Acetosyringone, a phenolic inducer of the virulence (vir) genes of Agrobacterium tumefaciens, inhibited the growth of the nopaline-type strains T37 and C58 incubated under acidic conditions. In the course of a 6-day incubation with acetosyringone, avirulent clones were produced in different proportions by strains T37 and C58 and also by a spontaneous variant of strain C58, denominated C58F. The proportion of avirulent clones in acetosyringone-treated cultures often exceeded 50%o for strains T37 and C58F and was of the order of 1% for strain C58. Control cultures not exposed to acetosyringone did not yield avirulent clones. Two other vir inducers, sinapinic acid and syringaldehyde, also inhibited growth and promoted accumulation of avirulent clones in cultures of strains C58F and T37. On the other hand, various acetosyringone analogs reported not to induce the vir genes did not act as growth inhibitors. All of the T37 and most of the C58F avirulent clones examined still carried a Ti plasmid. In all instances examined, avirulent clones still carrying a Ti plasmid were mutated in this plasmid. Mutants of strain C58F lacked the capacity to induce a virB::lacZ fusion in the presence of acetosyringone.The causal agent of crown gall, Agrobacterium tumefaciens, infects plant wounds where it detects the presence of specific compounds of plant origin. The virulence (vir) genes present on the Ti plasmids ofA. tumefaciens are then induced. Expression of the vir genes in the bacterium leads to the transfer to host plant cells of a particular region of the Ti plasmids, called the transferred (T-) DNA. This DNA of bacterial origin becomes stably integrated into the nuclear genome of the plant cells.

Agrobacterium tumefaciens C58F is a variant of strain C58 which generates a high proportion of avirulent mutants in the presence of the virulence (vir) gene inducer acetosyringone. These mutants are altered in the Ti plasmid and do not respond to the acetosyringone signal (C. Fortin, E. W. Nester, and P. Dion, J. Bacteriol. 174:5676-5685, 1992). The physical organization of the Ti plasmid was compared in strain C58 and its variant. One feature distinguishing pTiC58F from its parent plasmid was the presence of the insertion element IS426. Three copies of this element were detected in the strain C58 chromosome, whereas two additional copies were found in strain C58F, including one copy in the Ti plasmid. This particular copy of IS426 was associated with the region of arginine and nopaline catabolism of pTiC58F. Most of the avirulent mutants recovered following growth of strain C58F in the presence of acetosyringone were complemented by clones carrying either virA or virG. Element IS426 was no longer found in the arginine and nopaline catabolism region of the Ti plasmids from the virA and virG mutants, but it resided in the particular KpnI fragment containing the modified vir locus.Behavior of a strain C58F derivative, which was inactivated in a chromosomal component required for the response to acetosyringone, was consistent with the possibility that vir gene induction is essential to the massive production of avirulent mutants.The causal agent of crown gall disease, Agrobacterium tumefaciens, infects plant wounds where it detects the presence of specific compounds of plant origin. These compounds, which include acetosyringone and various analogs, induce the virulence (vir) genes carried by A. tumefaciens tumor-inducing (Ti) plasmids. Expression of the vir genes leads to the transfer to host plant cells of part of the Ti plasmids. The transferred DNA (T-DNA) becomes stably integrated in the nuclear genome of the plant, where it directs hormone-independent growth and the production of specific secondary metabolites, called opines (3,18). A nontransferred region on the Ti plasmids confers on the bacterium the ability to utilize opines as the sole carbon and nitrogen source. Octopine and nopaline are two particular types of opines (9).The vir region of A. tumefaciens comprises at least six operons which are required for full virulence. The vir operons virA and virG code for a two-component regulatory system (23, 37) responding to acetosyringone and other phenolic inducers (31). Chromosomal genes are also involved in tumorigenesis. One of these, the chvE gene, encodes a periplasmic galactose-binding protein which interacts with VirA to stimulate vir gene induction in the presence of sugars and low concentrations of acetosyringone (17).The various biological activities ofA. tumefaciens, including plant cell transformation, can be influenced by endogenous insertion sequence (IS) elements. Insertion of element IS66 in the tms region of T-DNA from pTiA6 results in an attenuated phenotype of this octopine-type strain (2, ...

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