phytologist © 2024 New Phytologist Foundation.)

Next-generation sequencing demands high-quality nucleic acid, yet isolating DNA and RNA is often challenging, particularly from plant tissues. Despite advances in developing various kits and reagents, these products are tailored to isolation of nucleic acid from model plant tissues. Here we introduce a universal lysis buffer to separate nucleic acid from various plant species, including recalcitrant plants, to facilitate molecular analyses, such as quantitative PCR (qPCR), transcriptomics, and whole-genome sequencing (WGS). The protocol is a modification of the original CTAB methods, which leads to nucleic acid isolation from many plant species, including monocots and eudicots. The lysis buffer consists of hexadecyltrimethylammonium bromide (CTAB), sodium chloride (NaCl), Tris base, ethylenediaminetetraacetic acid (EDTA) and β-mercaptoethanol (βME). The modified CTAB method enables the isolation of nucleic acid from small amounts of plant tissues (e.g., 15-100 mg) in a timely manner, which is well-suited for a large number of samples and also when adequate sample collection is a limiting factor. The protocol isolates not only DNA from various plant species but also RNA. This makes it highly effective for molecular analyses compared to previously described CTAB methods optimised for DNA isolation. The appropriate concentration of the components enables high-quality DNA and RNA isolation from plant tissues simultaneously. Additionally, this protocol is compatible with commercially available columns. For DNA and RNA to be qualified for next-generation sequencing platforms, the protocol is supplemented with columns to purify either DNA or RNA from the same tissue to meet high standards for sequencing analyses. This protocol provides an ideal approach to overcome potential obstacles in isolating high-quality DNA or RNA from a wide range of plant species for downstream molecular analysis., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Masoomi-Aladizgeh et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Multiple abiotic stress is known as a type of environmental unfavourable condition maximizing the yield and growth gap of crops compared with the optimal condition in both natural and cultivated environments. Rice is the world's most important staple food, and its production is limited the most by environmental unfavourable conditions. In this study, we investigated the pre-treatment of abscisic acid (ABA) on the tolerance of the IAC1131 rice genotype to multiple abiotic stress after a 4-day exposure to combined drought, salt and extreme temperature treatments. A total of 3285 proteins were identified and quantified across the four treatment groups, consisting of control and stressed plants with and without pre-treatment with ABA, with 1633 of those proteins found to be differentially abundant between groups. Compared with the control condition, pre-treatment with the ABA hormone significantly mitigated the leaf damage against combined abiotic stress at the proteome level. Furthermore, the application of exogenous ABA did not affect the proteome profile of the control plants remarkably, while the results were different in stress-exposed plants by a greater number of proteins changed in abundance, especially those which were increased. Taken together, these results suggest that exogenous ABA has a potential priming effect for enhancing the rice seedlings' tolerance against combined abiotic stress, mainly by affecting stress-responsive mechanisms dependent on ABA signalling pathways in plants.

Food safety of staple crops such as rice is of global concern and is at the top of the policy agenda worldwide. Abiotic stresses are one of the main limitations to optimizing yields for sustainability, food security and food safety. We analyzed proteome changes in Oryza sativa cv. Nipponbare in response to five adverse abiotic treatments, including three levels of drought (mild, moderate, and severe), soil salinization, and non-optimal temperatures. All treatments had modest, negative effects on plant growth, enabling us to identify proteins that were common to all stresses, or unique to one. More than 75% of the total of differentially abundant proteins in response to abiotic stresses were specific to individual stresses, while fewer than 5% of stress-induced proteins were shared across all abiotic constraints. Stress-specific and non-specific stress-responsive proteins identified were categorized in terms of core biological processes, molecular functions, and cellular localization.

Drought is responsible for major losses in rice production. Root tips contain meristematic and elongation zones that play major roles in determination of root traits and adaptive strategies to drought. In this study we analysed two contrasting genotypes of rice: IR64, a lowland, drought-susceptible, and shallow-rooting genotype; and Azucena, an upland, drought-tolerant, and deep-rooting genotype. Samples were collected of root tips of plants grown under control and water deficit stress conditions. Quantitative proteomics analysis resulted in the identification of 7294 proteins from the root tips of IR64 and 6307 proteins from Azucena. Data are available via ProteomeXchange with identifier PXD033343. Using a Partial Least Square Discriminant Analysis on 4170 differentially abundant proteins, 1138 statistically significant proteins across genotypes and conditions were detected. Twenty two enriched biological processes showing contrasting patterns between two genotypes in response to stress were detected through gene ontology enrichment analysis. This included identification of novel proteins involved in root elongation with specific expression patterns in Azucena, including four Expansins and seven Class III Peroxidases. We also detected an antioxidant network and a metallo-sulfur cluster assembly machinery in Azucena, with roles in reactive oxygen species and iron homeostasis, and positive effects on root cell cycle, growth and elongation., (© 2022 The Authors. Proteomics published by Wiley-VCH GmbH.)

Key Message: A wild relative of rice from the Australian savannah was compared with cultivated rice, revealing thermotolerance in growth and photosynthetic processes and a more robust carbon economy in extreme heat. Above ~ 32 °C, impaired photosynthesis compromises the productivity of rice. We compared leaf tissues from heat-tolerant wild rice (Oryza australiensis) with temperate-adapted O. sativa after sustained exposure to heat, as well as diurnal heat shock. Leaf elongation and shoot biomass in O. australiensis were unimpaired at 45 °C, and soluble sugar concentrations trebled during 10 h of a 45 °C shock treatment. By contrast, 45 °C slowed growth strongly in O. sativa. Chloroplastic CO 2 concentrations eliminated CO 2 supply to chloroplasts as the basis of differential heat tolerance. This directed our attention to carboxylation and the abundance of the heat-sensitive chaperone Rubisco activase (Rca) in each species. Surprisingly, O. australiensis leaves at 45 °C had 50% less Rca per unit Rubisco, even though CO 2 assimilation was faster than at 30 °C. By contrast, Rca per unit Rubisco doubled in O. sativa at 45 °C while CO 2 assimilation was slower, reflecting its inferior Rca thermostability. Plants grown at 45 °C were simultaneously exposed to 700 ppm CO 2 to enhance the CO 2 supply to Rubisco. Growth at 45 °C responded to CO 2 enrichment in O. australiensis but not O. sativa, reflecting more robust carboxylation capacity and thermal tolerance in the wild rice relative., (© 2022. Crown.)

