Characterization and Safener Induction of Multiple Glutathione S-Transferases in Three Genetic Lines of Rice

Abstract

Multiple glutathione S-transferase (GST; EC 2.5.1.18) activity peaks were characterized by fast protein liquid chromatography (FPLC) in three genetic lines of rice (Oryza sativa L.). Root-applied pretilachlor (0.15 μM) inhibited root growth of 5-day-old etiolated seedlings by 67% in Teqing (indica type), 54% in Koshihikari (japonica type), and 34% in Lemont (indica x japonica type). The safener fenclorim (1 μM) prevented the injury caused by pretilachlor in all genotypes. GST activity from 5-day-old untreated or fenclorim-treated seedlings of the three genetic lines was assayed using 1-chloro-2,4-dinitrobenzene (CDNB), radiolabeled pretilachlor, and radiolabeled fenclorim as substrates. GST activity with each substrate was greater in roots than in shoots of the three genotypes. Treatment with fenclorim increased GST activity in roots and shoots by 20 to 90%. GST activity in all genotypes was greater with CDNB, followed by pretilachlor, and then fenclorim. GST(CDNB) and GST(pretilachlor) activities were greater in Lemont, followed by Koshihikari, and then Teqing. In contrast, GST(fenclorim) activity was greater in Koshihikari and least in Lemont. FPLC elution patterns (MonoQ column) of partially purified GST extracts from untreated and fenclorim-treated seedlings of the three rice genotypes revealed the presence of multiple GST activities. Teqing shoots had six GST activity peaks with fenclorim. Four of them were also active with CDNB and pretilachlor. Koshihikari shoots had six GST activity peaks with pretilachlor. Five of them were also active with fenclorim, but only three were active with CDNB. Lemont shoots exhibited three GST activity peaks with fenclorim, five with CDNB, and seven with pretilachlor. Multiple GST activity peaks using CDNB and pretilachlor as substrates were also present in the roots of Teqing, Koshihikari, and Lemont seedlings. Fenclorim increased the activity of GST peaks conjugating pretilachlor and CDNB more than those active with fenclorim. Induction of GST peaks was more evident in Lemont, followed by Koshihikari, and then Teqing. The differential patterns of GST peaks active toward pretilachlor show that rice japonica lines may be more tolerant to pretilachlor than indica lines.