INDOLE-3-ACETIC-ACID HOMEOSTASIS IN TRANSGENIC TOBACCO PLANTS EXPRESSING THE AGROBACTERIUM-RHIZOGENES ROLB GENE

The roIB gene of the plant pathogen Agrobacterium rhizogenes has an im portant role in the establishment of hairy root disease in infected pl ant tissues. When expressed as a single gene in transgenic plants the RoIB protein gives rise to effects indicative of increased auxin activ ity. It has been reported that the RoIB product is a beta-glucosidase and proposed that the physiological and developmental alterations in t ransgenic plants expressing the roIB gene are the result of this enzym e hydrolysing bound auxins, in particular (indole-3-acetyl)-beta-D-glu coside (IAGluc), and thereby bringing about an increase in the intrace llular concentration of indole-3-acetic acid (IAA). Using tobacco plan ts as a test system, this proposal has been investigated in detail. Co mparisons have been made between the RoIB phenotype and that of iaaM/i aaH transformed plants overproducing IAA. In addition, the levels of I AA and IAA amide and IAA ester conjugates were determined in wild-type and transgenic 35S-roIB tobacco plants and metabolic studies were car ried out with [C-13(6)]IAA, [2'-C-14]IAA, [C-14]IAGluc, [5-H-3]-2-O-(i ndole-3-acetyl)myo-inositol and [C-14]indole-3-acetylaspartic acid. Th e data obtained demonstrate that expression of the roIB encoded protei n in transgenic tobacco does not produce a phenotype that resembles th at of IAA over-producing plants, does not alter the size of the free I AA pool, has no significant effect on the rate of IAA metabolism, and, by implication, appears not to influence the overall rate of IAA bios ynthesis. Furthermore, the in vivo hydrolysis of IAGluc, and that of t he other IAA conjugates that were tested, is not affected. On the basi s of these findings, it is concluded that the RoIB phenotype is not th e consequence of an increase in the size of the free IAA pool mediated by an enhanced rate of hydrolysis of IAA conjugates.