Deciphering glyoxalase gene families and their coordinated regulation with the ascorbate-glutathione cycle during salinity stress in Artemisia annua.

Abstract

Salinity stress disrupts plant metabolism by causing excessive buildup of methylglyoxal and reactive oxygen species, both of which demand efficient detoxification and redox balance. In this study, we combined genome-wide in silico analyses with molecular, biochemical, metabolic, and physiological approaches to explore how the glyoxalase system contributes to salinity stress adaptation in Artemisia annua. We identified 20 AaGLYI, 13 AaGLYII, and 7 AaGLYIII genes, all characterised by conserved domain structures, distinct motif patterns, and stress-responsive cis-regulatory elements. Phylogenetic analysis highlighted both evolutionary conservation and functional diversification of these gene families across plant species. Expression profiling of Arabidopsis orthologues, together with qRT-PCR validation of selected AaGLYs (AaGLYI18, AaGLYI20, AaGLYII05, AaGLYII11, AaGLYIII01, AaGLYIII02), revealed strong, time-dependent induction under salinity stress. This was coupled with enhanced glyoxalase enzyme activities and a transient spike in MG levels followed by efficient detoxification. Concurrent activation of the ascorbate-glutathione (AsA-GSH) cycle maintained redox equilibrium during early stress phases, though prolonged exposure imposed redox constraints. Physiological assessments indicated early reductions in stomatal conductance and photochemical efficiency, partially compensated by increased non-photochemical quenching and preserved PSII integrity. Interestingly, glyoxalase activation under stress paralleled elevated expression of key artemisinin biosynthetic genes and higher in vivo artemisinin accumulation, suggesting a functional link between MG detoxification, redox regulation, and secondary metabolism during salinity stress in A. annua.

Graphical abstract: Schematic representation demonstrating cellular and physiological reactions of Artemisia annua to salt stress. Salinity-induced ionic and osmotic instability results in increased generation of reactive oxygen species and methylglyoxal, leading to oxidative and carbonyl stress. MG is detoxified by the glyoxalase route by the sequential actions of GLYI and GLYII in a glutathione (GSH)-dependent manner, while GLYIII facilitates glutathione-independent detoxification under prolonged stress conditions. The activation of the glyoxalase system is precisely synchronised with the ascorbate-glutathione (AsA-GSH) cycle, which includes APX, GR, MDHAR, and DHAR, to maintain redox homeostasis and mitigate oxidative damage. Metabolites generated from MG integrate into central carbon metabolism, facilitating metabolic reprogramming during stress. Simultaneously, physiological modifications such as proline buildup, increased non-photochemical quenching, and alterations in gas exchange and photosynthetic parameters facilitate the preservation of photosystem II integrity and adaptability to stress. The graphical abstract collectively emphasises the coordinated regulation of detoxification pathways, redox metabolism, and physiological responses that contribute to salinity tolerance. ROS, reactive oxygen species; MG, methylglyoxal; GLYI/II/III, glyoxalase I/II/III; GSH, reduced glutathione; GSSG, oxidised glutathione; AsA, ascorbate; APX, ascorbate peroxidase; GR, glutathione reductase; MDHAR, monodehydroascorbate reductase; DHAR, dehydroascorbate reductase; NPQ, non-photochemical quenching; PSII, photosystem II.

Supplementary information: The online version contains supplementary material available at 10.1007/s12298-026-01751-1.