Oryza australiensis is a wild rice native to monsoonal northern Australia. The International Oryza Map Alignment Project emphasises its significance as the sole representative of the EE genome clade. Assembly of the O. australiensis genome has previously been challenging due to its high Long Terminal Repeat (LTR) retrotransposon (RT) content. Oxford Nanopore long reads were combined with Illumina short reads to generate a high-quality ~ 858 Mbp genome assembly within 850 contigs with 46× long read coverage. Reference-guided scaffolding increased genome contiguity, placing 88.2% of contigs into 12 pseudomolecules. After alignment to the Oryza sativa cv. Nipponbare genome, we observed several structural variations. PacBio Iso-Seq data were generated for five distinct tissues to improve the functional annotation of 34,587 protein-coding genes and 42,329 transcripts. We also report SNV numbers for three additional O. australiensis genotypes based on Illumina re-sequencing. Although genetic similarity reflected geographical separation, the density of SNVs also correlated with our previous report on variations in salinity tolerance. This genome re-confirms the genetic remoteness of the O. australiensis lineage within the O. officinalis genome complex. Assembly of a high-quality genome for O. australiensis provides an important resource for the discovery of critical genes involved in development and stress tolerance., (© 2022. Crown.)

Heat stress specifically affects fertility by impairing pollen viability but cotton wild relatives successfully reproduce in hot savannas where they evolved. An Australian arid-zone cotton (Gossypium robinsonii) was exposed to heat events during pollen development then mature pollen was subjected to deep proteomic analysis using 57 023 predicted genes from a genomic database we assembled for the same species. Three stages of pollen development, including tetrads (TEs), uninucleate microspores (UNs) and binucleate microspores (BNs) were exposed to 36°C or 40°C for 5 days and the resulting mature pollen was collected at anthesis (p-TE, p-UN and p-BN, respectively). Using the sequential windowed acquisition of all theoretical mass spectra proteomic analysis, 2704 proteins were identified and quantified across all pollen samples analysed. Proteins predominantly decreased in abundance at all stages in response to heat, particularly after exposure of TEs to 40°C. Functional enrichment analyses demonstrated that extreme heat increased the abundance of proteins that contributed to increased messenger RNA splicing via spliceosome, initiation of cytoplasmic translation and protein refolding in p-TE40. However, other functional categories that contributed to intercellular transport were inhibited in p-TE40, linked potentially to Rab proteins. We ascribe the resilience of reproductive processes in G. robinsonii at temperatures up to 40°C, relative to commercial cotton, to a targeted reduction in protein transport., (© 2022 John Wiley & Sons Ltd.)

Rice crops are often subject to multiple abiotic stresses simultaneously in both natural and cultivated environments, resulting in yield reductions beyond those expected from single stress. We report physiological changes after a 4 day exposure to combined drought, salt and extreme temperature treatments, following a 2 day salinity pre-treatment in two rice genotypes-Nipponbare (a paddy rice) and IAC1131 (an upland landrace). Stomata closed after two days of combined stresses, causing intercellular CO2 concentrations and assimilation rates to diminish rapidly. Abscisic acid (ABA) levels increased at least five-fold but did not differ significantly between the genotypes. Tandem Mass Tag isotopic labelling quantitative proteomics revealed 6215 reproducibly identified proteins in mature leaves across the two genotypes and three time points (0, 2 and 4 days of stress). Of these, 987 were differentially expressed due to stress ( cf. control plants), including 41 proteins that changed significantly in abundance in all stressed plants. Heat shock proteins, late embryogenesis abundant proteins and photosynthesis-related proteins were consistently responsive to stress in both Nipponbare and IAC1131. Remarkably, even after 2 days of stress there were almost six times fewer proteins differentially expressed in IAC1131 than Nipponbare. This contrast in the translational response to multiple stresses is consistent with the known tolerance of IAC1131 to dryland conditions.

Reproductive performance in plants is impaired as maximum temperatures consistently approach 40°C. However, the timing of heatwaves critically affects their impact. We studied the molecular responses during pollen maturation in cotton to investigate the vulnerability to high temperature. Tetrads (TEs), uninucleate and binucleate microspores, and mature pollen were subjected to SWATH-MS and RNA-seq analyses after exposure to 38/28°C (day/night) for 5 days. The results indicated that molecular signatures were downregulated progressively in response to heat during pollen development. This was even more evident in leaves, where three-quarters of differentially changed proteins decreased in abundance during heat. Functional analysis showed that translation of genes increased in TEs after exposure to heat; however, the reverse pattern was observed in mature pollen and leaves. For example, proteins involved in transport were highly abundant in TEs whereas in later stages of pollen formation and leaves, heat suppressed synthesis of proteins involved in cell-to-cell communication. Moreover, a large number of heat shock proteins were identified in heat-affected TEs, but these proteins were less abundant in mature pollen and leaves. We speculate that the sensitivity of TE cells to heat is related to high rates of translation targeted to pathways that might not be essential for thermotolerance. Molecular signatures during stages of pollen development after heatwaves could provide markers for future genetic improvement., (© 2021 Society for Experimental Biology and John Wiley & Sons Ltd.)

Mesophyll conductance (gm) is a crucial leaf trait contributing to the photosynthetic rate (AN). Plant domestication typically leads to an enhancement of AN that is often associated with profound anatomical modifications, but it is unclear which of these structural alterations influence gm. We analyzed the implication of domestication on leaf anatomy and its effect on gm in 26 wild and 31 domesticated cotton genotypes (Gossypium sp.) grown under field conditions. We found that domesticated genotypes had higher AN but similar gm to wild genotypes. Consistent with this, domestication did not translate into significant differences in the fraction of mesophyll occupied by intercellular air spaces (fias) or mesophyll and chloroplast surface area exposed to intercellular air space (Sm/S and Sc/S, respectively). However, leaves of domesticated genotypes were significantly thicker, with larger but fewer mesophyll cells with thinner cell walls. Moreover, domesticated genotypes had higher cell wall conductance (gcw) but smaller cytoplasmic conductance (gcyt) than wild genotypes. It appears that domestication in cotton has not generally led to significant improvement in gm, in part because their thinner mesophyll cell walls (increasing gcw) compensate for their lower gcyt, itself due to larger distance between plasmalemma and chloroplast envelopes., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

The scarcity of freshwater is an increasing concern in flood-irrigated rice, whilst excessive use of nitrogen fertilizers is costly and contributes to environmental pollution. To co-ordinate growth adaptation under prolonged exposure to limited water or excess nitrogen supply, plants employ complex systems for signalling and regulation of metabolic processes. There is limited information on the involvement of one of the most important post-translational modifications (PTMs), protein phosphorylation, in plant adaptation to long-term changes in resource supply. Oryza sativa cv. Nipponbare was grown under two regimes of nitrogen from the time of germination to final harvest. Twenty-five days after germination, water was withheld from half the pots in each nitrogen treatment and low water supply continued for an additional 26 days, while the remaining pots were well watered. Leaves from all four groups of plants were harvested after 51 days in order to test whether phosphorylation of leaf proteins responded to prior abiotic stress events. The dominant impact of these resources is exerted in leaves, where PTMs have been predicted to occur. Proteins were extracted and phosphopeptides were analysed by nanoLC-MS/MS analysis, coupled with label-free quantitation. Water and nitrogen regimes triggered extensive changes in phosphorylation of proteins involved in membrane transport, such as the aquaporin OsPIP2-6, a water channel protein. Our study reveals phosphorylation of several peptides belonging to proteins involved in RNA-processing and carbohydrate metabolism, suggesting that phosphorylation events regulate the signalling cascades that are required to optimize plant response to resource supply.

