Expressing the sunflower transcription factor HaHB11 in maize improves waterlogging and defoliation tolerance.

The sunflower (Helianthus annuus) transcription factor HaHB11 (H. annuus  Homeobox 11) belongs to the homeodomain-leucine zipper family and confers improved yield to maize (Zea mays) hybrids (HiII × B73) and lines. Here we report that transgenic maize lines expressing HaHB11 exhibited better performance under waterlogging, both in greenhouse and field trials carried out during three growth cycles. Transgenic plants had increased chlorophyll content, wider stems, more nodal roots, greater total aerial biomass, a higher harvest index, and increased plant grain yield. Under severe defoliation caused by a windstorm during flowering, transgenic genotypes were able to set more grains than controls. This response was confirmed in controlled defoliation assays. Hybrids generated by crossing B73 HaHB11 lines with the contrasting Mo17 lines were also tested in the field and exhibited the same beneficial traits as the parental lines, compared with their respective controls. Moreover, they were less penalized by stress than commercial hybrids. Waterlogging tolerance increased via improvement of the root system, including more xylem vessels, reduced tissue damage, less superoxide accumulation, and altered carbohydrate metabolism. Multivariate analyses corroborated the robustness of the differential traits observed. Furthermore, canopy spectral reflectance data, computing 29 vegetation indices associated with biomass, chlorophyll, and abiotic stress, helped to distinguish genotypes as well as their growing conditions. Altogether the results reported here indicate that this sunflower gene constitutes a suitable tool to improve maize plants for environments prone to waterlogging and/or wind defoliation.

An Interdisciplinary Approach to Study the Performance of Second-generation Genetically Modified Crops in Field Trials: A Case Study With Soybean and Wheat Carrying the Sunflower HaHB4 Transcription Factor.

Research, production, and use of genetically modified (GM) crops have split the world between supporters and opponents. Up to now, this technology has been limited to the control of weeds and pests, whereas the second generation of GM crops is expected to assist farmers in abiotic stress tolerance or improved nutritional features. Aiming to analyze this subject holistically, in this presentation we address an advanced technology for drought-tolerant GM crops, upscaling from molecular details obtained in the laboratory to an extensive network of field trials as well as the impact of the introduction of this innovation into the market. Sunflower has divergent transcription factors, which could be key actors in the drought response orchestrating several signal transduction pathways, generating an improved performance to deal with water deficit. One of such factors, HaHB4, belongs to the homeodomain-leucine zipper family and was first introduced in Arabidopsis. Transformed plants had improved tolerance to water deficits, through the inhibition of ethylene sensitivity and not by stomata closure. Wheat and soybean plants expressing the HaHB4 gene were obtained and cropped across a wide range of growing conditions exhibiting enhanced adaptation to drought-prone environments, the most important constraint affecting crop yield worldwide. The performance of wheat and soybean, however, differed slightly across mentioned environments; whereas the improved behavior of GM wheat respect to controls was less dependent on the temperature regime (cool or warm), differences between GM and wild-type soybeans were remarkably larger in warmer compared to cooler conditions. In both species, these GM crops are good candidates to become market products in the near future. In anticipation of consumers' and other stakeholders' interest, spectral analyses of field crops have been conducted to differentiate these GM crops from wild type and commercial cultivars. In this paper, the potential impact of the release of such market products is discussed, considering the perspectives of different stakeholders.

Field-grown transgenic wheat expressing the sunflower gene HaHB4 significantly outyields the wild type.

HaHB4 is a sunflower transcription factor belonging to the homeodomain-leucine zipper I family whose ectopic expression in Arabidopsis triggers drought tolerance. The use of PCR to clone the HaHB4 coding sequence for wheat transformation caused unprogrammed mutations producing subtle differences in its activation ability in yeast. Transgenic wheat plants carrying a mutated version of HaHB4 were tested in 37 field experiments. A selected transgenic line yielded 6% more (P<0.001) and had 9.4% larger water use efficiency (P<0.02) than its control across the evaluated environments. Differences in grain yield between cultivars were explained by the 8% improvement in grain number per square meter (P<0.0001), and were more pronounced in stress (16% benefit) than in non-stress conditions (3% benefit), reaching a maximum of 97% in one of the driest environments. Increased grain number per square meter of transgenic plants was accompanied by positive trends in spikelet numbers per spike, tillers per plant, and fertile florets per plant. The gene transcripts associated with abiotic stress showed that HaHB4's action was not dependent on the response triggered either by RD19 or by DREB1a, traditional candidates related to water deficit responses. HaHB4 enabled wheat to show some of the benefits of a species highly adapted to water scarcity, especially in marginal regions characterized by frequent droughts.

