Best practices for the execution, analysis, and data storage of plant single-cell/nucleus transcriptomics.

Single-cell and single-nucleus RNA-sequencing technologies capture the expression of plant genes at an unprecedented resolution. Therefore, these technologies are gaining traction in plant molecular and developmental biology for elucidating the transcriptional changes across cell types in a specific tissue or organ, upon treatments, in response to biotic and abiotic stresses, or between genotypes. Despite the rapidly accelerating use of these technologies, collective and standardized experimental and analytical procedures to support the acquisition of high-quality data sets are still missing. In this commentary, we discuss common challenges associated with the use of single-cell transcriptomics in plants and propose general guidelines to improve reproducibility, quality, comparability, and interpretation and to make the data readily available to the community in this fast-developing field of research.

Regulation of hair cell and stomatal size by a hair cell-specific peroxidase in the grass Brachypodium distachyon.

The leaf epidermis is the outermost cell layer forming the interface between plants and the atmosphere that must both provide a robust barrier against (a)biotic stressors and facilitate carbon dioxide uptake and leaf transpiration.1 To achieve these opposing requirements, the plant epidermis developed a wide range of specialized cell types such as stomata and hair cells. Although factors forming these individual cell types are known,2,3,4,5 it is poorly understood how their number and size are coordinated. Here, we identified a role for BdPRX76/BdPOX, a class III peroxidase, in regulating hair cell and stomatal size in the model grass Brachypodium distachyon. In bdpox mutants, prickle hair cells were smaller and stomata were longer. Because stomatal density remained unchanged, the negative correlation between stomatal size and density was disrupted in bdpox and resulted in higher stomatal conductance and lower intrinsic water-use efficiency. BdPOX was exclusively expressed in hair cells, suggesting that BdPOX cell-autonomously promotes hair cell size and indirectly restricts stomatal length. Cell-wall autofluorescence and lignin stainings indicated a role for BdPOX in the lignification or crosslinking of related phenolic compounds at the hair cell base. Ectopic expression of BdPOX in the stomatal lineage increased phenolic autofluorescence in guard cell (GC) walls and restricted stomatal elongation in bdpox. Together, we highlight a developmental interplay between hair cells and stomata that optimizes epidermal functionality. We propose that cell-type-specific changes disrupt this interplay and lead to compensatory developmental defects in other epidermal cell types.