Response of sulfate transporters, thiol metabolism, and antioxidant defense system was studied in roots of two lentil (Lens culinaris Medik.) genotypes grown in arsenic (10, 25, and 40 μM AsV)-supplemented nutrient solution, and significant changes compared to control (0 μM AsV) were observed mainly at 25 and 40 μM. In L 414, high glutathione (GSH) redox (0.8-0.9) was maintained with elevated thiol synthesis, powered by transcriptional up-regulation of LcSultr1;1 and LcSultr1;2 sulfate transporters and significant induction of LcSAT1;1 and LcSAT1;2 (serine acetyltransferase), OAS-TL (O-acetylserine(thiol)-lyase), γ-ECS (γ-glutamylcysteine synthetase), and PCS (phytochelatin synthase) genes predominantly within 12-24 h of As exposure at 25 μM and within 6-12 h at 40 μM. This thiolic potency in L 414 roots was effectively complemented by up-regulation of gene expressions and consequent enhanced activities of superoxide dismutase, ascorbate peroxidase (APX), dehydroascorbate reductase, glutathione reductase (GR), and glutathione-S-transferase (GST) isoforms at 25 and 40 μMAs, efficiently scavenging excess reactive oxygen species to prevent onset of As-induced oxidative stress and consequent inhibition of root growth in L 414. In contrast, down-regulation of vital sulfate-uptake transporters as well as entire thiol-metabolizing system and considerably low APX, GST, and GR expressions in DPL 59 not only resulted in reduced GSH redox but also led to over-accumulation of H2O2. This triggered membrane lipid peroxidations as the marks of As-induced oxidative damage. Results indicated coordinated response of thiol-metabolism and antioxidant defense in conferring As-tolerance in lentil, and GSH is the key point in this cascade. © 2013 Springer-Verlag Wien.