The development of gametes in plants is acutely susceptible to heatwaves as brief as a few days, adversely affecting pollen maturation and reproductive success. Pollen in cotton (Gossypium hirsutum) was differentially affected when tetrad and binucleate stages were exposed to heat, revealing new insights into the interaction between heat and pollen development. Squares were tagged and exposed to 36/25°C (day/night, moderate heat) or 40/30°C (day/night, extreme heat) for 5 days. Mature pollen grains and leaves were collected for physiological and proteomic responses. While photosynthetic competence was not compromised even at 40°C, leaf tissues became leakier. In contrast, pollen grains were markedly smaller after the tetrad stage was exposed to 40°C and boll production was reduced by 65%. Sugar levels in pollen grains were elevated after exposure to heat, eliminating carbohydrate deficits as a likely cause of poor reproductive capacity. Proteomic analysis of pure pollen samples revealed a particularly high abundance of 70-kDa heat shock (Hsp70s) and cytoskeletal proteins. While short-term bursts of heat had a minor impact on leaves, male gametophyte development was profoundly damaged. Cotton acclimates to maxima of 36°C at both the vegetative and reproductive stages but 5-days exposure to 40°C significantly impairs reproductive development., (© 2020 John Wiley & Sons Ltd.)

Elevated atmospheric CO 2 (eC a ) may benefit plants during drought by reducing stomatal conductance (g s ) but any 'water savings effect' could be neutralized by concurrent stimulation of leaf area. We investigated whether eC a enhanced water savings, thereby ameliorating the impact of drought on carbon and water relations in trees. We report leaf-level gas exchange and whole-plant and soil water relations during a short-term dry-down in two Eucalyptus species with contrasting drought tolerance. Plants had previously been established for 9 to 11 months in steady-state conditions of ambient atmospheric CO 2 (aC a ) and eC a , with half of each treatment group exposed to sustained drought for 5 to 7 months. The lower stomatal conductance under eC a did not lead to soil moisture savings during the dry-down due to the counteractive effect of increased whole-plant leaf area. Nonetheless, eC a -grown plants maintained higher photosynthetic rates and leaf water potentials, making them less stressed during the dry-down, despite being larger. These effects were more pronounced in the xeric species than the mesic species, and in previously water-stressed plants. Our findings indicate that eC a may enhance plant performance during drought despite a lack of soil water savings, especially in species with more conservative growth and water-use strategies., (© 2021 The Authors New Phytologist © 2021 New Phytologist Foundation.)

Drought often compromises yield in non-irrigated crops such as rainfed rice, imperiling the communities that depend upon it as a primary food source. In this study, two cultivated species ( Oryza sativa cv. Nipponbare and Oryza glaberrima cv. CG14) and an endemic, perennial Australian wild species ( Oryza australiensis ) were grown in soil at 40% field capacity for 7 d (drought). The hypothesis was that the natural tolerance of O. australiensis to erratic water supply would be reflected in a unique proteomic profile. Leaves from droughted plants and well-watered controls were harvested for label-free quantitative shotgun proteomics. Physiological and gene ontology analysis confirmed that O. australiensis responded uniquely to drought, with superior leaf water status and enhanced levels of photosynthetic proteins. Distinctive patterns of protein accumulation in drought were observed across the O. australiensis proteome. Photosynthetic and stress-response proteins were more abundant in drought-affected O. glaberrima than O. sativa , and were further enriched in O. australiensis . In contrast, the level of accumulation of photosynthetic proteins decreased when O. sativa underwent drought, while a narrower range of stress-responsive proteins showed increased levels of accumulation. Distinctive proteomic profiles and the accumulated levels of individual proteins with specific functions in response to drought in O. australiensis indicate the importance of this species as a source of stress tolerance genes.

Rice is a critically important food source but yields worldwide are vulnerable to periods of drought. We exposed eight genotypes of upland and lowland rice ( Oryza sativa L. ssp. japonica and indica ) to drought stress at the late vegetative stage, and harvested leaves for label-free shotgun proteomics. Gene ontology analysis was used to identify common drought-responsive proteins in vegetative tissues, and leaf proteins that are unique to individual genotypes, suggesting diversity in the metabolic responses to drought. Eight proteins were found to be induced in response to drought stress in all eight genotypes. A total of 213 proteins were identified in a single genotype, 83 of which were increased in abundance in response to drought stress. In total, 10 of these 83 proteins were of a largely uncharacterized function, making them candidates for functional analysis and potential biomarkers for drought tolerance.

