Soil salinity is a main abiotic stress in agriculture worldwide. The Na⁺/H⁺ antiporters (NHXs) play pivotal roles in intracellular Na⁺ excretion and vacuolar Na⁺ compartmentalization, which are important for plant salt stress resistance (SSR). However, few systematic analyses of NHXs has been reported in allotetraploid rapeseed so far. Here, a total of 18 full-length NHX homologs, representing seven subgroups (NHX1-NHX8 without NHX5), were identified in the rapeseed genome (A_nA_nC_nC_n). Number variations of BnaNHXs might indicate their significantly differential roles in the regulation of rapeseed SSR. BnaNHXs were phylogenetically divided into three evolutionary clades, and the members in the same subgroups had similar physiochemical characteristics, gene/protein structures, and conserved Na⁺ transport motifs. Darwin´s evolutionary pressure analysis suggested that BnaNHXs suffered from strong purifying selection. The cis-element analysis revealed the differential transcriptional regulation of NHXs between the model Arabidopsis and B. napus. Differential expression of BnaNHXs under salt stress, different nitrogen forms (ammonium and nitrate), and low phosphate indicated their potential involvement in the regulation of rapeseed SSR. Global landscapes of BnaNHXs will give an integrated understanding of their family evolution and molecular features, which will provide elite gene resources for the genetic improvement of plant SSR through regulating the NHX-mediated Na⁺ transport.

Cadmium (Cd) is a highly toxic heavy metal that quickly enters cereals, such as wheat, via the roots and is translocated to the shoots and grains, posing high risks to human health. However, the vast and complex genome of allohexaploid wheat makes it challenging to understand Cd resistance and accumulation. Herein, a Cd-resistant cultivar (‘ZM1860’) and a Cd-sensitive cultivar (‘ZM32’) of wheat, selected from 442 accessions, exhibited significantly differential plant Cd resistance and grain Cd accumulation. An integrated comparative analysis of the morpho-physiological traits, ionomic and phytohormone profiles, genomic variations, transcriptomic landscapes, and gene functionality was performed to identify the mechanisms underlying differential Cd resistance and accumulation among the wheat genotypes. Under Cd toxicity, ‘ZM1860’ outperformed ‘ZM32’, which showed more severe leaf chlorosis, weaker root architecture, higher reactive oxygen species accumulation, and disordered phytohormone homeostasis. Ionomics showed that ‘ZM32’ had a higher root-to-shoot translocation coefficient of Cd and accumulated more Cd in the grains than ‘ZM1860’. Whole-genome re-sequencing (WGS) and transcriptome sequencing identified numerous DNA variants and differentially expressed genes involved in abiotic stress responses and ion transport between the genotypes. Combined ionomics, transcriptomics, and functional gene analysis identified the plasma membrane-localized TaHMA2b-7A as a crucial Cd exporter regulating long-distance Cd translocation in wheat. WGS- and PCR-based analysis of sequence polymorphisms revealed a 25-bp InDel site in the promoter region, probably responsible for the differential expression of TaHMA2b-7A between the wheat genotypes. The multiomics approach eased identifying a core transporter involved in long-distance Cd translocation in wheat, and it may provide an elite genetic resource for improving plant Cd resistance and grain Cd accumulation in wheat and other cereal crops.

Background: Nitrogen (N), referred to as a “life element”, is a macronutrient essential for optimal plant growth and yield production. Amino acid (AA) permease (AAP) genes play pivotal roles in root import, long-distance translocation, remobilization of organic amide-N from source organs to sinks, and other environmental stress responses. However, few systematic analyses of AAPs have been reported in Brassica napus so far. Results: In this study, we identified a total of 34 full-length AAP genes representing eight subgroups (AAP1–8) from the allotetraploid rapeseed genome (A_nA_nC_nC_n, 2n = 4x = 38). Great differences in the homolog number among the BnaAAP subgroups might indicate their significant differential roles in the growth and development of rapeseed plants. The BnaAAPs were phylogenetically divided into three evolutionary clades, and the members in the same subgroups had similar physiochemical characteristics, gene/protein structures, and conserved AA transport motifs. Darwin’s evolutionary analysis suggested that BnaAAPs were subjected to strong purifying selection pressure. Cis-element analysis showed potential differential transcriptional regulation of AAPs between the model Arabidopsis and B. napus. Differential expression of BnaAAPs under nitrate limitation, ammonium excess, phosphate shortage, boron deficiency, cadmium toxicity, and salt stress conditions indicated their potential involvement in diverse nutrient stress responses. Conclusions: The genome-wide identification of BnaAAPs will provide a comprehensive insight into their family evolution and AAP-mediated AA transport under diverse abiotic stresses. The molecular characterization of core AAPs can provide elite gene resources and contribute to the genetic improvement of crop stress resistance through the modulation of AA transport.

