Abstract
As one of the pioneer crops widely planted in saline-alkaline areas, Gossypium provides daily necessities, including natural fiber, vegetable proteins, and edible oils. However, cotton fiber yield and quality are highly influenced by salt stress. Therefore, elucidating the molecular mechanisms of cotton in response to salinity stress is importance to breed new cultivars with high tolerance. In this study, we first developed a method for single-cell RNA-seq based on isolating protoplast from cotton root tips; then, we studied the impact of salinity stress on gene expression profiling and their dynamic changes using the developed high-efficiency method for protoplast dissociation suitable for single-cell RNA-seq. A total of 3391 and 2826 differentially expressed genes (DEGs) were identified in salt-treated samples before and after protoplast dissociation, respectively, which were enriched into several molecular components, including response to stimulus, response to stress, and cellular macromolecule metabolic process by gene ontology (GO) analysis. Plant hormone signal transduction, phenylpropanoid biosynthesis, and MAPK signaling pathway were found to be enriched via Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Twenty-two and nine salinity-responsive DEGs participated in plant hormone signaling and MAPK signaling in roots, before and after protoplast dissociation, respectively; six upregulated DEGs were involved in ABA signaling transduction, namely, Ga04G2111, Ga07G0142, Ga09G2061, Ga10G0262, Ga01G0063, and Ga08G1915 which indicates their potential functions on plants adapting to salt stress. Additionally, 384 and 257 transcription factors (TFs) were differentially expressed in salt-stress roots before and after protoplast dissociation, respectively, of which significantly up-regulated TFs mainly belonged to the AP2/ERF-ERF family, which implied their potential roles responding to salt stress. These results not only provide novel insights to reveal the regulatory networks in plant’s root response to salt stress, but also lay the solid foundation for further exploration on cellular heterogeneity by single-cell transcriptome sequencing.
Highlights
During each stage of the life cycle, the growth and development of plants are uninterruptedly affected by various biotic and abiotic stresses
We report the highly-efficient method for protoplast dissociation from lateral roots of cotton under salt stress
RNA-seq was conducted on the cotton root tips of 5-day-old seedlings and its dissociated protoplasts under salt and control conditions, and 22 and 9 common differentially expressed genes (DEGs) responsible for salt stress were found to participate in plant hormone signaling and MAP kinase (MAPK) signaling between the samples before and after dissociation
Summary
During each stage of the life cycle, the growth and development of plants are uninterruptedly affected by various biotic and abiotic stresses. Drought and salt stresses are the leading contributors for yield loss. Plenty of attempts had been conducted to investigate the biological processes during plant response to salt stress; results indicated that, in the initial stages, plants underwent water loss, which inhibited leaf growth; subsequently, osmotic effect could be quickly observed, which gradually inhibited cell division and cell growth, and led to stomatal closure [2]. Plants undergo osmotic stress and ion stress, of which the former inhibits the growth of young leaves, while the latter promotes the senescence of mature leaves. Salt stress could promote the transport of abscisic acid (ABA) to guard cells, resulting in reduced stomatal conductance and, further reducing photosynthesis and photoinhibition to resist stress, which inhibits plants growth and leads to leaf senescence. Salt stress induced oxidative stress by triggering high accumulation of reactive oxygen species in the aerobic process of cells, which plays an important role in maintaining homeostasis for protecting plants against abiotic stress, including salinity stress [3]
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