Christine Granier

Plant aerial development is well known to be affected by day length in terms of the timing and developmental stage of floral transition. Arabidopsis thaliana is a ‘long day’ plant in which the time to flower is delayed by short days and leaf number is increased. The aim of the work presented here was to determine the effects of different day lengths on individual leaf area expansion. The effect of flower emergence per se on the regulation of leaf expansion was also tested in this study. Care was taken to ensure that day length was the only source of micro-meteorological variation. The dynamics of individual leaf expansion were analysed in Ler and Col-0 plants grown under five day lengths in five independent experiments. Responses at cellular level were analysed in Ler plants grown under various day lengths and treatments to alter the onset of flowering. When the same leaf position was compared, the final leaf area and both the relative and absolute rates of leaf expansion were decreased by short days, whereas the duration of leaf expansion was increased. Epidermal cell number and cell area were also altered by day-length treatments and some of these responses could be mimicked by manipulating the date of flowering. Both the dynamics and cellular bases of leaf development are altered by differences in day length even when visible phenotypes are absent. To some extent, cell area and its response to day length are controlled by whole plant control mechanisms associated with the onset of flowering.

Genetic variability in the plasticity of leaf area expansion in response to water deficit has been reported in Arabidopsis thaliana. Here, the objective was to identify the underlying dynamic and cellular processes involved in this variability. Twenty‐five accessions were subjected to identical soil water deficit treatments. In all accessions, the plasticity of leaf production was low compared with that of individual leaf expansion. A subset of accessions was selected for further dissection of individual leaf expansion into its underlying variables: the rate and duration of leaf expansion and epidermal cell number and area. In all accessions, water deficit had opposite effects on the rate and duration of leaf expansion. The accumulation of these effects was reflected in changes in final leaf area. At the cellular level, moderate water deficits had opposite effects on cell number and cell size, but more severe ones reduced both variables. The importance of these opposing effects is highlighted by the behaviour of the accession An‐1, for which the compensation between the decrease in leaf expansion rate and the increase in the duration of expansion is total. This dynamic plasticity in response to water deficit is not detectable when only final measurements are done.

Leaf area expansion is affected by environmental conditions because of differences in cell number and/or cell size. Increases in the DNA content (ploidy) of a cell by endoreduplication are related to its size. The aim of this work was to determine how cell ploidy interacts with the regulation of cell size and with leaf area expansion. The approach used was to grow Arabidopsis thaliana plants performing increased or decreased rounds of endoreduplication under shading and water deficit. The shading and water deficit treatments reduced final leaf area and cell number; however, cell area was increased and decreased, respectively. These differences in cell size were unrelated to alterations of the endocycle, which was reduced by these treatments. The genetic modification of the extent of endoreduplication altered leaf growth responses to shading and water deficit. An increase in the extent of endoreduplication in a leaf rendered it more sensitive to the shade treatment but less sensitive to water deficit conditions. The link between the control of whole organ and individual cell expansion under different environmental conditions was demonstrated by the correlation between the plasticity of cell size and the changes in the duration of leaf expansion.

• Background and Aims It is well known that plant aerial development is affected by light intensity in terms of the date of flowering, the length of stems and petioles, and the final individual leaf area. The aim of the work presented here was to analyse how shade-induced changes in leaf development occur on a dynamic basis from the whole rosette level to that of the cells. • Methods Care was taken to ensure that light intensity was the only source of micro-meteorological variation in the study. The dynamics of leaf production, rosette expansion, individual leaf area expansion and epidermal cell expansion were analysed in Arabidopsis thaliana plants grown under two light intensities in three independent experiments. • Key Results The total area of rosette leaves was reduced by the shading treatment. Both the number of leaves produced and their individual leaf areas were reduced. The reduction in leaf number was associated with a reduction in leaf initiation rate and the duration of the phase of leaf production. The reduction in individual leaf area was associated with a reduction in leaf expansion rate and an increase in the duration of leaf expansion. The changes in leaf expansion dynamics were accompanied by a decrease in epidermal cell number which was partly compensated for by an increase in epidermal cell area. Overall, the whole rosette leaf expansion rate was reduced by shading, whereas the total duration of rosette leaf expansion was unaffected. This was mainly due to the accumulation of the increases in the durations of expansion of each individual leaf which was associated with an increase in cell expansion. • Conclusions The dynamic analysis presented here reveals a new shade-adaptative response mediated via the control of area expansion at the cell, organ and whole plant levels.