Salinity is a major constraint on rice productivity worldwide. However, mechanisms of salt tolerance in wild rice relatives are unknown. Root microsomal proteins are extracted from two Oryza australiensis accessions contrasting in salt tolerance. Whole roots of 2-week-old seedlings are treated with 80 mM NaCl for 30 days to induce salt stress. Proteins are quantified by tandem mass tags (TMT) and triple-stage Mass Spectrometry. More than 200 differentially expressed proteins between the salt-treated and control samples in the two accessions (p-value <0.05) are found. Gene Ontology (GO) analysis shows that proteins categorized as "metabolic process," "transport," and "transmembrane transporter" are highly responsive to salt treatment. In particular, mitochondrial ATPases and SNARE proteins are more abundant in roots of the salt-tolerant accession and responded strongly when roots are exposed to salinity. mRNA quantification validated the elevated protein abundances of a monosaccharide transporter and an antiporter observed in the salt-tolerant genotype. The importance of the upregulated monosaccharide transporter and a VAMP-like protein by measuring salinity responses of two yeast knockout mutants for genes homologous to those encoding these proteins in rice are confirmed. Potential new mechanisms of salt tolerance in rice, with implications for breeding of elite cultivars are also discussed., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Plants require a distinctive cohort of enzymes to coordinate cell division and expansion. Proteomic analysis now enables interrogation of immature leaf bases where these processes occur. Hence, proteins in tissues sampled from leaves of a drought-tolerant rice (IAC1131) are investigated to provide insights into the effect of soil drying on gene expression relative to the drought-sensitive genotype Nipponbare. Shoot growth zones are dissected to estimate the proportion of dividing cells and extract protein for subsequent tandem mass tags quantitative proteomic analysis. Gene ontology annotations of differentially expressed proteins provide insights into responses of Nipponbare and IAC1131 to drought. Soil drying does not affect the percentage of mitotic cells in IAC1131. More than 800 proteins across most functional categories increase in drought (and decrease on rewatering) in IAC1131, including proteins involved in "organizing the meristem" and "new cell formation". On the other hand, the percentage of dividing cells in Nipponbare is severely impaired during drought and fewer than 200 proteins respond in abundance when growing zones undergo a drying cycle. Remarkably, the proteomes of the growing zones of each genotype respond in a highly distinctive manner, reflecting their contrasting drought tolerance even at the earliest stages of leaf development., (© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Soil salinity is widespread in rice-producing areas globally, restricting both vegetative growth and grain yield. Attempts to improve the salt tolerance of Asian rice, Oryza sativa-the most salt sensitive of the major cereal crops-have met with limited success, due to the complexity of the trait and finite variation in salt responses among O. sativa lines. Naturally occurring variation among the more than 20 wild species of the Oryza genus has great potential to provide breeders with novel genes to improve resistance to salt. Here, through two distinct screening experiments, we investigated variation in salinity tolerance among accessions of two wild rice species endemic to Australia, O. meridionalis and O. australiensis, with O. sativa cultivars Pokkali and IR29 providing salt-tolerant and sensitive controls, respectively., Results: Rice plants were grown on soil supplemented with field-relevant concentrations of NaCl (0, 40, 80, and 100 mM) for 30 d, a period sufficient to reveal differences in growth and physiological traits. Two complementary screening approaches were used: destructive phenotyping and high-throughput image-based phenotyping. All genotypes displayed clear responses to salt treatment. In the first experiment, both salt-tolerant Pokkali and an O. australiensis accession (Oa-VR) showed the least reduction in biomass accumulation, SES score and chlorophyll content in response to salinity. Average shoot Na + /K + values of these plants were the lowest among the genotypes tested. In the second experiment, plant responses to different levels of salt stress were quantified over time based on projected shoot area calculated from visible red-green-blue (RGB) and fluorescence images. Pokkali grew significantly faster than the other genotypes. Pokkali and Oa-VR plants displayed the same absolute growth rate under 80 and 100 mM, while Oa-D grew significantly slower with the same treatments. Oa-VR showed substantially less inhibition of growth in response to salinity when compared with Oa-D. Senescence was seen in Oa-D after 30 d treatment with 40 mM NaCl, while the putatively salt-tolerant Oa-VR had only minor leaf damage, even at higher salt treatments, with less than a 40% increase in relative senescence at 100 mM NaCl compared to 120% for Oa-VR., Conclusion: The combination of our two screening experiments uncovered striking levels of salt tolerance diversity among the Australian wild rice accessions tested and enabled analysis of their growth responses to a range of salt levels. Our results validate image-based phenotyping as a valuable tool for quantitative measurement of plant responses to abiotic stresses. They also highlight the potential of exotic germplasm to provide new genetic variation for salinity tolerance in rice.

Two rice cultivars, IAC1131 (drought tolerant) and Nipponbare (drought sensitive), with contrasting genetic backgrounds and levels of tolerance to drought, were analysed using both label-free and tandem mass tags (TMTs) quantitative proteomics approaches, aiming to elucidate the mechanisms of drought tolerance. Four-week-old seedlings of both cultivars were grown in large soil volumes in the glasshouse under controlled conditions and then exposed to moderate and extreme drought for 7 days, followed by 3 days of re-watering period. Mature leaves were harvested from plants of each treatment for protein extraction and subsequent complementary shotgun proteomic analyses. The data from this study are related to the research article "Quantitative proteomic analysis of two different rice varieties reveals that drought tolerance is correlated with reduced abundance of photosynthetic machinery and increased abundance of ClpD1 protease" (Wu et al., 2016) [1].

Genes encoding thermostable variants of the photosynthesis heat-labile protein Rubisco activase (Rca) from a wild relative Oryza australiensis were overexpressed in domesticated rice ( Oryza sativa ). Proteomics was used to quantify the abundance of O. australiensis Rca (Rca- Oa ) in the resulting plants. Plants were grown to maturity in growth rooms and from early tillering until immediately prior to anthesis, they were exposed to daytime maximum temperatures of 28, 40, and 45°C and constant night temperatures of 22°C. Non-destructive measurements of leaf elongation and photosynthesis were used to compare the null segregant with a transfected line in which 19% of its total Rca content was the recombinant O. australiensis Rca (T- Oa -19). Height, fresh mass, panicle number, seed set, and seed number were measured at final harvest. Traits at maturity after heat stress at 45°C correlated strongly with recombinant protein abundance. Seed number was far the most responsive trait to an increase in Rca- Oa abundance, improving by up to 150%. Leaf elongation rates ( LER ) and tiller number were significantly greater in the transformed plants in the first two weeks of exposure to 45°C but tiller numbers later became equal in the two genotypes. Gas exchange measurements showed that T- Oa -19 had faster light induction of photosynthesis but not significantly higher CO 2 assimilation rates, indicating that the carbon gain that resulted in large yield improvement after growth at 45°C was not strongly correlated with an instantaneous measurement of steady-state photosynthesis. When plants were grown at 40°C daytime maximum, there was no improvement in the final biomass, panicle or seed number when compared with 28°C, indicating that the threshold for heat damage and beneficial effects of the thermostable Rca recombinant protein was between 40 and 45°C, which corresponded to leaf temperatures in the range 38-42°C. The results suggest that the thermotolerant form of Rca from O. australiensis was sufficient to enhance carbohydrate accumulation and storage by rice over the life of the plant, dramatically improving yields after exposure to heat throughout the vegetative phase.

