Drought induces distinct growth response, protection, and recovery mechanisms in the maize leaf growth zone

Drought is the most important crop yield-limiting factor, and detailed knowledge of its impact on plant growth regulation is crucial. The maize (Zea mays) leaf growth zone offers unique possibilities for studying the spatiotemporal regulation of developmental processes by transcriptional analyses and methods that require more material, such as metabolite and enzyme activity measurements. By means of a kinematic analysis, we show that drought inhibits maize leaf growth by inhibiting cell division in the meristem and cell expansion in the elongation zone. Through a microarray study, we observed the down- regulation of 32 of the 54 cell cycle genes, providing a basis for the inhibited cell division. We also found evidence for an up- regulation of the photosynthetic machinery and the antioxidant and redox systems. This was confirmed by increased chlorophyll content in mature cells and increased activity of antioxidant enzymes and metabolite levels across the growth zone, respectively. We demonstrate the functional significance of the identified transcriptional reprogramming by showing that increasing the antioxidant capacity in the proliferation zone, by overexpression of the Arabidopsis (Arabidopsis thaliana) iron-superoxide dismutase gene, increases leaf growth rate by stimulating cell division. We also show that the increased photosynthetic capacity leads to enhanced photosynthesis upon rewatering, facilitating the often-observed growth compensation.Fil: Avramova, Viktoriya. Universiteit Antwerp; BélgicaFil: Abdelgawad, Hamada. University of Beni-Suef; EgiptoFil: Zhang, Zhengfeng. Central China Normal University; ChinaFil: Fotschki, Bartosz. Institute of Animal Reproduction and Food Research; PoloniaFil: Casadevall, Romina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Centro de Estudios Fotosintéticos y Bioquímicos. Universidad Nacional de Rosario. Facultad de Ciencias Bioquímicas y Farmacéuticas. Centro de Estudios Fotosintéticos y Bioquímicos; ArgentinaFil: Vergauwen, Lucia. Universiteit Antwerp; BélgicaFil: Knapen, Dries. Universiteit Antwerp; BélgicaFil: Taleisnik, Edith. Instituto Nacional de Tecnología Agropecuaria. Centro de Investigaciones Agropecuarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Guisez, Yves. Universiteit Antwerp; BélgicaFil: Asard, Han. Universiteit Antwerp; BélgicaFil: Beemster, Gerrit T. S.. Universiteit Antwerp; Bélgic

Kinematic analysis of cell division and expansion : quantifying the cellular basis of growth and sampling developmental zones in **Zea mays** leaves

Growth analyses are often used in plant science to investigate contrasting genotypes and the effect of environmental conditions. The cellular aspect of these analyses is of crucial importance, because growth is driven by cell division and cell elongation. Kinematic analysis represents a methodology to quantify these two processes. Moreover, this technique is easy to use in non-specialized laboratories. Here, we present a protocol for performing a kinematic analysis in monocotyledonous maize (Zea mays) leaves. Two aspects are presented: (1) the quantification of cell division and expansion parameters, and (2) the determination of the location of the developmental zones. This could serve as a basis for sampling design and/or could be useful for data interpretation of biochemical and molecular measurements with high spatial resolution in the leaf growth zone. The growth zone of maize leaves is harvested during steady-state growth. Individual leaves are used for meristem length determination using a DAPI stain and cell-length profiles using DIC microscopy. The protocol is suited for emerged monocotyledonous leaves harvested during steady-state growth, with growth zones spanning at least several centimeters. To improve the understanding of plant growth regulation, data on growth and molecular studies must be combined. Therefore, an important advantage of kinematic analysis is the possibility to correlate changes at the molecular level to well-defined stages of cellular development. Furthermore, it allows for a more focused sampling of specified developmental stages, which is useful in case of limited budget or time

Generating Plants with Improved Water Use Efficiency

To improve sustainability of agriculture, high yielding crop varieties with improved water use efficiency (WUE) are needed. Despite the feasibility of assessing WUE using different measurement techniques, breeding for WUE and high yield is a major challenge. Factors influencing the trait under field conditions are complex, including different scenarios of water availability. Plants with C3 photosynthesis are able to moderately increase WUE by restricting transpiration, resulting in higher intrinsic WUE (iWUE) at the leaf level. However, reduced CO2 uptake negatively influences photosynthesis and possibly growth and yield as well. The negative correlation of growth and WUE could be partly disconnected in model plant species with implications for crops. In this paper, we discuss recent insights obtained for Arabidopsis thaliana (L.) and the potential to translate the findings to C3 and C4 crops. Our data on Zea mays (L.) lines subjected to progressive drought show that there is potential for improvements in WUE of the maize line B73 at the whole plant level (WUEplant). However, changes in iWUE of B73 and Arabidopsis reduced the assimilation rate relatively more in maize. The trade-off observed in the C4 crop possibly limits the effectiveness of approaches aimed at improving iWUE but not necessarily efforts to improve WUEplant