Improving crop nitrogen (N) limitation adaptation (NLA) is a core approach to enhance N use efficiency (NUE) and reduce N fertilizer application. Rapeseed has a high demand for N nutrients for optimal plant growth and seed production, but it exhibits low NUE. Epigenetic modification, such as DNA methylation and modification from small RNAs, is key to plant adaptive responses to various stresses. However, epigenetic regulatory mechanisms underlying NLA and NUE remain elusive in allotetraploid B. napus. In this study, we identified overaccumulated carbohydrate, and improved primary and lateral roots in rapeseed plants under N limitation, which resulted in decreased plant nitrate concentrations, enhanced root-to-shoot N translocation, and increased NUE. Transcriptomics and RT-qPCR assays revealed that N limitation induced the expression of NRT1.1, NRT1.5, NRT1.7, NRT2.1/NAR2.1, and Gln1;1, and repressed the transcriptional levels of CLCa, NRT1.8, and NIA1. High-resolution whole genome bisulfite sequencing characterized 5094 differentially methylated genes involving ubiquitin-mediated proteolysis, N recycling, and phytohormone metabolism under N limitation. Hypermethylation/hypomethylation in promoter regions or gene bodies of some key N-metabolism genes might be involved in their transcriptional regulation by N limitation. Genome-wide miRNA sequencing identified 224 N limitation-responsive differentially expressed miRNAs regulating leaf development, amino acid metabolism, and plant hormone signal transduction. Furthermore, degradome sequencing and RT-qPCR assays revealed the miR827-NLA pathway regulating limited N-induced leaf senescence as well as the miR171-SCL6 and miR160-ARF17 pathways regulating root growth under N deficiency. Our study provides a comprehensive insight into the epigenetic regulatory mechanisms underlying rapeseed NLA, and it will be helpful for genetic engineering of NUE in crop species through epigenetic modification of some N metabolism-associated genes.

Background: Plants worldwide are often stressed by low Fe availability around the world, especially in aerobic soils. Therefore, the plant growth, seed yield, and quality of crop species are severely inhibited under Fe deficiency. Fe metabolism in plants is controlled by a series of complex transport, storage, and regulatory mechanisms in cells. Allohexaploid wheat (Triticum aestivum L.) is a staple upland crop species that is highly sensitive to low Fe stresses. Although some studies have been previously conducted on the responses of wheat plants to Fe deficiency, the key mechanisms underlying adaptive responses are still unclear in wheat due to its large and complex genome. Results: Transmission electron microscopy showed that the chloroplast structure was severely damaged under Fe deficiency. Paraffin sectioning revealed that the division rates of meristematic cells were reduced, and the sizes of elongated cells were diminished. ICP-MS-assisted ionmics analysis showed that low-Fe stress significantly limited the absorption of nutrients, including N, P, K, Ca, Mg, Fe, Mn, Cu, Zn, and B nutrients. High-throughput transcriptome sequencing identified 378 and 2,619 genome-wide differentially expressed genes (DEGs) were identified in the shoots and roots between high-Fe and low-Fe conditions, respectively. These DEGs were mainly involved in the Fe chelator biosynthesis, ion transport, photosynthesis, amino acid metabolism, and protein synthesis. Gene coexpression network diagrams indicated that TaIRT1b-4A, TaNAS2-6D, TaNAS1a-6A, TaNAS1-6B, and TaNAAT1b-1D might function as key regulators in the adaptive responses of wheat plants to Fe deficiency. Conclusions: These results might help us fully understand the morpho-physiological and molecular responses of wheat plants to low-Fe stress, and provide elite genetic resources for the genetic modification of efficient Fe use.