LAUSR

All seed plants share a common basic vegetative body composed of three organs: root, stem, and leaf. The functionality and plasticity of each of these specialized organs are conferred by the complex organization of different tissues whose function depends, in turn, on the interplay of distinct cell populations (Tsukaya, 2013). Leaves are the primary photosynthetic organs and harbor a plethora of cell types organized in three functionally distinct tissues: epidermis, mesophyll, and vasculature. Whereas the epidermis functions as a shield to protect the internal tissues from the environment, the mesophyll represents the major photosynthetic tissue. Finally, the vascular tissue, composed of xylem and phloem and embedded by the bundle sheath within the mesophyll, transports water and products of absorption and assimilation (Kalve et al., 2014). The establishment of these specialized leaf tissues occurs gradually during leaf development from a population of undifferentiated cells forming the leaf primordium. In addition to developmental cues, leaf development is very sensitive to environmental stimuli, and developing leaves reprogram their growth and their metabolism in response to different stresses (Claeys and Inzé, 2013). Despite the recent progress in the elucidation of the genetic program controlling leaf growth and its environmentresponsive plasticity, our understanding of the molecular mechanisms coordinating growth within and among different leaf tissues remains limited. For instance, studies involving whole-transcriptome analyses have focused on leaf cell types that can be easily isolated, such as trichomes or stomatal guard cells (Jin et al., 2013; Yang and Ye, 2013), but studies reporting transcriptomic profiling of multiple leaf tissues simultaneously at single-cell resolution have not been reported. Recent methodological advances, including the development of high-throughput single-cell RNA-sequencing (scRNA-seq), now allow the elucidation of the transcriptomic landscape of the different cell types that build complex plant tissues, laying the foundations for revealing unknown aspects of the molecular mechanisms controlling plant development. In this issue of Plant Physiology, Berrı́o et al. (2021) report the generation of a single-cell transcriptomic atlas of developing Arabidopsis (Arabidopsis thaliana) leaves, which was used to characterize the genetic basis of the identity and functions of the distinct leaf cell types. Protoplasting of leaf cells was performed in actively growing leaves to include both proliferating and differentiating cells from plants under well-watered and mild drought conditions to capture specific adaptative responses during leaf development. Droplet-based scRNA-seq performed on 1,887 high-quality protoplasts, 1,330 obtained from well-watered leaves and 557 from stressed leaves, and preliminary analysis based on global expression profiles and unsupervised clustering identified a total of 11 distinct cell populations (Figure 1A). Differential expression (DE) analysis between samples from the two analyzed environmental conditions depicted differences in their expression profiles, confirming that mild drought stress was effective in altering the leaf transcriptome. Once different cell type populations were identified, the authors first aimed at delineating which of these populations formed the three main leaf tissues: epidermis, mesophyll, and vasculature. Accordingly, the expression of marker genes for each of these tissues was analyzed. For instance, genes belonging to the GLYCOSYL-PHOSPHATIDYLINOSITOL-ANCHORED LIPID PROTEIN TRANSFER (LPTG) and the 3-KETOACYL-CoASYNTHASE (KCS) families are involved in cuticular wax production and are specifically expressed in the epidermis (Kim et al., 2013). Based on their expression specifically observed in cells from clusters #4, #7, and part of cluster #9, the populations of epidermal cells were defined (Figure 1B). Similar analyses led to the identification of cell populations from N ew s an d V ie w s