Soybean Root Nodule cDNA Encoding Glutathione Reductase

GSH is widely distributed in organisms and has diverse functions in protein synthesis, sulfur storage, and protection against a variety of stresses (Meister and Anderson, 1983). GR (EC 1.6.4.2) is a flavoprotein that catalyzes the reduction of GSSG to reduced GSH using NADPH as the reducing cofactor. It is a key enzyme in the GSH-ascorbate cycle, which provides protection against oxidative stress, particularly in photosynthetic tissues of plants (Foyer and Halliwell, 1976). Tobacco transformed with the gor gene, encoding GR in Escherichia coli, showed mixed results in resistance to oxidative stress (Aono et al., 1991; Foyer et al., 1991). Recently, a cDNA clone encoding GR was isolated from a cDNA library of pea leaves (Creissen et al., 1991). An N-terminal leader sequence encoded by the pea cDNA was consistent with targeting of the gene product to the chloroplast. In root nodules, enzymes of the GSH-ascorbate cycle have been suggested to function in peroxide scavenging and protection against other oxidative stresses (Dalton et al., 1986). In soybean, GR activity was much higher in nodules than in uninfected roots and was enhanced in nodules exposed to higher than normal p 0 2 (Dalton et al., 1991). GR activity also was about 4-fold higher in effective nodules than in ineffective nodules, with a range of factors controlling effectiveness (Dalton et al., 1993). Soybean (Glycine max) nodules contain both homo-GSH as well as GSH, and the combined concentration of thiol tripeptides in nodules was calculated to be about 1 m~ (Dalton et al., 1993). Recent studies examining the role of GSH in relation to pathogen defense responses (Wingate et al., 1988; Edwards et al., 1991) have implications for GSH function during nodule development as well. A full-length cDNA encoding soybean nodule GR (Table I) was isolated serendipitously in screening a soybean nodule cDNA library with antibodies against nodule allantoinase. The deduced amino acid sequence of the cDNA was found to share a high degree of homology to sequences for GR in other organisms and was 90% similar and 83% identical to pea GR (Creissen et al., 1991). The soybean GR sequence also had an N-terminal leader sequence characteristic of a chloroplast transit peptide, which would target the gene product to nonphotosynthetic plastids in root nodules. South’