Changes in phenotype and gene expression under lead stress revealed key genetic responses to lead tolerance in Medicago sativa L.

Lead (Pb) is a serious heavy metal soil pollutant. It can be absorbed and accumulated by plant roots and impact plant growth. Medicago sativa L. (alfalfa) is a low-input forage and potential bioenergy crop, and improving its yield and quality has always been a focus of the alfalfa breeding industry. Little is known about the mechanism by which alfalfa responds to Pb stress at the molecular level. In this study, three alfalfa genotypes (a lead-resistant type (LR), a lead-sensitive type (LS) and an intermediate type (IN)) with contrasting abilities to resist lead were exposed to different durations of Pb treatment. Next-generation sequencing (NGS)-based RNA-seq technology was employed to characterize the root transcriptomes of three genotypes of alfalfa and identify differentially expressed genes (DEGs) during Pb stress. Genotypes LR and LS displayed different mechanisms of tolerance. In LR, the accumulation of more resistant substances was induced by the upregulation of sucrose synthase, glucan endo-1,3-beta-glucosidase, beta-amylase 3, probable trehalose-phosphate phosphatase J, 6-phosphofructo-2-kinase delta-1-pyrroline-5-carboxylate synthase (P5CS) and δ-ornithine aminotransferase (δ-OAT). In addition, flavin monooxygenase (YUCCA), 4-coumarate:CoA ligase-like protein (4CL), cinnamoyl-CoA reductase-like protein (CCR), ferulate 5-hydroxylase (F5H) and caffeic acid O-methyltransferase (COMT) were upregulated, leading to root development in a short time under Pb stress. Further study of the expression levels of metal transport-related genes, such as NRAMP (metal transporter), MATE (multidrug and toxin extrusion), HIPPs (heavy metal-associated isoprenylated plant proteins), MTP (metal tolerance protein), and ABC transporter, suggested that these genes were differentially expressed after lead treatment in the three alfalfa genotypes. Our research provides useful information for further studies on the molecular mechanism of Pb resistance in Medicago sativa L.