Soil waterlogging and subsequent ethylene release from cotton (Gossypium hirsutum L.) tissues has been linked with abscission of developing cotton fruits. This glasshouse study investigates the effect of a 9-day waterlogging event and CO2 enrichment (eCO2, 700 parts per million (ppm)) on a fully linted cultivar 'Empire' and a lintless cotton mutant (5B). We hypothesised that cotton performance in extreme environments such as waterlogging can be improved through mitigating ethylene action. Plants were grown at 28:20°C day:night temperature, 50-70% relative humidity and a 14:10 light:dark photoperiod under natural light and were exposed to waterlogging and eCO2 at early reproductive growth. Ethylene synthesis was inhibited by spraying aminoethoxyvinylglycine (830ppm) 1 day before waterlogging. Waterlogging significantly increased ethylene release from both cotton genotypes, although fruit production was significantly inhibited only in Empire. Aminoethoxyvinylglycine consistently reduced waterlogging-induced abscission of fruits, mainly in Empire. Limited damage to fruits in 5B, despite increased ethylene production during waterlogging, suggested that fruit abscission in 5B was inhibited by disrupting ethylene metabolism genetically. Elevated CO2 promoted fruit production in both genotypes and was more effective in 5B than in Empire plants. Hence 5B produced more fruits than Empire, providing additional sinks (existing and new fruit) that enhanced the response to CO2 enrichment.

We investigated the role of ethylene in the response of cotton to high temperature using cotton genotypes with genetically interrupted ethylene metabolism. In the first experiment, Sicot 71BRF and 5B (a lintless variant with compromised ethylene metabolism) were exposed to 45°C, either by instantaneous heat shock or by ramping temperatures by 3°C daily for 1 week. One day prior to the start of heat treatment, half the plants were sprayed with 0.8 mM of the ethylene synthesis inhibitor, aminoethoxyvinylglycine (AVG). In a subsequent experiment, Sicot 71BRF and a putatively heat-tolerant line, CIM 448, were exposed to 36 or 45°C for 1 week, and half the plants were sprayed with 20 μM of the ethylene precursor, 1-aminocyclopropane-1-carboxylic acid, (ACC). High temperature exposure of plants in both experiments was performed at the peak reproductive phase (65-68 days after sowing). Elevated temperature (heat shock or ramping to 45°C) significantly reduced production and retention of fruits in all cotton lines used in this study. At the termination of heat treatment, cotton plants exposed to 45°C had at least 50% fewer fruits than plants under optimum temperature in all three genotypes, while plants at 36°C remained unaffected. Heat-stressed plants continued producing new squares (fruiting buds) after termination of heat stress but these squares did not turn into cotton bolls due to pollen infertility. In vitro inhibition of pollen germination by high temperatures supported this observation. Leaf photosynthesis ( P n ) of heat-stressed plants (45°C) measured at the end of heat treatments remained significantly inhibited, despite an increased leaf stomatal conductance ( g s ), suggesting that high temperature impairs P n independently of stomatal behavior. Metabolic injury was supported by high relative cellular injury and low photosystem II yield of the heat-stressed plants, indicating that high temperature impaired photosynthetic electron transport. Both heat shock and ramping of heat significantly reduced ethylene release from cotton leaf tissues measured at the end of heat treatment but modulating ethylene production via AVG or ACC application had no significant effect on fruit production or retention in heat-stressed cotton plants. Instead, high temperature accelerated fruit abortion by impairing pollen development and/or restricting leaf photosynthesis.

The mechanistic basis of tolerance to heat stress was investigated in Oryza sativa and two wild rice species, Oryza meridionalis and Oryza australiensis. The wild relatives are endemic to the hot, arid Australian savannah. Leaf elongation rates and gas exchange were measured during short periods of supra-optimal heat, revealing species differences. The Rubisco activase (RCA) gene from each species was sequenced. Using expressed recombinant RCA and leaf-extracted RCA, the kinetic properties of the two isoforms were studied under high temperatures. Leaf elongation was undiminished at 45°C in O. australiensis. The net photosynthetic rate was almost 50% slower in O. sativa at 45°C than at 28°C, while in O. australiensis it was unaffected. Oryza meridionalis exhibited intermediate heat tolerance. Based on previous reports that RCA is heat-labile, the Rubisco activation state was measured. It correlated positively with leaf elongation rates across all three species and four periods of exposure to 45°C. Sequence analysis revealed numerous polymorphisms in the RCA amino acid sequence from O. australiensis. The O. australiensis RCA enzyme was thermally stable up to 42°C, contrasting with RCA from O. sativa, which was inhibited at 36°C. We attribute heat tolerance in the wild species to thermal stability of RCA, enabling Rubisco to remain active., (© 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.)

Unlabelled: Rice is the major staple food for more than half of world's population. As global climate changes, we are observing more floods, droughts and severe heat waves. Two rice cultivars with contrasting genetic backgrounds and levels of tolerance to drought, Nipponbare and IAC1131, were used in this study. Four-week-old seedlings of both cultivars were grown in large soil volumes and then exposed to moderate and extreme drought for 7days, followed by 3days of re-watering. Mature leaves were harvested from plants from each treatment for protein extraction and subsequent shotgun proteomic analysis, with validation of selected proteins by western blotting. Gene Ontology (GO) annotations of differentially expressed proteins provide insights into the metabolic pathways that are involved in drought stress resistance. Our data indicate that IAC1131 appears to be better able to cope with stressful conditions by upregulating a suite of stress and defence response related proteins. Nipponbare, in contrast, lacks the range of stress responses shown by the more stress tolerant variety, and responds to drought stress by initiating a partial shutdown of chlorophyll biosynthesis in an apparent attempt to preserve resources., Significance: In this study, two rice genotypes with contrasting drought tolerance were exposed to soil water deficits, and proteomic changes were observed in mature leaf laminae. Plants were well watered and then switched to conditions of either moderate drought or extreme drought followed by three days of recovery. Proteins were identified and quantified using both label-free and Tandem Mass Tag multiplexing approaches. Several biochemical pathways were significantly altered in response to water deficit. Most notably, the up-regulation of ClpD1 protease responded strongly in the drought-tolerant landrace; this protein is typically involved in heat and osmotic stress response. In contrast, porphyrin and chlorophyll biosynthesis pathways were down-regulated, indicating suppression of the photosynthetic machinery., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Models of tree responses to climate typically project that elevated atmospheric CO2 concentration (eCa ) will reduce drought impacts on forests. We tested one of the mechanisms underlying this interaction, the 'low Ci effect', in which stomatal closure in drought conditions reduces the intercellular CO2 concentration (Ci ), resulting in a larger relative enhancement of photosynthesis with eCa , and, consequently, a larger relative biomass response. We grew two Eucalyptus species of contrasting drought tolerance at ambient and elevated Ca for 6-9 months in large pots maintained at 50% (drought) and 100% field capacity. Droughted plants did not have significantly lower Ci than well-watered plants, which we attributed to long-term changes in leaf area. Hence, there should not have been an interaction between eCa and water availability on biomass, and we did not detect one. The xeric species did have higher Ci than the mesic species, indicating lower water-use efficiency, but both species exhibited similar responses of photosynthesis and biomass to eCa , owing to compensatory differences in the photosynthetic response to Ci . Our results demonstrate that long-term acclimation to drought, and coordination among species traits may be important for predicting plant responses to eCa under low water availability., (© 2015 The Authors. New Phytologist © 2015 New Phytologist Trust.)

