Transcriptome analysis of Medicago lupulina seedlings leaves treated by high calcium provides insights into calcium oxalate formation

Abstract

Calcium oxalate (CaOx) is a common biomineral found in the plant kingdom. Crystals of CaOx occur in different plant tissues, such as leaves and stems. However, little is known about the biosynthesis of CaOx in oxalate-accumulating plants. Moreover, the literature on genes related to CaOx formation under high-calcium environment is scarce. In the present study, the physiological parameters and the transcript profiles of Medicago lupulina leaves treated with 0.1 and 25 mM Ca2+ were analyzed to study the genes involved in the biosynthesis of CaOx. We demonstrated that exposure to high external calcium concentration induced H2O2 production, ascorbic acid degradation, and CaOx accumulation in M. lupulina leaves. Moreover, we identified 1715 differentially expressed genes (DEGs) (1322 up-regulated and 393 down-regulated genes) in leaves treated with 25 mM Ca2+ compared with the leaves treated with 0.1 mM Ca2+. We further demonstrated the involvement of DEGs in oxalic acid production, calcium transport, and calcium buffering. These results revealed that a high calcium promoted oxalic acid biosynthesis by inducing the expression of NADPH oxidase and ascorbate oxidase genes. In addition, several genes encoding cyclic nucleotide-gated channel, Ca2+-ATPase, H+/Ca2+ exchangers, and calcium-binding proteins were found to be differentially expressed and involved in calcium transport and calcium buffering. Our transcriptome analyses provide a comprehensive insight into the biosynthesis of CaOx in oxalate-accumulating plants.