Plant responses to drought stress in future climate CO2: A multi-level analysis

AbstractFuture climate scenarios indicate increasing CO2 levels in combination with more frequent drought spells. Recent studies suggest that elevated CO2 reduces the effect of drought stress, but the mechanisms underlying the reduced stress effects remain unclear. Therefore we investigated the growth, physiological, biochemical and genome-wide transcriptional responses of Arabidopsis thaliana to mild (MD) and severe drought (SD) under ambient (aCO2) and elevated CO2 (eCO2), 360 and 620ppm respectively. Kinematic analysis showed that drought reduced leaf growth by inhibiting cell division and expansion. High CO2 and drought showed opposite and interactive effects on growth. Where, eCO2 reduced the impact of MD on growth by limiting the effect on both cell number and size, and the impact of SD by reducing the effect on cell size only. Genome-wide transcriptional analysis showed that growth, photosynthesis and defense processes are more strongly affected by SD as compared to MD. These results were mirrored at the growth, physiological and biochemical levels, where SD inhibited photosynthesis and stomatal conductance and induced defense related parameters. Accumulation of antioxidants and osmolytes were not sufficient to completely recover the SD damage impact. In contrast, in SD conditions in particular, eCO2 significantly mitigated the detrimental effects of drought stress. The transcriptome and metabolic analyses did not implicate the antioxidant defense metabolism in protective effect of eCO2, with down regulation or no significant impact on most of measured antioxidant parameters. On the other hand, we found that eCO2 alleviated photorespiration; a stress-induced H2O2 generation process thus, the relaxation in antioxidant system could be a consequence of less oxidative pressure. Further induction of osmolytes was also observed and that could be associated with improved leaf relative water content under the drought and eCO2. We therefore concluded that elevated CO2 mitigates the effects of drought by maintaining plant turgor and reduced hydrogen peroxide production

Screening for drought tolerance of maize hybrids by multi-scale analysis of root and shoot traits at the seedling stage

We studied the drought response of eight commercial hybrid maize lines with contrasting drought sensitivity together with the reference inbred line B73 using a non-invasive platform for root and shoot phenotyping and a kinematics approach to quantify cell level responses in the leaf. Drought treatments strongly reduced leaf growth parameters including projected leaf area, elongation rate, final length and width of the fourth and fifth leaf. Physiological measurements including water use efficiency, chlorophyll fluorescence and photosynthesis were also significantly affected. By performing a kinematic analysis, we show that leaf growth reduction in response to drought is mainly due to a decrease in cell division rate, whereas a marked reduction in cell expansion rate is compensated by increased duration of cell expansion. Detailed analysis of root growth in rhizotrons under drought conditions revealed a strong reduction in total root length as well as rooting depth and width. This was reflected by corresponding decreases in fresh and dry weight of the root system. We show that phenotypic differences between lines differing in geographic origin (African vs. European) and in drought tolerance under field conditions can already be identified at the seedling stage by measurements of total root length and shoot dry weight of the plants. Moreover, we propose a list of candidate traits that could potentially serve as traits for future screening strategies

High antioxidant activity facilitates maintenance of cell division in leaves of drought tolerant maize hybrids

We studied the impact of drought on growth regulation in leaves of 13 maize varieties with different drought sensitivity and geographic origins (Western Europe, Egypt, South Africa) and the inbred line B73. Combining kinematic analysis of the maize leaf growth zone with biochemical measurements at a high spatial resolution allowed us to examine the correlation between the regulation of the cellular processes cell division and elongation, and the molecular redox-regulation in response to drought. Moreover, we demonstrated differences in the response of the maize lines to mild and severe levels of water deficit. Kinematic analysis indicated that drought tolerant lines experienced less impact on leaf elongation rate due to a smaller reduction of cell production, which, in turn, was due to a smaller decrease of meristem size and number of cells in the leaf meristem. Clear differences in growth responses between the groups of lines with different geographic origin were observed in response to drought. The difference in drought tolerance between the Egyptian hybrids was significantly larger than between the European and South-African hybrids. Through biochemical analyses, we investigated whether antioxidant activity in the growth zone, contributes to the drought sensitivity differences. We used a hierarchical clustering to visualize the patterns of lipid peroxidation, H(2)O(2) and antioxidant concentrations, and enzyme activities throughout the growth zone, in response to stress. The results showed that the lines with different geographic region used different molecular strategies to cope with the stress, with the Egyptian hybrids responding more at the metabolite level and African and the European hybrids at the enzyme level. However, drought tolerance correlated with both, higher antioxidant levels throughout the growth zone and higher activities of the redox-regulating enzymes CAT, POX, APX, and GR specifically in leaf meristems. These findings provide evidence for a link between antioxidant regulation in the leaf meristem, cell division, and drought tolerance