Climatic variability, typified by erratic heavy-rainfall events, causes waterlogging in intensively irrigated crops and is exacerbated under warm temperature regimes on soils with poor internal drainage. Irrigated cotton is often grown in precisely these conditions, exposing it to waterlogging-induced yield losses after substantial summer rainfall. This calls for a deeper understanding of mechanisms of waterlogging tolerance and its relevance to cotton. Hence this review suggests possible causes of waterlogging-induced yield loss in cotton and approaches to improvement of waterlogging tolerance, drawing upon the slight body of published data in cotton and principles from other species. The yield penalty depends on soil type, phenological stage and cumulative period of root exposure to air-filled porosities below 10 %. Events in the soil include O2 deficiency in the root zone that changes the redox state of nutrients, making them unavailable (e.g. nitrogen) or potentially toxic for plants. Furthermore, root-derived hormones that are transported in the xylem have long been associated with oxygen deficits. These belowground effects (impaired root growth, nutrient uptake and transport, hormonal signalling) affect the shoots, interfering with canopy development, photosynthesis and radiation-use efficiency. Compared with the more waterlogging-tolerant cereals, cotton does not have identified adaptations to waterlogging in the root zone, forming no conspicuous root aerenchyma and having low fermentative activity. We speculate that these factors contribute substantially to the sensitivity of cotton to sustained periods of waterlogging. We discuss the impact of these belowground factors on shoot performance, photosynthesis and yield components. Management practices, i.e. soil aeration, scheduling irrigation and fertilizer application, can reduce waterlogging-induced damage. Limiting ethylene biosynthesis using anti-ethylene agents and down-regulating expression of genes controlling ethylene biosynthesis are strong candidates to minimize yield losses in waterlogged cotton crops. Other key pathways of anoxia tolerance are also cited as potential tools towards waterlogging-tolerant cotton genotypes., (Published by Oxford University Press on behalf of the Annals of Botany Company.)

Anoxia tolerance in plants is distinguished by direction of the sparse supply of energy to processes crucial to cell maintenance and sometimes to growth, as in rice seedlings. In anoxic rice coleoptiles energy is used to synthesise proteins, take up K(+) , synthesise cell walls and lipids, and in cell maintenance. Maintenance of electrochemical H(+) gradients across the tonoplast and plasma membrane is crucial for solute compartmentation and thus survival. These gradients sustain some H(+) -solute cotransport and regulate cytoplasmic pH. Pyrophosphate (PPi ), the alternative energy donor to ATP, allows direction of energy to the vacuolar H(+) -PPi ase, sustaining H(+) gradients across the tonoplast. When energy production is critically low, operation of a biochemical pHstat allows H(+) -solute cotransport across plasma membranes to continue for at least for 18 h. In active (e.g. growing) cells, PPi produced during substantial polymer synthesis allows conversion of PPi to ATP by PPi -phosphofructokinase (PFK). In quiescent cells with little polymer synthesis and associated PPi formation, the PPi required by the vacuolar H(+) -PPi ase and UDPG pyrophosphorylase involved in sucrose mobilisation via sucrose synthase might be produced by conversion of ATP to PPi through reversible glycolytic enzymes, presumably pyruvate orthophosphate dikinase. These hypotheses need testing with species characterised by contrasting anoxia tolerance., (© 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.)

Coleoptiles of rice (Oryza sativa) seedlings grown under water commonly elongate by up to 1 mm h(-1) to reach the atmosphere. We initially analysed this highly specialized phenomenon by measuring epidermal cell lengths along the coleoptile axis to determine elongation rates. This revealed a cohort of cells in the basal zone that elongated rapidly following emergence from the embryo, reaching 200 μm within 12 h. After filming coleoptiles in vivo for a day, kinematic analysis was applied. Eight time-sliced 'segments' were defined by their emergence from the embryo at four-hourly intervals, revealing a mathematically simple growth model. Each segment entering the coleoptile from the embryo elongated at a constant velocity, resulting in accelerating growth for the entire organ. Consistent with the epidermal cell lengths, relative rates of elongation (mm mm(-1) h(-1)) were tenfold greater in the small, newly emerged basal segments than the older distal tip segments. This steep axial gradient defined two contrasting growth zones (bases versus tips) in which we measured ATP production and protein, RNA and DNA content, and analysed the global transcriptome under steady-state normoxia, hypoxia (3% O2) and anoxia. Determination of the transcriptome revealed tip-specific induction of genes encoding TCP [Teosinte Branched1 (Tb1) of maize, Cycloidea (Cyc), and Proliferating Cell Factor (Pcf)] transcription factors, RNA helicases, ribosomal proteins and proteins involved in protein folding, whilst expression of F-box domain-containing proteins in the ubiquitin E3-SCF complex (Skp, Cullin, F-box containing complex) was induced specifically in bases under low oxygen conditions. We ascribed the sustained elongation under hypoxia to hypoxia-specific responses such as controlled suppression of photosystem components and induction of RNA binding/splicing functions, indicating preferential allocation of energy to cell extension., (© 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.)

The effect of drought on plant isoprene emission varies tremendously across species and environments. It was recently shown that an increased ratio of photosynthetic electron transport rate (ETR) to net carbon assimilation rate (NAR) consistently supported increased emission under drought. In this commentary, we highlight some of the physiological aspects of drought tolerance that are central to the observed variability. We briefly discuss some of the issues that must be addressed in order to refine our understanding of plant isoprene emission response to drought and increasing global temperature.

Plants undergoing heat and low-CO2 stresses emit large amounts of volatile isoprenoids compared with those in stress-free conditions. One hypothesis posits that the balance between reducing power availability and its use in carbon assimilation determines constitutive isoprenoid emission rates in plants and potentially even their maximum emission capacity under brief periods of stress. To test this, we used abiotic stresses to manipulate the availability of reducing power. Specifically, we examined the effects of mild to severe drought on photosynthetic electron transport rate (ETR) and net carbon assimilation rate (NAR) and the relationship between estimated energy pools and constitutive volatile isoprenoid emission rates in two species of eucalypts: Eucalyptus occidentalis (drought tolerant) and Eucalyptus camaldulensis (drought sensitive). Isoprenoid emission rates were insensitive to mild drought, and the rates increased when the decline in NAR reached a certain species-specific threshold. ETR was sustained under drought and the ETR-NAR ratio increased, driving constitutive isoprenoid emission until severe drought caused carbon limitation of the methylerythritol phosphate pathway. The estimated residual reducing power unused for carbon assimilation, based on the energetic status model, significantly correlated with constitutive isoprenoid emission rates across gradients of drought (r(2) > 0.8) and photorespiratory stress (r(2) > 0.9). Carbon availability could critically limit emission rates under severe drought and photorespiratory stresses. Under most instances of moderate abiotic stress levels, increased isoprenoid emission rates compete with photorespiration for the residual reducing power not invested in carbon assimilation. A similar mechanism also explains the individual positive effects of low-CO2, heat, and drought stresses on isoprenoid emission., (© 2014 American Society of Plant Biologists. All Rights Reserved.)

