The cytokinin-active isoprenoid nucleosides of Agrobacterium tumefaciens transfer ribonucleic acid were identified by high-pressure liquid chromatography, permethylation, and mass spectroscopy. Besides the expected 6-[(3-methylbut-2-enyl)amino]-9-(beta-D-ribofuranosyl)purine (i6A) and its 2-methylthio derivative (ms2i6A), substantial amounts of cis- and trans-ribosylzeatin (io6A) and cis-2-(methylthio)ribosylzeatin (c-ms2io6A) were present. These hydroxylated side chain derivatives are normally characteristic of plant tRNA. Fractionation of the total bacterial tRNA on BD-cellulose and RPC-5 allowed isolation of purified iso-accepting species whose cytokinin nucleoside contents were then determined. Distribution of the isoprenoid nucleosides among the U-group tRNA species was not uniform. cis-Ribosylzeatin was found almost exclusively in one tRNASer while ms2io6A was found predominantly in tRNAPhe, tRNASer, and tRNATyr. Not all cytokinin-active species were found in every member of the U-group tRNAs. The only species present in tRNATrp was i6A; it contained no zeatin derivatives. The hydroxylation and methylthiolation processes appear to be highly specific and dependent upon tRNA structure or sequence.

The distribution of cytokinin activity in wheat ( Tnticum aesdivum) germ tRNA fractionated by BD-cellulose and RPC-5 chromatography has been examined. As In other organisms, the cytokinia moieties in wheat germ tRNA appear to be restricted to tRNA species that would be expected to respond to codons ginnn with U. Only a few of the wheat germ tRNA species in this coding group actually contain cytokinin modifications.Cytokinin activity was associated with isoaccepting tRNAsr species and with a minor tRNALU species from wheat germ. All other wheat germ tRNA species corresponding to codons bnning with U were devoid of cytokinin activity in the tobacco callus bioassay.Trace amounts of the trans-isomers of io6A and ms2io6A have also been reported to be present (3,21,22). Information concerning the distribution of cytokinins with respect to particular plant tRNA species is much more limited. Ribosyl-cis-zeatin has been identified as a constituent of a minor tRNALu species from peas (8), but the distribution of cytokinin moieties in other tRNA species from this source was not examined.The (12,20). Because not all tRNA species within the U-group necessarily contain a cytokinin modification, the distribution of cytokinin moieties with respect to particular tRNA species may vary in different organisms. The cytokinin modifications of plant tRNA species are of particular interest in view of the hormonal function of cytokinins in plant systems. Cytokinin-active ribonucleosides have been isolated and identified from hydrolysates of a number of plant tRNA preparations (5,6,13,22). In addition to i6A, plant tRNA preparations typically contain ribosyl-cis-zeatin (io6A) and the 2-methylthio derivatives of these two compounds (ms2io6A and ms2i6A).