Genetic Components of Root Architecture Remodeling in Response to Salt Stress.

Salinity of the soil is highly detrimental to plant growth. Plants respond by a redistribution of root mass between main and lateral roots, yet the genetic machinery underlying this process is still largely unknown. Here, we describe the natural variation among 347 Arabidopsis thaliana accessions in root system architecture (RSA) and identify the traits with highest natural variation in their response to salt. Salt-induced changes in RSA were associated with 100 genetic loci using genome-wide association studies. Two candidate loci associated with lateral root development were validated and further investigated. Changes in CYP79B2 expression in salt stress positively correlated with lateral root development in accessions, and cyp79b2 cyp79b3 double mutants developed fewer and shorter lateral roots under salt stress, but not in control conditions. By contrast, high HKT1 expression in the root repressed lateral root development, which could be partially rescued by addition of potassium. The collected data and multivariate analysis of multiple RSA traits, available through the Salt_NV_Root App, capture root responses to salinity. Together, our results provide a better understanding of effective RSA remodeling responses, and the genetic components involved, for plant performance in stress conditions.

Strategies for engineering improved nitrogen use efficiency in crop plants via redistribution and recycling of organic nitrogen.

Global use of nitrogen (N) fertilizers has increased sevenfold from 1960 to 1995 but much of the N applied is lost to the environment. Modifying the temporal and spatial distribution of organic N within the plant can lead to improved grain yield and/or grain protein content for the same or reduced N fertilizer inputs. Biotechnological approaches to modify whole plant distribution of amino acids and ureides has proven successful in several crop species. Manipulating selective autophagy pathways in crops has also improved N remobilization efficiency to sink tissues whilst the contribution of ribophagy, RNA and purine catabolism to N recycling in crops is still too early to foretell. Improved recycling and remobilization of N must exploit N-stress responsive transcriptional regulators, N-sensing or phloem-localized promotors and genetic variation for N-responsive traits.

Investigation of a His-rich arabinogalactan-protein for micronutrient biofortification of cereal grain.

The micronutrient content of most cereal grains is low and responsible for malnutrition deficiencies in millions of people who rely on grains as their primary food source. Any strategy that can increase the micronutrient content of grain will have significant benefits to world health. We identified a gene from barley encoding a cell wall protein with multiple histidine (His)-rich motifs interspersed with short arabinogalactan-protein (AGP) domains and have called it Hordeum vulgare His-rich AGP (HvHRA1). Sequence analysis shows that His-rich AGPs are rare in plants and that the number of His-rich and AGP domains differ between cereals and dicots. The barley and wheat encoded proteins have more than 13 His-rich domains, whereas the putative rice orthologue has only 5 His-rich regions. His-rich motifs are well-established metal-binding motifs; therefore, we developed transgenic (Tx) rice plants that constitutively overexpress barley HvHRA1. There was no significant effect on plant growth or grain yield in Tx plants. Purification of AGPs from wild-type and Tx plants showed that only Tx plants contained detectable levels of a His-rich AGP. Calcein assay shows that the AGP fraction from Tx plants had increased binding affinity for Cu(2+) . Micronutrient analysis of brown and white rice showed that the grain nutrient yield for Fe, Zn and Cu was higher in two Tx lines compared to their respective nulls, although the differences were not statistically significant. This approach highlights the potential of the plant apoplast (cell wall) for storage of key nutrients through overexpression of genes for metal-binding proteins.

The Use of High-Throughput Phenotyping for Assessment of Heat Stress-Induced Changes in Arabidopsis.

The worldwide rise in heatwave frequency poses a threat to plant survival and productivity. Determining the new marker phenotypes that show reproducible response to heat stress and contribute to heat stress tolerance is becoming a priority. In this study, we describe a protocol focusing on the daily changes in plant morphology and photosynthetic performance after exposure to heat stress using an automated noninvasive phenotyping system. Heat stress exposure resulted in an acute reduction of the quantum yield of photosystem II and increased leaf angle. In longer term, the exposure to heat also affected plant growth and morphology. By tracking the recovery period of the WT and mutants impaired in thermotolerance (hsp101), we observed that the difference in maximum quantum yield, quenching, rosette size, and morphology. By examining the correlation across the traits throughout time, we observed that early changes in photochemical quenching corresponded with the rosette size at later stages, which suggests the contribution of quenching to overall heat tolerance. We also determined that 6 h of heat stress provides the most informative insight in plant's responses to heat, as it shows a clear separation between treated and nontreated plants as well as the WT and hsp101. Our work streamlines future discoveries by providing an experimental protocol, data analysis pipeline, and new phenotypes that could be used as targets in thermotolerance screenings.