Light-dependent de novo volatile isoprene emission by terrestrial plants (approximately 2% of carbon fixed during photosynthesis) contributes as much as 0.5 PgC/year to the global carbon cycle. Although most plant taxa exhibit either constitutive or inducible monoterpene emissions, the evolution of isoprene emission capacity in multiple lineages has remained unexplained. Based on the predominant occurrence of isoprene emission capacity in long-lived, fast-growing woody plants; the relationship between 'metabolic scope' of tree genera and their species richness; and the proposed role of high growth rates and long generation times in accelerating molecular evolution, we hypothesise that long-lived plant genera with inherently high speciation rates have repeatedly acquired and lost the capacity to emit isoprene in their evolutionary history., (Crown Copyright © 2014. Published by Elsevier Ltd. All rights reserved.)

Substantial reductions in yield caused by drought stress can occur when parts of the root system experience water deficit even though other parts have sufficient access to soil water. To identify proteins associated to drought signaling, rice (Oryza sativa L. cv. IR64.) plants were transplanted into plastic pots with an internal wall dividing each pot into two equal compartments, allowing for equal distribution of soil and the root system between these compartments. The following treatments were applied: either both compartments were watered daily ("wet" roots), or water was withheld from both compartments ("dry" roots), or water was withheld from only one of the two compartments in each pot ("wet" and "dry" roots). The substantial differences in physiological parameters of different growth conditions were accompanied by differential changes in protein abundances. Label-free quantitative shotgun proteomics have resulted in identification of 1383 reproducible proteins across all three conditions. Differentially expressed proteins were categorized within 17 functional groups. The patterns observed were interesting in that in some categories such as protein metabolism and oxidation-reduction, substantial numbers of proteins were most abundant when leaves were receiving signals from "wet" and "dry" roots. In yet other categories such as transport, several key transporters were surprisingly abundant in leaves supported by partially or completely droughted root systems, especially plasma membrane and vacuolar transporters. Stress-related proteins behaved very consistently by increasing in droughted plants but notably some proteins were most abundant when roots of the same plant were growing in both wet and dry soils. Changes in carbohydrate-processing proteins were consistent with the passive accumulation of soluble sugars in shoots under drought, with hydrolysis of sucrose and starch synthesis both enhanced. These results suggest that drought signals are complex interactions and not simply the additive effect of water supply to the roots.

Oryza sativa and Oryza glaberrima have been selected to acquire and partition resources efficiently as part of the process of domestication. However, genetic diversity in cultivated rice is limited compared to wild Oryza species, in spite of 120,000 genotypes being held in gene banks. By contrast, there is untapped diversity in the more than 20 wild species of Oryza, some having been collected from just a few coastal locations (e.g. Oryza schlechteri), while others are widely distributed (e.g. Oryza nivara and Oryza rufipogon). The extent of DNA sequence diversity and phenotypic variation is still being established in wild Oryza, with genetic barriers suggesting a vast range of morphologies and function even within species, such as has been demonstrated for Oryza meridionalis. With increasing climate variability and attempts to make more marginal land arable, abiotic and biotic stresses will be managed over the coming decades by tapping into the genetic diversity of wild relatives of O. sativa. To help create a more targeted approach to sourcing wild rice germplasm for abiotic stress tolerance, we have created a climate distribution map by plotting the natural occurrence of all Oryza species against corresponding temperature and moisture data. We then discuss interspecific variation in phenotype and its significance for rice, followed by a discussion of ways to integrate germplasm from wild relatives into domesticated rice., (Crown Copyright © 2013. Published by Elsevier Ireland Ltd. All rights reserved.)

Low root temperature causes a decrease in water uptake, which leads to mineral and nutrient deficiencies with potentially decreased root and shoot growth. Differential temperature effects in plants have been studied extensively, however, the effect of root chilling on the global protein expression in shoots has not been explored. In this study, we imposed chilling temperatures on roots of rice plants while maintaining shoots at optimum atmospheric temperature. Shoot materials (growing zones and leaves) were harvested at five points over a time course of four days, including a two-day recovery period. Proteins were quantified by tandem mass tags and triple stage MS, using a method developed to overcome ratio compression in isobaric-labelled quantitation. Over 3000 proteins in each of the tissues were quantified by multiple peptides. Proteins significantly differentially expressed as compared with the control included abscisic acid-responsive and drought-associated proteins. The data also contained evidence of a possible induction of a sugar signalling pathway., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: Most members of the serpin family of proteins are potent, irreversible inhibitors of specific serine or cysteine proteinases. Inhibitory serpins are distinguished from members of other families of proteinase inhibitors by their metastable structure and unique suicide-substrate mechanism. Animal serpins exert control over a remarkable diversity of physiological processes including blood coagulation, fibrinolysis, innate immunity and aspects of development. Relatively little is known about the complement of serpin genes in plant genomes and the biological functions of plant serpins., Results: A structurally refined amino-acid sequence alignment of the 14 full-length serpins encoded in the genome of the japonica rice Oryza sativa cv. Nipponbare (a monocot) showed a diversity of reactive-centre sequences (which largely determine inhibitory specificity) and a low degree of identity with those of serpins in Arabidopsis (a eudicot). A new convenient and functionally informative nomenclature for plant serpins in which the reactive-centre sequence is incorporated into the serpin name was developed and applied to the rice serpins. A phylogenetic analysis of the rice serpins provided evidence for two main clades and a number of relatively recent gene duplications. Transcriptional analysis showed vastly different levels of basal expression among eight selected rice serpin genes in callus tissue, during seedling development, among vegetative tissues of mature plants and throughout seed development. The gene OsSRP-LRS (Os03g41419), encoding a putative orthologue of Arabidopsis AtSerpin1 (At1g47710), was expressed ubiquitously and at high levels. The second most highly expressed serpin gene was OsSRP-PLP (Os11g11500), encoding a non-inhibitory serpin with a surprisingly well-conserved reactive-centre loop (RCL) sequence among putative orthologues in other grass species., Conclusions: The diversity of reactive-centre sequences among the putatively inhibitory serpins of rice point to a range of target proteases with different proteolytic specificities. Large differences in basal expression levels of the eight selected rice serpin genes during development further suggest a range of functions in regulation and in plant defence for the corresponding proteins.

