Unique presence of 2-methylthio-ribosylzeatin in the transfer ribonucleic acid of the bacterium Pseudomonas aeruginosa

Thimmappaya, Bayar; Cherayil, Joseph D.

doi:10.1016/0006-291x(74)90292-7

Cited by 32 publications

(23 citation statements)

References 17 publications

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“…Their presence in saprophytic microorganisms has been restricted to pathogens and symbionts or species otherwise associated with overgrowth in plants. However, one apparent exception to such a functional association, the presence of ms2io6Ado in Pseudomonas aeruginosa tRNA, has been reported (26 …”

Section: Resultsmentioning

confidence: 99%

Cytokinins in Corynebacterium fascians Cultures

et al. 1976

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In addition to the four cytokinins, 6-(3-methyl-2-butenylamino)purine, 6-methylaminopunne and the cis and trans isomers of 6-(4-hydroxy-3-methyl-2-butenylamino)purine, reported earlier from our laboratories, three cytokinin-active fractions have been obtained from the aqueous medium of 6-day-old Corynebacterium fascians cultures. One of these has been identified as 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-2-methylthiopurine (2-methylthio-cis-zeatin, c-ms2io6Ade).The elution volumes of the other two frctions correspond to those of authentic 6-(4-hydroxy-3-methyl-2-butenylamino)-9-p-D-ribofuranosylpunne and 6-(3-methyl-2-butenylamino)-9-j3-D-ribofuranosylpurine, indicating the presence of trace amounts of these two ribonudeosides.Fasciation and modified growth of seedlings closely resembling symptoms of infection with Corynebacterium fascians have been induced in Pisum by soaking the seeds in high concentrations of kinetin (20). That in fact these symptoms of infection by the bacterium might be due to its production of cytokinin was indicated by the ability of extracts of pure cultures to release lateral buds of Pisum from apical dominance in the same manner as kinetin (25). Isolation of cytokinin-active material from aqueous cultures of the same stock5 was achieved by solvent extraction and chromatography. Three crystalline fractions with cytokinin activity as monitored in the tobacco bioassay were obtained (8, 10). The most active fraction was identified as 6(3-methyl-2-butenylamino)purine (i6Ade) (6), known at that time only as a synthetic, highly active cytokinin (5, 9). The second fraction contained nicotinamide. Inasmuch as synthetic nicotinamide was inactive in the tobacco bioassay, the activity of the isolated material was ascribed to a contaminant. This cytokinin was recently identified as 6-(4-hydroxy-3-methyl-cis-2-buteny- for the activity of this fraction in the tobacco bioassay. Hence, another more potent cytokinin was assumed to be present as a contaminant and to be mainly responsible for the cytokinin activity in this fraction. We now report the purification and characterization of this cytokinin from Corynebacterium fascians culture medium as 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-2-methylthiopurine (2-methylthio-cis-zeatin, c-ms2io6Ade). Paper Chromatography. Whatman No. 1 filter paper, washed as described (21), was used. Cytokinin samples were dissolved in small volumes of 95% ethanol and applied to the paper as 5-cm streaks. The chromatograms were developed by ascending chromatography over 30 cm. UV-absorbipg bands were recovered from the chromatograms in small volumes (about 0.5-1 ml) by 749 www.plantphysiol.org on May 11, 2018 -Published by Downloaded from MATERIALS AND METHODS Growth of Corynebacterium fascians

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Section: Resultsmentioning

confidence: 99%

Cytokinins in Corynebacterium fascians Cultures

et al. 1976

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“…iPA: isopentenyladenosine: 6-(3-methyl-2-butenylamino)-9-,8-D-ribofuranosylpurine; msiPA: methylthio-isopentenyladenosine: 6-(3-methyl-2-butenylamino)-2 -methylthio-9-/,3-D-ribofuranosylpurine; CTAB: cetyltrimethylammonium bromide; KE: kinetin equivalent, deand msZR (25). However, msZR has been found in tRNA of Pseudomonas aeruginosa (27) and recently Chapman et al (8) have reported the presence of t-ZR in the tRNAs of Agrobacterium tumefaciens. Unlike bacterial tRNA, plant tRNA contains predominantly c-ZR.…”

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Cytokinins in tRNA Obtained from Spinacia oleracea L. Leaves and Isolated Chloroplasts

Vreman

Thomas²,

Corse³

et al. 1978

Plant Physiol.