Predicting Biomass and Yield in a Tomato Phenotyping Experiment Using UAV Imagery and Random Forest.

Biomass and yield are key variables for assessing the production and performance of agricultural systems. Modeling and predicting the biomass and yield of individual plants at the farm scale represents a major challenge in precision agriculture, particularly when salinity and other abiotic stresses may play a role. Here, we evaluate a diversity panel of the wild tomato species (Solanum pimpinellifolium) through both field and unmanned aerial vehicle (UAV)-based phenotyping of 600 control and 600 salt-treated plants. The study objective was to predict fresh shoot mass, tomato fruit numbers, and yield mass at harvest based on a range of variables derived from the UAV imagery. UAV-based red-green-blue (RGB) imageries collected 1, 2, 4, 6, 7, and 8 weeks before harvest were also used to determine if prediction accuracies varied between control and salt-treated plants. Multispectral UAV-based imagery was also collected 1 and 2 weeks prior to harvest to further explore predictive insights. In order to estimate the end of season biomass and yield, a random forest machine learning approach was implemented using UAV-imagery-derived predictors as input variables. Shape features derived from the UAV, such as plant area, border length, width, and length, were found to have the highest importance in the predictions, followed by vegetation indices and the entropy texture measure. The multispectral UAV imagery collected 2 weeks prior to harvest produced the highest explained variances for fresh shoot mass (87.95%), fruit numbers (63.88%), and yield mass per plant (66.51%). The RGB UAV imagery produced very similar results to those of the multispectral UAV dataset, with the explained variance reducing as a function of increasing time to harvest. The results showed that predicting the yield of salt-stressed plants produced higher accuracies when the models excluded control plants, whereas predicting the yield of control plants was not affected by the inclusion of salt-stressed plants within the models. This research demonstrates that it is possible to predict the average biomass and yield up to 8 weeks prior to harvest within 4.23% of field-based measurements and up to 4 weeks prior to harvest at the individual plant level. Results from this work may be useful in providing guidance for yield forecasting of healthy and salt-stressed tomato plants, which in turn may inform growing practices, logistical planning, and sales operations.

Unmanned Aerial Vehicle-Based Phenotyping Using Morphometric and Spectral Analysis Can Quantify Responses of Wild Tomato Plants to Salinity Stress.

With salt stress presenting a major threat to global food production, attention has turned to the identification and breeding of crop cultivars with improved salt tolerance. For instance, some accessions of wild species with higher salt tolerance than commercial varieties are being investigated for their potential to expand food production into marginal areas or to use brackish waters for irrigation. However, assessment of individual plant responses to salt stress in field trials is time-consuming, limiting, for example, longitudinal assessment of large numbers of plants. Developments in Unmanned Aerial Vehicle (UAV) sensing technologies provide a means for extensive, repeated and consistent phenotyping and have significant advantages over standard approaches. In this study, 199 accessions of the wild tomato species, Solanum pimpinellifolium, were evaluated through a field assessment of 600 control and 600 salt-treated plants. UAV imagery was used to: (1) delineate tomato plants from a time-series of eight RGB and two multi-spectral datasets, using an automated object-based image analysis approach; (2) assess four traits, i.e., plant area, growth rates, condition and Plant Projective Cover (PPC) over the growing season; and (3) use the mapped traits to identify the best-performing accessions in terms of yield and salt tolerance. For the first five campaigns, >99% of all tomato plants were automatically detected. The omission rate increased to 2-5% for the last three campaigns because of the presence of dead and senescent plants. Salt-treated plants exhibited a significantly smaller plant area (average control and salt-treated plant areas of 0.55 and 0.29 m2, respectively), maximum growth rate (daily maximum growth rate of control and salt-treated plant of 0.034 and 0.013 m2, respectively) and PPC (5-16% difference) relative to control plants. Using mapped plant condition, area, growth rate and PPC, we show that it was possible to identify eight out of the top 10 highest yielding accessions and that only five accessions produced high yield under both treatments. Apart from showcasing multi-temporal UAV-based phenotyping capabilities for the assessment of plant performance, this research has implications for agronomic studies of plant salt tolerance and for optimizing agricultural production under saline conditions.

Variation for N Uptake System in Maize: Genotypic Response to N Supply.

