ORIGINAL PAPER Molecular Cloning and Characterization of γ-Glutamyl Cysteine Synthetase (VrγECS) from Roots of Vigna radiata (L.) Wilczek Under Progressive Drought Stress and Recovery Debashree Sengupta & Golla Ramesh & Shalini Mudalkar & Koppolu Raja Rajesh Kumar & Pulugurtha Bharadwaja Kirti & Attipalli R. Reddy Published online: 12 January 2012 # Springer-Verlag 2012 Abstract Glutathione is an essential redox buffer and an antioxidant in majority of higher plants, imparting tolerance against abiotic stress. The rate-limiting enzyme, gamma- glutamyl cysteine synthetase (γECS), plays an important role in regulation of glutathione biosynthesis under adverse environmental conditions including drought. To understand the role of γECS in an economically important food legume, Vigna radiata (L.) Wilczek, under progressive drought stress, we cloned and derived the full-length cDNA sequence and denoted it as VrγECS. Real-time PCR analysis of VrγECS in the roots of V. radiata, during progressive drought stress and recovery, indicated a stable expression pattern of the gene. However, the VrγECS enzyme activity altered differentially during varying water-deficit con- ditions and recovery period, reflecting the existence of some post-transcriptional or post-translational regulatory system for the enzyme. Linear regression analysis between H 2 O 2 and lipid peroxidation as well as H 2 O 2 and VrγECS enzyme activity during drought stress and recovery demonstrates the delicate inter-relationships and putative regulatory mecha- nisms operating in the root system under adverse conditions. The present study could contribute towards understanding the complex regulation of γECS in glutathione biosynthesis in an important food legume under drought stress. Keywords Drought . Gamma-glutamyl cysteine synthetase . Glutathione . Mungbean . Regulatory mechanisms Abbreviations DAS Days after onset of stress treatment DAR Days after re-watering GSH Glutathione PPFD Photosynthetic photon flux density RACE Rapid amplification of cDNA ends ROS Reactive oxygen species γECS Gamma-glutamyl cysteine synthetase Introduction In plants, the tripeptide glutathione (γ-L-glutamyl-L-cysteinyl- glycine) is well known for its multipurpose role as an efficient redox buffer (Foyer and Noctor 2005), xenobiotic detoxifier (Cummins et al. 2011), sulphur assimilator (Kopriva 2006), heavy metal detoxifier (Cobbett 2000), cell signalling compo- nent (Foyer and Noctor 2005), antioxidant (Mittova et al. 2003) and a key component of various plant developmental processes (Vernoux et al. 2000). Owing to its diversified importance, the biosynthesis, recycling, degradation and regulation of glutathione during adverse environmental conditions have been actively worked upon (Foyer et al. 1995; Xiang et al. 2001; Ball et al. 2004; Chen et al. 2004; Aravind and Prasad 2005). Glutathione is synthesized in two ATP-dependent steps involving two different enzymes: γ-glutamyl cysteine synthetase or γECS (EC 6.3.2.2) and Electronic supplementary material The online version of this article (doi:10.1007/s11105-011-0398-y) contains supplementary material, which is available to authorized users. D. Sengupta : S. Mudalkar : K. R. R. Kumar : P. B. Kirti : A. R. Reddy (*) Department of Plant Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, India e-mail: arrsl@uohyd.ernet.in G. Ramesh Division of Haematology and Oncology, School of Medicine, University of Pennsylvania, Philadelphia 19104, USA Plant Mol Biol Rep (2012) 30:894–903 DOI 10.1007/s11105-011-0398-y