Identification of Ice Plant (Mesembryanthemum crystallinum L.) MicroRNAs Using RNA-Seq and Their Putative Roles in High Salinity Responses in Seedlings

Abstract

The halophyte Mesembryanthemum crystallinum (common or crystalline ice plant) is a useful model for studying molecular mechanisms of salt tolerance. The morphology, physiology, metabolism, and gene expression of ice plant have been studied and large-scale analyses of gene expression profiling have drawn an outline of salt tolerance in ice plant. A rapid root growth to a sudden increase in salinity was observed in ice plant seedlings. Using a fluorescent dye to detect Na+, we found that ice plant roots respond to an increased flux of Na+ by either secreting or storing Na+ in specialized cells. High-throughput sequencing was used to identify small RNA profiles in 3-day-old seedlings treated with or without 200 mM NaCl. In total, 135 conserved miRNAs belonging to 21 families were found. The hairpin precursor of 19 conserved mcr-miRNAs and 12 novel mcr-miRNAs were identified. After 6 h of salt stress, the expression of most mcr-miRNAs showed decreased relative abundance, whereas the expression of their corresponding target genes showed increased mRNA relative abundance. The cognate target genes are involved in a broad range of biological processes: transcription factors that regulate growth and development, enzymes that catalyze miRNA biogenesis for the most conserved mcr-miRNA, and proteins that are involved in ion homeostasis and drought-stress responses for some novel mcr-miRNAs. Analyses of the functions of target genes revealed that cellular processes, including growth and development, metabolism, and ion transport activity are likely to be enhanced in roots under salt stress. The expression of eleven conserved miRNAs and two novel miRNAs were correlated reciprocally with predicted targets within hours after salt stress exposure. Several conserved miRNAs have been known to regulate root elongation, root apical meristem activity, and lateral root formation. Based upon the expression pattern of miRNA and target genes in combination with the observation of Na+ distribution, ice plant likely responds to increased salinity by using Na+ as an osmoticum for cell expansion and guard cell opening. Excessive Na+ could either be secreted through the root epidermis or stored in specialized leaf epidermal cells. These responses are regulated in part at the miRNA-mediated post-transcriptional level.

Making Plants Break a Sweat: the Structure, Function, and Evolution of Plant Salt Glands

“…The critical role played by salt bladders in M. crystallinum for development and survival under high NaCl was further confirmed by the creation of growth impaired mutant plants without bladder cells (Agarie et al, 2007). The remarkable salt and drought tolerance capacity exhibited by M. crystallinum has also led to its use as a model halophyte in multiple gene expression studies using ESTs and RNAseq from bulk tissues to discover gene regulatory mechanisms related to salt tolerance (Bohnert and Cushman, 2000; Cushman et al, 2008; Tsukagoshi et al, 2015; Chiang et al, 2016). Additionally, the recent cell specific targeted transcriptome, proteome, and metabolome analyses have reported the type of genes, proteins, and metabolites expressed specifically in salt glands in M. crystallinum (Barkla et al, 2012; Barkla and Vera-Estrella, 2015; Oh et al, 2015).…”

Physiological Changes in Mesembryanthemum crystallinum During the C3 to CAM Transition Induced by Salt Stress

“…Differences in gene expression, protein, and metabolite levels, as well as phenotypic changes have been studied in C 3 and CAM plants (Aragón et al, 2012;Cosentino et al, 2013;Abraham et al, 2016;Chiang et al, 2016). For example, weekly morphological and physiological changes in micropropagated pineapple under CAM-inducing conditions were characterized (Aragón et al, 2012).…”

Model approaches to advance crassulacean acid metabolism system integration

“…The availability of CAM genomes has provided a springboard for analysis of orthologues and shared transcriptional control elements (Ming et al, 2015;Yang et al, 2017;Wai et al, 2019), whereas emerging comparative 'omics' analysis of various CAM species has also contributed to a more in-depth understanding of regulatory networks (Abraham et al, 2016;Chiang et al, 2016;Zhang et al, 2016;Yin et al, 2018;Heyduk et al, 2019). Comparative transcriptomic studies have provided insights into the evolutionary trajectory of CAM and suggest that transcriptional regulation is primarily associated with specific expression profiles of key CAM genes.…”