An understanding of the adaptations made by plants in their nitrogen (N) uptake systems in response to reduced N supply is important to the development of cereals with enhanced N uptake efficiency (NUpE). Twenty seven diverse genotypes of maize (Zea mays, L.) were grown in hydroponics for 3 weeks with limiting or adequate N supply. Genotypic response to N was assessed on the basis of biomass characteristics and the activities of the nitrate ([Formula: see text]) and ammonium ([Formula: see text]) high-affinity transport systems. Genotypes differed greatly for the ability to maintain biomass with reduced N. Although, the N response in underlying biomass and N transport related characteristics was less than that for biomass, there were clear relationships, most importantly, lines that maintained biomass at reduced N maintained net N uptake with no change in size of the root relative to the shoot. The root uptake capacity for both [Formula: see text] and [Formula: see text] increased with reduced N. Transcript levels of putative [Formula: see text] and [Formula: see text] transporter genes in the root tissue of a subset of the genotypes revealed that predominately ZmNRT2 transcript levels responded to N treatments. The correlation between the ratio of transcripts of ZmNRT2.2 between the two N levels and a genotype's ability to maintain biomass with reduced N suggests a role for these transporters in enhancing NUpE. The observed variation in the ability to capture N at low N provides scope for both improving NUpE in maize and also to better understand the N uptake system in cereals.

Free oxygen radicals regulate plasma membrane Ca2+- and K+-permeable channels in plant root cells.

Free oxygen radicals are an irrefutable component of life, underlying important biochemical and physiological phenomena in animals. Here it is shown that free oxygen radicals activate plasma membrane Ca(2+)- and K(+)-permeable conductances in Arabidopsis root cell protoplasts, mediating Ca(2+) influx and K(+) efflux, respectively. Free oxygen radicals generate increases in cytosolic Ca(2+) mediated by a novel population of nonselective cation channels that differ in selectivity and pharmacology from those involved in toxic Na(+) influx. Analysis of the free oxygen radical-activated K(+) conductance showed its similarity to the Arabidopsis root K(+) outward rectifier. Significantly larger channel activation was found in cells responsible for perceiving environmental signals and undergoing elongation. Quenching root free oxygen radicals inhibited root elongation, confirming the role of radical-activated Ca(2+) influx in cell growth. Net free oxygen radical-stimulated Ca(2+) influx and K(+) efflux were observed in root cells of monocots, dicots, C3 and C4 plants, suggesting conserved mechanisms and functions. In conclusion, two functions for free oxygen radical cation channel activation are proposed: initialization/amplification of stress signals and control of cell elongation in root growth.

Arabidopsis thaliana root non-selective cation channels mediate calcium uptake and are involved in growth.

Calcium is a critical structural and regulatory nutrient in plants. However, mechanisms of its uptake by root cells are poorly understood. We have found that Ca2+ influx in Arabidopsis root epidermal protoplasts is mediated by voltage-independent rapidly activating Ca2+-permeable non-selective cation channels (NSCCs). NSCCs showed the following permeability (P) sequence: PCa (1.00) = PBa (0.93) > PZn (0.51), PCa/PNa = 0.19, PCa/PK = 0.14. They were inhibited by quinine, Gd3+, La3+ and the His modifier diethylpyrocarbonate, but not by the Ca2+ or K+ channel antagonists, verapamil and tetraethylammonium (TEA+). Single channel conductance measured in 20 mm external Ca2+ was 5.9 pS. Calcium-permeable NSCCs co-existed with hyperpolarisation-activated Ca2+ channels (HACCs), which activated 40-60 min after forming the whole-cell configuration. HACCs activated at voltages <-130 to -150 mV, showed slow activation kinetics and were regulated by cytosolic Ca2+ ([Ca2+]cyt). Using aequorin-expressing plants, a linear relationship between membrane potential (Vm) and resting [Ca2+]cyt was observed, indicating the involvement of NSCCs. Intact root 45Ca2+ influx was reduced by Gd3+ (NSCC blocker) but was verapamil and TEA+ insensitive. In the root elongation zone, both root net Ca2+ influx (measured by Ca2+-selective vibrating microelectrode) and NSCC activity were increased compared to the mature epidermis, suggesting the involvement of NSCC in growth. A Ca2+ acquisition system based on NSCC and HACC co-existence is proposed. In mature epidermal cells, NSCC-mediated Ca2+ influx dominates whereas in specialised root cells (root hairs and elongation zone cells) where elevated [Ca2+]cyt activates HACCs, HACC-mediated Ca2+ influx predominates.