A New Phenotyping Pipeline Reveals Three Types of Lateral Roots and a Random Branching Pattern in Two Cereals.

Recent progress in root phenotyping has focused mainly on increasing throughput for genetic studies, while identifying root developmental patterns has been comparatively underexplored. We introduce a new phenotyping pipeline for producing high-quality spatiotemporal root system development data and identifying developmental patterns within these data. The SmartRoot image-analysis system and temporal and spatial statistical models were applied to two cereals, pearl millet (Pennisetum glaucum) and maize (Zea mays). Semi-Markov switching linear models were used to cluster lateral roots based on their growth rate profiles. These models revealed three types of lateral roots with similar characteristics in both species. The first type corresponds to fast and accelerating roots, the second to rapidly arrested roots, and the third to an intermediate type where roots cease elongation after a few days. These types of lateral roots were retrieved in different proportions in a maize mutant affected in auxin signaling, while the first most vigorous type was absent in maize plants exposed to severe shading. Moreover, the classification of growth rate profiles was mirrored by a ranking of anatomical traits in pearl millet. Potential dependencies in the succession of lateral root types along the primary root were then analyzed using variable-order Markov chains. The lateral root type was not influenced by the shootward neighbor root type or by the distance from this root. This random branching pattern of primary roots was remarkably conserved, despite the high variability of root systems in both species. Our phenotyping pipeline opens the door to exploring the genetic variability of lateral root developmental patterns.

Phenotyping oilseed rape growth-related traits and their responses to water deficit: the disturbing pot size effect.

Following the recent development of high-throughput phenotyping platforms for plant research, the number of individual plants grown together in a same experiment has raised, sometimes at the expense of pot size. However, root restriction in excessively small pots affects plant growth and carbon partitioning, and may interact with other stresses targeted in these experiments. In work reported here, we investigated the interactive effects of pot size and soil water deficit on multiple growth-related traits from the cellular to the whole-plant scale in oilseed rape (Brassica napus L.). The effects of pot size on responses to water deficit and allometric relationships revealed strong, multilevel interactions between pot size and watering regime. Notably, water deficit increased the root:shoot ratio in large pots, but not in small pots. At the cellular scale, water deficit decreased epidermal leaf cell area in large pots, but not in small pots. These results were consistent with changes in the level of endoreduplication factor in leaf cells. Our study illustrates the disturbing interaction of pot size with water deficit and raises the need to carefully consider this factor in the frame of the current development of high-throughput phenotyping experiments.

Heterosis and inbreeding depression of epigenetic Arabidopsis hybrids.

We have addressed the possible epigenetic contribution to heterosis using epigenetic inbred lines (epiRILs) with varying levels and distributions of DNA methylation. One line consistently displayed parent-of-origin heterosis for growth-related traits. Genome-wide transcription profiling followed by a candidate gene approach revealed 33 genes with altered regulation in crosses of this line that could contribute to the observed heterosis. Although none of the candidate genes could explain hybrid vigour, we detected intriguing, hybrid-specific transcriptional regulation of the RPP5 gene, encoding a growth suppressor. RPP5 displayed intermediate transcript levels in heterotic hybrids; surprisingly however, with global loss of fitness of their F2 progeny, we observed striking under-representation of the hybrid-like intermediate levels. Thus, in addition to genetic factors contributing to heterosis, our results strongly suggest that epigenetic diversity and epigenetic regulation of transcription play a role in hybrid vigour and inbreeding depression, and also in the absence of parental genetic diversity.

Phenotyping the kinematics of leaf development in flowering plants: recommendations and pitfalls.

Leaves of flowering plants are produced from the shoot apical meristem at regular intervals and they grow according to a developmental program that is determined by both genetic and environmental factors. Detailed frameworks for multiscale dynamic analyses of leaf growth have been developed in order to identify and interpret phenotypic differences caused by either genetic or environmental variations. They revealed that leaf growth dynamics are non-linearly and nonhomogeneously distributed over the lamina, in the leaf tissues and cells. The analysis of the variability in leaf growth, and its underlying processes, has recently gained momentum with the development of automated phenotyping platforms that use various technologies to record growth at different scales and at high throughput. These modern tools are likely to accelerate the characterization of gene function and the processes that underlie the control of shoot development. Combined with powerful statistical analyses, trends have emerged that may have been overlooked in low throughput analyses. However, in many examples, the increase in throughput allowed by automated platforms has led to a decrease in the spatial and/or temporal resolution of growth analyses. Concrete examples presented here indicate that simplification of the dynamic leaf system, without consideration of its spatial and temporal context, can lead to important misinterpretations of the growth phenotype.

Disentangling the intertwined genetic bases of root and shoot growth in Arabidopsis.

Root growth and architecture are major components of plant nutrient and water use efficiencies and these traits are the matter of extensive genetic analysis in several crop species. Because root growth relies on exported assimilate from the shoot, and changes in assimilate supply are known to alter root architecture, we hypothesized (i) that the genetic bases of root growth could be intertwined with the genetic bases of shoot growth and (ii) that the link could be either positive, with alleles favouring shoot growth also favouring root growth, or negative, because of competition for assimilates. We tested these hypotheses using a quantitative genetics approach in the model species Arabidopsis thaliana and the Bay-0 × Shahdara recombinant inbred lines population. In accordance with our hypothesis, root and shoot growth traits were strongly correlated and most root growth quantitative trait loci (QTLs) colocalized with shoot growth QTLs with positive alleles originating from either the same or the opposite parent. In order to identify regions that could be responsible for root growth independently of the shoot, we generated new variables either based on root to shoot ratios, residuals of root to shoot correlations or coordinates of principal component analysis. These variables showed high heritability allowing genetic analysis. They essentially all yielded similar results pointing towards two regions involved in the root--shoot balance. Using Heterogeneous Inbred Families (a kind of near-isogenic lines), we validated part of the QTLs present in these two regions for different traits. Our study thus highlights the difficulty of disentangling intertwined genetic bases of root and shoot growth and shows that this difficulty can be overcome by using simple statistical tools.