Expression in Vitro and During Plant Pathogenesis of the syrB Gene Required for Syringomycin Production by Pseudomonas syringae pv. syringae. Yin- Yuan Mo. Department of Plant Pathology, Washington State University, Pullman 99164–6430 U.S.A. Dennis C. Gross. Department of Plant Pathology, Washington State University, Pullman 99164–6430 U.S.A. MPMI 4:28-36. Accepted 20 August 1990. This article is in the public domain and not copyrightable. It may be freely reprinted with customary crediting of the source. The American Phytopathological Society, 1991.

The syrB gene required for syringomycin production by Pseudomonas syringae pv. syringae B301D was subjected to Tn3HoHo1 mutagenesis to generate random transcriptional fusions to a promoterless lac operon. Approximately 80 Tn3HoHo1 insertions were mapped to the 3.1-kilobase syrB locus, and the direction of transcription for syrB was determined from those inserts expressing Β -galactosidase activity. Besides nontoxigenicity, Tn3HoHo1 mutations caused deficiencies in proteins SR4 and SR5, postulated to constitute part of the syringomycin synthetase complex in strain B301D. Three inserts (i.e., 132, 257, and 329) that were marker exchanged into the genome of strain B301D-R each expressed ~1,300 Β-galactosidase units after incubation on an agar medium conducive to syringomycin production. Tn3HoHo1 insert 132 was marker exchanged into strains B301D-R and B3A-R to yield BR132 and B3AR132, respectively. The syrB::lacZ fusion in strain BR132 expressed Β-galactosidase activity in the nutritionally defined SRM liquid medium (D. C. Gross, Journal of Applied Bacteriology 58:167-174, 1985) as compared to nonexpression of the same fusion in strain B3AR132. In contrast, both syrB::lacZ recombinant strains expressed activity in a medium containing a potato extract; activity was expressed about 24 hr before the initial detection of toxin production in parental strains B301D-R and B3A-R. Like syringomycin production itself, the syrB::lacZ fusion was regulated in a positive manner by iron, since quantities of Fe³⁺ >2 µM were required for maximum expression. In immature cherry fruits, the syrB::lacZ fusions in strains BR132 and B3AR132 were expressed soon after inoculation, although peak Β-galactosidase activity of ~400 units occurred at day 1 for strain B3AR132 versus day 3 for BR132. Addition of a water-soluble extract from cherry leaves to SRM liquid medium induced strain B3AR132 to express Β-galactosidase activity. Signal molecules in cherry tissues were inferred to induce the syrB::lacZ fusion in strain B3AR132.