A D-lactate dehydrogenase from rice is involved in conferring tolerance to multiple abiotic stresses by maintaining cellular homeostasis

Abstract

D-lactate dehydrogenase (D-LDH) converts D-lactate (the end product of glyoxalase system) to pyruvate and thereby completes the detoxification process of methylglyoxal. D-LDH detoxifies and diverts the stress induced toxic metabolites, MG and D-lactate, towards energy production and thus, protects the cell from their deteriorating effects. In this study, a D-LDH enzyme from rice (OsD-LDH2, encoded by Os07g08950.1) was characterized for its role in abiotic stress tolerance. For this, a combination of in silico, molecular, genetic and biochemical approaches was used. The kinetic analysis revealed OsD-LDH2 to be the most efficient D-LDH enzyme in comparison to D-LDHs from other plant species. Heterologous overexpression of OsD-LDH2 provides tolerance against multiple abiotic stresses in E. coli, yeast and plant system. The analysis of D-LDH mutant and OsD-LDH2 overexpressing transgenic plants uncovered the crucial role of D-LDH in mitigation of abiotic stresses. OsD-LDH2 overexpressing plants maintained lower level of ROS and other toxic metabolites along with better functioning of antioxidant system. This is the first report on correlation of D-LDH with multiple abiotic stress tolerance. Overall, OsD-LDH2 emerged as a promising candidate which can open a new direction for engineering stress tolerant crop varieties by maintaining their growth and yield in unfavorable conditions. Abiotic stress is defined as the impact of non-living factors on the living organisms in a specific environment. Abiotic stress leads to a series of morphological, physiological, biochemical and molecular changes and it has been predicted that environmental factors may limit crop production by approximately 70% 1. So, to overcome the toxic effects of abiotic stress, a network of events is triggered that starts with stress perception and ends with the expression of a large array of genes. To engineer stress tolerance, various stress responsive genes, proteins and transcription factors have been studied for their role in conferring stress tolerance 2-7. The present study is aimed at identifying the role of enzyme D-lactate dehydrogenase (D-LDH) in abiotic stress mitigation. D-lactate dehydrogenase (D-LDH) catalyzes the conversion of D-lactate to pyruvate. D-LDH belongs to the class of D-lactate ferricytochrome c oxidoreductase (EC 1.1.2.4) or D-lactate NAD oxidoreductase (EC 1.1.1.28) depending on the cofactor used, either cytochrome c or NAD. D-LDH enzymes have earlier been identified in prokaryotes and lower fungi where they are involved in anaerobic energy metabolism 8-16. In S. cerevisiae and K. lactis, D-lactate cytochrome c oxidoreductase was found important for utilization of D-lactate 13,17,18. But, extensive information about functioning of D-LDH in higher organisms or plants is yet to be explored. The major source of D-lactate accumulation in plants is the glyoxalase pathway for methylglyoxal (MG) detoxification. MG is a cytotoxic metabolite produced as a by-product of various metabolic reactions and is accumulated during stress cond...

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