RNA-seq analysis of laser micro-dissected Arabidopsis thaliana leaf epidermis, mesophyll and vasculature defines tissue-specific transcriptional responses to multiple stress treatments

Abstract

Acclimation of plants to adverse environmental conditions requires the coordination of gene expression and signalling pathways between tissues and cell types. As the energy and carbon capturing organs, leaves are significantly affected by abiotic and biotic stresses. However, tissue- or cell type-specific analyses of stress responses have largely focussed on the Arabidopsis root. Here, we comparatively explore the transcriptomes of three leaf tissues (epidermis, mesophyll, vasculature) after induction of diverse stress pathways by chemical stimuli (antimycin A, 3-amino-1,2,4-triazole, methyl viologen, salicylic acid) and UV light in Arabidopsis. Profiling stimuli-dependent changes after treatments revealed an overall reduction in the tissue-specific expression of genes, with only a limited number gaining or changing their tissue-specificity. We find no evidence of a common stress response, with only a few genes responsive to two or more treatments in the analysed tissues. However, differentially expressed genes overlap across tissues for individual treatments. Further analyses provided evidence for an interaction of auxin and ethylene that mediates retrograde signalling during mitochondrial dysfunction specifically in the epidermis, and a gene regulatory network defined the hierarchy of interactions. Taken together, we generated an extensive reference data set and results enable the tailoring of the tissue-specific engineering of stress tolerant plants.

The single-cell stereo-seq reveals region-specific cell subtypes and transcriptome profiling in Arabidopsis leaves

“…WAX2, which has a metabolic function associated with both cuticle membrane and wax synthesis (Chen et al, 2003), was highly expressed in upper epidermal cells, whereas DIN6, which is involved in nitrogen metabolism repressed by light (Lam et al, 1998), was highly expressed in the lower epidermis (Figure 3D). Since guard cells were excluded from the epidermal cell in our dataset, this result indicates that epidermal pavement cells have photosynthetic capacity, consistent with the finding that pavement cells of Arabidopsis have chloroplasts (Barton et al, 2016;Berkowitz et al, 2021). For mesophyll cell subtypes, we observed that photosynthesis-related GO terms were significantly enriched in the palisade mesophyll, while stress-related GO terms were enriched in spongy mesophyll cells, which is suggestive of functional distinction as well (Figure S4D).…”

“…Based on the DEG analysis results, we captured cell-type-specific clusters of genes (Figures 3D and S4C; Table S3). The following Gene Ontology (GO) term analysis of DEGs for upper versus lower epidermal cells revealed that biological processes associated with photosynthesis (Berkowitz et al, 2021), cuticle development, and fatty acid biosynthesis were significantly enriched in upper epidermal cells, while biological terms related to immune response, response to absence of light, and regulation of cell death were significantly enriched in lower epidermal cells (Figure 3E). WAX2, which has a metabolic function associated with both cuticle membrane and wax synthesis (Chen et al, 2003), was highly expressed in upper epidermal cells, whereas DIN6, which is involved in nitrogen metabolism repressed by light (Lam et al, 1998), was highly expressed in the lower epidermis (Figure 3D).…”

“…Leaves are the main photosynthetic organs and are also involved in responses to biotic and abiotic stresses (Maugarny-Cale `s and Laufs, 2018). Different cell types in plant leaves exhibit various molecular signatures and perform specific functions (Berkowitz et al, 2021). Cell-type-specific characterizations are essential for a better understanding of the developmental mechanisms of leaves and their responses to environmental stimuli.…”

“…Embedded in the mesophyll cells are the vascular cells, which are responsible for substance transmission and exchange with other types of cells (Svozil et al, 2015;Tsukaya, 2013). Since individual cell types have specialized functions during leaf development and response to environmental stimuli (Berkowitz et al, 2021), tools for systematic research are needed to understand how distinct types of cells are regulated and how they communicate with each other to build the complex functional network in plant leaves.…”