Oryza meridionalis is a wild species of rice, endemic to tropical Australia. It shares a significant genome homology with the common domesticated rice Oryza sativa. Exploiting the fact that the two species are highly related but O. meridionalis has superior heat tolerance, experiments were undertaken to identify the impact of temperature on key events in photosynthesis. At an ambient CO(2) partial pressure of 38 Pa and irradiance of 1500 µmol quanta m(-2) s(-1), the temperature optimum of photosynthesis was 33.7 ± 0.8°C for O. meridionalis, significantly higher than the 30.6 ± 0.7°C temperature optimum of O. sativa. To understand the basis for this difference, we measured gas exchange and rubisco activation state between 20 and 42°C and modeled the response to determine the rate-limiting steps of photosynthesis. The temperature response of light respiration (R(light)) and the CO(2) compensation point in the absence of respiration (Γ(*)) were determined and found to be similar for the two species. C3 photosynthesis modeling showed that despite the difference in susceptibility to high temperature, both species had a similar temperature-dependent limitation to photosynthesis. Both rice species were limited by ribulose-1,5-bisphosphate (RuBP) regeneration at temperatures of 25 and 30°C but became RuBP carboxylation limited at 35 and 40°C. The activation state of rubisco in O. meridionalis was more stable at higher temperatures, explaining its greater heat tolerance compared with O. sativa., (Copyright © Physiologia Plantarum 2012.)

Oxygen deprivation limits the energy available for cellular processes and yet no comprehensive ATP budget has been reported for any plant species under O(2) deprivation, including Oryza sativa. Using 3-d-old coleoptiles of a cultivar of O. sativa tolerant to flooding at germination, (i) rates of ATP regeneration in coleoptiles grown under normoxia (aerated solution), hypoxia (3% O(2)), and anoxia (N(2)) and (ii) rates of synthesis of proteins, lipids, nucleic acids, and cell walls, as well as K(+) transport, were determined. Based on published bioenergetics data, the cost of synthesizing each class of polymer and the proportion of available ATP allocated to each process were then compared. Protein synthesis consumed the largest proportion of ATP synthesized under all three oxygen regimes, with the proportion of ATP allocated to protein synthesis in anoxia (52%) more than double that in normoxic coleoptiles (19%). Energy allocation to cell wall synthesis was undiminished in hypoxia, consistent with preferential elongation typical of submerged coleoptiles. Lipid synthesis was also conserved strongly in O(2) deficits, suggesting that membrane integrity was maintained under anoxia, thus allowing K(+) to be retained within coleoptile cells. Rates of protein synthesis in coleoptiles from rice cultivars with contrasting tolerance to oxygen deficits (including mutants deficient in fermentative enzymes) confirmed that synthesis and turnover of proteins always accounted for most of the ATP consumed under anoxia. It is concluded that successful establishment of rice seedlings under water is largely due to the capacity of coleoptiles to allocate energy to vital processes, particularly protein synthesis.

Mechanisms of drought tolerance are complex, interacting, and polygenic. This paper describes patterns of gene expression at precise physiological stages of drought in 35-day-old seedlings of Oryza sativa cv. Nipponbare. Drought was imposed gradually for up to 15 days, causing abscisic acid levels to rise and growth to cease, and plants were then re-watered. Proteins were identified from leaf samples after moderate drought, extreme drought, and 3 and 6 days of re-watering. Label-free quantitative shotgun proteomics resulted in identification of 1548 non-redundant proteins. More proteins were down-regulated in early stages of drought but more were up-regulated as severe drought developed. After re-watering, there was notable down regulation, suggesting that stress-related proteins were being degraded. Proteins involved in signalling and transport became dominant as severe drought took hold but decreased again on re-watering. Most of the nine aquaporins identified were responsive to drought, with six decreasing rapidly in abundance as plants were re-watered. Nine G-proteins appeared in large amounts during severe drought and dramatically degraded once plants were re-watered. We speculate that water transport and drought signalling are critical elements of the overall response to drought in rice and might be the key to biotechnological approaches to drought tolerance., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

• Plant light interception efficiency is a crucial determinant of carbon uptake by individual plants and by vegetation. Our aim was to identify whole-plant variables that summarize complex crown architecture, which can be used to predict light interception efficiency. • We gathered the largest database of digitized plants to date (1831 plants of 124 species), and estimated a measure of light interception efficiency with a detailed three-dimensional model. Light interception efficiency was defined as the ratio of the hemispherically averaged displayed to total leaf area. A simple model was developed that uses only two variables, crown density (the ratio of leaf area to total crown surface area) and leaf dispersion (a measure of the degree of aggregation of leaves). • The model explained 85% of variation in the observed light interception efficiency across the digitized plants. Both whole-plant variables varied across species, with differences in leaf dispersion related to leaf size. Within species, light interception efficiency decreased with total leaf number. This was a result of changes in leaf dispersion, while crown density remained constant. • These results provide the basis for a more general understanding of the role of plant architecture in determining the efficiency of light harvesting., (© 2011 The Authors. New Phytologist © 2011 New Phytologist Trust.)

Rice (Oryza sativa L. cv. IR64) was grown in split-root systems to analyze long-distance drought signaling within root systems. This in turn underpins how root systems in heterogeneous soils adapt to drought. The approach was to compare four root tissues: (1) fully watered; (2) fully droughted and split-root systems where (3) one-half was watered and (4) the other half was droughted. This was specifically aimed at identifying how droughted root tissues altered the proteome of adjacent wet roots by hormone signals and how wet roots reciprocally affected dry roots hydraulically. Quantitative label-free shotgun proteomic analysis of four different root tissues resulted in identification of 1487 nonredundant proteins, with nearly 900 proteins present in triplicate in each treatment. Drought caused surprising changes in expression, most notably in partially droughted roots where 38% of proteins were altered in level compared to adjacent watered roots. Specific functional groups changed consistently in drought. Pathogenesis-related proteins were generally up-regulated in response to drought and heat-shock proteins were totally absent in roots of fully watered plants. Proteins involved in transport and oxidation-reduction reactions were also highly dependent upon drought signals, with the former largely absent in roots receiving a drought signal while oxidation-reduction proteins were strongly present during drought. Finally, two functionally contrasting protein families were compared to validate our approach, showing that nine tubulins were strongly reduced in droughted roots while six chitinases were up-regulated, even when the signal arrived remotely from adjacent droughted roots.