A 5-methyltryptophan resistant mutant of rice has an altered regulation of anthranilate synthase gene expression

The rice mutant, TR-1, was previously selected for resistance to growth inhibition by the tryptophan (Trp) analog 5-methyltryptophan (5MT). This mutant has an elevated Trp level. Since anthranilate synthase (AS) catalyzes the first step of Trp biosynthesis and is normally subject to feedback regulation by Trp, alteration of AS causes elevated Trp levels. In this study, we performed a physiological, biochemical and molecular characterization of the 5MT resistant mutant TR-68, a derivative of TR-1. The extracts of TR-68 exhibited 6-fold higher AS activity than that of the wild-type under standard conditions. However, the AS activity of TR-68 was as sensitive to feedback inhibition by Trp as that of the wild-type. We determined the nucleotide sequences of a part of rice AS gene (OASA1) corresponding to the site of mutation in 5MT resistant mutant in Arabidopsis and to the site of naturally feedback insensitive AS in Nicotiana tabacum and Ruta graveolens, however, there were no differences in these regions between TR-68 and wild-type. In the wild-type, the expression of two AS genes (OASA1 and OASA2) was inhibited simultaneously by 5MT and the addition of Trp partly suppressed this inhibition. In contrast, 5MT did not affect the expression of AS genes in TR-68. These data indicate that the 5MT resistance of TR-68 may be caused by mutation of a regulator of the expression of the AS genes.