Single‐cell RNA‐seq describes the transcriptome landscape and identifies critical transcription factors in the leaf blade of the allotetraploid peanut (Arachis hypogaea L.)

Abstract

Summary Single‐cell RNA‐seq (scRNA‐seq) has been highlighted as a powerful tool for the description of human cell transcriptome, but the technology has not been broadly applied in plant cells. Herein, we describe the successful development of a robust protoplast cell isolation system in the peanut leaf. A total of 6,815 single cells were divided into eight cell clusters based on reported marker genes by applying scRNA‐seq. Further, a pseudo‐time analysis was used to describe the developmental trajectory and interaction network of transcription factors (TFs) of distinct cell types during leaf growth. The trajectory enabled re‐investigation of the primordium‐driven development processes of the mesophyll and epidermis. These results suggest that palisade cells likely differentiate into spongy cells, while the epidermal cells originated earlier than the primordium. Subsequently, the developed method integrated multiple technologies to efficiently validate the scRNA‐seq result in a homogenous cell population. The expression levels of several TFs were strongly correlated with epidermal ontogeny in accordance with obtained scRNA‐seq values. Additionally, peanut AHL23 (AT‐HOOK MOTIF NUCLEAR LOCALIZED PROTEIN 23), which is localized in nucleus, promoted leaf growth when ectopically expressed in Arabidopsis by modulating the phytohormone pathway. Together, our study displays that application of scRNA‐seq can provide new hypotheses regarding cell differentiation in the leaf blade of Arachis hypogaea. We believe that this approach will enable significant advances in the functional study of leaf blade cells in the allotetraploid peanut and other plant species.

Single‐cell transcriptome atlas reveals developmental trajectories and a novel metabolic pathway of catechin esters in tea leaves

“…Based on the specific unique molecular identification of single‐cell, scRNA‐seq enable monitoring of intracellular transcriptional activity at the single‐cell level (Seyfferth et al ., 2021 ). Currently, scRNA‐seq of several plant species, such as Arabidopsis, strawberry, rice and maize, peanuts, and one woody plant populus were reported (Bai et al ., 2022 ; Chen et al ., 2021 ; Kim et al ., 2021 ; Li et al ., 2021 ; Liu et al ., 2020 , 2021 ; Marand et al ., 2021 ; Xie et al ., 2021 ; Zhang et al ., 2021a , b ). Through these studies, a database of cell‐specific markers and a single‐cell atlas of various tissues and organs from different species has been established, providing new insights into organ development, cell differentiation and cell division (Jin et al ., 2021 ).…”

“…Moreover, the relatively long growth cycles and lack of effective transgenic technologies to validate the spatial and temporal expression profiles of genes have resulted in few cell type‐specific marker genes being detected in the non‐model woody plants. This exacerbates the difficulty of cell type identification and largely explains why most scRNA‐seq analyses were done in model plants (Jin et al ., 2021 ; Liu et al ., 2021 ; Seyfferth et al ., 2021 ; Zhang et al ., 2021a ). Leaf is one of the principal vegetative organs of vascular plants for photosynthesis and transpiration.…”

Developmental Analysis of Compound Leaf Development in Arachis hypogaea

“…Leaves are vital to plants for their ability to process photosynthesis. The diversity of leaf shapes has evolved to adapt to the environment, by maximizing the ability to absorb sunlight ( Liu et al, 2021 ). Most legume species have compound leaf structures.…”

Protoplast Dissociation and Transcriptome Analysis Provides Insights to Salt Stress Response in Cotton

“…Therefore, protoplasts have been widely applied in the transient genetic transformation and single-cell RNA-seq (sc-RNA-seq) research, of which the former is devoted to conduct subcellular localization and interaction protein identification on targeted genes, and the later focus on revealing the cellular heterogeneity at the single cell level. Currently, protoplast dissociation has been successfully obtained from different tissues of Arabidopsis [ 16 , 17 , 18 , 19 , 20 , 21 , 22 , 23 ], maize [ 24 ], peanut [ 25 ], rice [ 26 ] and poplar [ 27 ], most of which were subsequently utilized to perform scRNA-seq for cell type identification. To date, numerous protoplast-based studies in cotton have been done on leaves, while few studies focused on the optimization method of protoplast isolation and its transcriptome analysis under salt stress at the single cell level in roots.…”