Reduced expression of selected FASCICLIN-LIKE ARABINOGALACTAN PROTEIN genes associates with the abortion of kernels in field crops of Zea mays (maize) and of Arabidopsis seeds.

Abortion of fertilized ovaries at the tip of the ear can generate significant yield losses in maize crops. To investigate the mechanisms involved in this process, 2 maize hybrids were grown in field crops at 2 sowing densities and under 3 irrigation regimes (well-watered control, drought before pollination, and drought during pollination), in all possible combinations. Samples of ear tips were taken 2-6 days after synchronous hand pollination and used for the analysis of gene expression and sugars. Glucose and fructose levels increased in kernels with high abortion risk. Several FASCICLIN-LIKE ARABINOGALACTAN PROTEIN (FLA) genes showed negative correlation with abortion. The expression of ZmFLA7 responded to drought only at the tip of the ear. The abundance of arabinogalactan protein (AGP) glycan epitopes decreased with drought and pharmacological treatments that reduce AGP activity enhanced the abortion of fertilized ovaries. Drought also reduced the expression of AthFLA9 in the siliques of Arabidopsis thaliana. Gain- and loss-of-function mutants of Arabidopsis showed a negative correlation between AthFLA9 and seed abortion. On the basis of gene expression patterns, pharmacological, and genetic evidence, we propose that stress-induced reductions in the expression of selected FLA genes enhance abortion of fertilized ovaries in maize and Arabidopsis.

Water deficit stress tolerance in maize conferred by expression of an isopentenyltransferase (IPT) gene driven by a stress- and maturation-induced promoter.

Senescence can be delayed in transgenic plants overexpressing the enzyme isopentenyltransferase (IPT) due to stress-induced increased levels of endogenous cytokinins. This trait leads to sustained photosynthetic activity and improved tolerance to abiotic stress. The aim of this study was to generate and characterize transgenic plants of maize (Zea mays L.) transformed with the IPT gene sequence under the regulation of SARK promoter (protein kinase receptor-associated senescence). Three independent transgenic events and their segregating null controls were evaluated in two watering regimes (WW: well watered; WD: water deficit) imposed for two weeks around anthesis. Our results show that the WD treatment induced IPT expression with the concomitant increase in cytokinin levels, which prolonged the persistence of total green leaf area, and maintained normal photosynthetic rate and stomatal conductance. These trends were accompanied by a minor decrease in number of grains per plant, individual grain weight and plant grain yield as compared to WW plants. Plants expressing the IPT gene under WD had PGR, anthesis and silking dates and biomass levels similar to WW plants. Our results demonstrate that expression of the IPT gene under the regulation of the SARK promoter helps improve productivity under WD conditions in C4 plants like maize.

Maize leaves turn away from neighbors.

In commercial crops, maize (Zea mays) plants are typically grown at a larger distance between rows (70 cm) than within the same row (16-23 cm). This rectangular arrangement creates a heterogeneous environment in which the plants receive higher red light (R) to far-red light (FR) ratios from the interrow spaces. In field crops, the hybrid Dekalb 696 (DK696) showed an increased proportion of leaves toward interrow spaces, whereas the experimental hybrid 980 (Exp980) retained random leaf orientation. Mirrors reflecting FR were placed close to isolated plants to simulate the presence of neighbors in the field. In addition, localized FR was applied to target leaves in a growth chamber. During their expansion, the leaves of DK696 turned away from the low R to FR ratio signals, whereas Exp980 leaves remained unaffected. On the contrary, tillering was reduced and plant height was increased by low R to FR ratios in Exp980 but not in DK696. Isolated plants preconditioned with low R/FR-simulating neighbors in a North-South row showed reduced mutual shading among leaves when the plants were actually grouped in North-South rows. These observations contradict the current view that phytochrome-mediated responses to low R/FR are a relic from wild conditions, detrimental for crop yield.