• The high‐throughput phenotypic analysis of Arabidopsis thaliana collections requires methodological progress and automation. Methods to impose stable and reproducible soil water deficits are presented and were used to analyse plant responses to water stress. • Several potential complications and methodological difficulties were identified, including the spatial and temporal variability of micrometeorological conditions within a growth chamber, the difference in soil water depletion rates between accessions and the differences in developmental stage of accessions the same time after sowing. Solutions were found. • Nine accessions were grown in four experiments in a rigorously controlled growth‐chamber equipped with an automated system to control soil water content and take pictures of individual plants. One accession, An1, was unaffected by water deficit in terms of leaf number, leaf area, root growth and transpiration rate per unit leaf area. • Methods developed here will help identify quantitative trait loci and genes involved in plant tolerance to water deficit.

Leaf development is affected by both internal (genetic) and external (environmental) regulatory factors. The aim of this work was to investigate how leaf growth variables are related to one another in a range of environments. The leaf growth variables of wild‐type Arabidopsis thaliana and leaf development mutants (ang4, ron2‐1, elo1, elo2 and elo4) were studied under different incident light treatments (light and shade). The leaves studied were altered in various leaf development variables, such as the duration of expansion, relative and absolute expansion rates, epidermal cell size, epidermal cell number and initiation rate. Final leaf area was correlated to maximal absolute leaf expansion rate and cell number, but not to duration of leaf expansion or cell size. These relationships were common to all studied genotypes and light conditions, suggesting that leaf size is determined early in development. In addition, the early variables involved in leaf development were correlated to one another, and initial relative expansion rate was negatively correlated to the duration of expansion. These relationships between the leaf development variables were used to construct a conceptual model of leaf size control.

In crop species, the impact of temperature on plant development is classically modelled using thermal time. We examined whether this method could be used in a non‐crop species, Arabidopsis thaliana, to analyse the response to temperature of leaf initiation rate and of the development of two leaves of the rosette. The results confirmed the large plant‐to‐plant variability in the studied isogenic line of the Columbia ecotype: 100‐fold differences in leaf area among plants sown on the same date were commonly observed at a given date. These differences disappeared in mature leaves, suggesting that they were due to a variability in plant developmental stage. The whole population could therefore be represented by any group of synchronous plants labelled at the two‐leaf stage and followed during their development. Leaf initiation rate, duration of leaf expansion and maximal relative leaf expansion rate varied considerably among experiments performed at different temperatures (from 6 to 26 °C) but they were linearly related to temperature in the range 6–26 °C, with a common x‐intercept of 3 °C. Expressing time in thermal time with a threshold temperature of 3 °C unified the time courses of leaf initiation and of individual leaf development for plants grown at different temperatures and experimental conditions. The two leaves studied (leaf 2 and leaf 6) had a two‐phase development, with an exponential phase followed by a phase with decreasing relative elongation rate. Both phases had constant durations for a given leaf position if expressed in thermal time. Changes in temperature caused changes in both the rate of development and in the expansion rate which mutually compensated such that they had no consequence on leaf area at a given thermal time. The resulting model of leaf development was applied to ten experiments carried out in a glasshouse or in a growth chamber, with plants grown in soil or hydroponically. Because it predicts accurately the stage of development and the relative expansion rate of any leaf of the rosette, this model facilitates precise planning of sampling procedures and the comparison of treatments in growth analyses.

We have investigated the spatial distributions of cell division rate, p34^cdc2 kinase activity, and amount of p34^cdc2a in maize (Zea mays) leaves grown at contrasting temperatures and soil water conditions. An original method for calculating cell division rate in all leaf tissues is proposed. In all studied conditions, cell division rate was stable and maximum in the first 2 cm beyond the leaf insertion point, declined afterward, and reached zero at 7 cm from the insertion point. The spatial distribution of p34^cdc2 kinase activity, expressed on a per cell basis, followed the same pattern. In contrast, the amount of p34^cdc2a was maximum in the first centimeter of the leaf, declined afterward, but remained at 20% of maximum in more distal zones with a near-zero cell division rate. A mild water deficit caused a reduction in cell division rate and p34^cdc2 kinase activity by approximately 45% in all leaf zones, but did not affect the amount of p34^cdc2a. Growth temperature affected to the same extent cell division rate and p34^cdc2 kinase activity, but only if p34^cdc2 kinase activity was assayed at growth temperature, and not if a standard temperature was used in all assays. A common linear relationship between cell division rate and p34^cdc2 kinase activity applied to all causes of changes in cell division rate, i.e. cell aging, water deficit, or changes in temperature. It is shown that temperature has two distinct and additive effects on p34^cdc2 kinase activity; first, an effect on the rate of the reaction, and second, an effect on the amount of p34^cdc2a.