Transfer RNA Is the Source of Extracellular Isopentenyladenine in a Ti-Plasmidless Strain of Agrobacterium tumefaciens

Even in the absence of the classical Ti plasmid-encoded cytokinin biosynthetic genes ipt and tzs, Agrobacterium tumefaciens strains still release significant amounts of the cytokinin isopentenyladenine (iP) into the culture medium (R.W. Kaiss-Chapman and R.O. Morris [1977] Biochem Biophys Res Commun 76: 453-459). A potential source of the iP is isopentenylated transfer RNA (tRNA), which, in turn, is synthesized by the activity of tRNA:isopentenyltransferase encoded by the bacterial miaA gene. To determine whether secreted iP had its origin in isopentenylated tRNA, a miaA- deletion/insertion mutant was prepared and reconstructed in Agrobacterium tumefaciens in vivo. The mutant no longer possessed tRNA:isopentenylation activity and no longer released iP into the extracellular medium. Transfer RNA therefore makes a small but significant contribution to the total amount of cytokinin normally secreted by Agrobacterium strains. tRNA-mediated synthesis may also account for cytokinin production by other plant-associated bacteria, such as Rhizobia, that have been reported to secrete similarly low levels of nonhydroxylated cytokinins.