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Cytokinin-active ribonudeosides have been isolated from tRNA of whole spinach (Spinacia oleracea L.) leaves and isolated spinach chloroplasts. The tRNA from spinach leaf blades contained: 6-(4-hydroxy-3-methyl-2-butenylamino)-9-fi-D-ribofuranosylpurine (cis and trans isomers), 6-(3-methyl-2-butenylamino)-9-fi-D-ribofuranosylpurine, and 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthio-9-.8-Dribofuranosylpurine (cis and trans isomers). A method for isolation of large amounts of intact chloroplasts was developed and subsequently used for the isolation of chloroplast tRNA. The chloroplast tRNA contained 6-(3-methyl-2-butenylamino)-9-fi-D-ribofuranosylpurine and 6-(4-hydroxy-3-methyl-2-butenylamino)-2 -methylthio-9-P-D-ribofunrnosylpurine (the cis isomer only). The structures of these compounds were assigned on the basis of their chromatographic properties and mass spectra of trimethylsilyl derivatives which were identical with those of the corresponding synthetic compounds. The results of this study indicate that ribosylzeatin was present in spinach leaf tRNA, but absent from the purified chloroplast tRNA preparation.Cytokinin-active compounds have been demonstrated to occur in tRNA preparations from various phylogenetically divergent groups of organisms, such as bacteria, fungi, plants, and animals (25). Studies on the distribution of cytokinins in the tRNAs of these organisms show that whereas microbial tRNAs contain mainly iPA3 and msiPA, plant tRNAs contain in addition ZR I This research was performed when H.

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“…Evidence from various microorganisms shows that i6A and ms2i6A are the two major cytokinins found in their tRNAs (2,6,27). However, c-io6A and ms2io6A have been found in culture filtrates of certain plant pathogens (16,17,20), and recently Thimmappaya and Cherayil (29) CTAB: cetyltrimethylammonium bromide; KE: kinetin equivalent, defined as the ,ug of kinetin (6-furfurylaminopurine) required to give the same activity as the test sample under the specified conditions. tRNA preparations.…”

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Subcellular Localization of Cytokinins in Transfer Ribonucleic Acid

1977

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Evidence on the localization of cytokinins in chloroplast tRNA was obtained by comparison of Euglena gracilis var. bacillaris light-grown and dark-grown wild type cultures and chloroplast-bleached mutant strains. The several cytokinins characteristic of tRNA were separated by Sephadex LH-20 column chromatography of the hydrolysates and were quantitatively determined by tobacco bioassays of the eluates. The results indicate that 6-(3-methyl-2-butenylamino)-9-l3-D-ribofuranosylpurine (i6A) is formed in both the cytoplasmic and chloroplast tRNA, whereas 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-9-fl-D-ribofuranosylpurine (c-io6A) is produced mainly in the cytoplasmic tRNA and 6-(4-hydroxy-3-methyl-2-butenylamiao)-2-methylthio-9-fi-D-ribofuranosylpurine (ms2io6A) is localized exdusively in chloroplast tRNA. The restriction of the methiolation reaction to the chloropbst is supported by results of radioisotope experiments showing that 3S-labeled MgSO4 is incorported into ms2io6A in the wild type cultures, but not in the chloroplast-bleached mutant strains.Preparations of unfractionated tRNAs from various sources of wide evolutionary divergence have been shown to contain cytokinin-active nucleosides (3,12,26). To date, the four major active nucleosides which have been identified in tRNAs are i6A,3 c-io6A, ms2i6A, and ms2io6A. Evidence from various microorganisms shows that i6A and ms2i6A are the two major cytokinins found in their tRNAs (2,6,27). However, c-io6A and ms2io6A have been found in culture filtrates of certain plant pathogens (16,17,20), and recently Thimmappaya and Cherayil (29) 3 Abbreviations: i6A: N6-(A2-isopentenyl)adenosine or 6-(3-methyl-2-butenylamino)-9-/3-D-ribofuranosylpurine; t-io6A: ribosyl-trans-zeatin or 6-(4-hydroxy-3-methyl-trans-2-butenylamino)-9-/3-D-ribofuranosylpurine; c-io6A: ribosyl-cis-zeatin or 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-9-,8--ribofuranosylpurine; ms2i6A: 2-methylthio-NO-isopentenyladenosine or 6-(3-methyl-2-butenylamino)-2-methylthio-9-13D-ribofuranosylpurine; ms2io6A: ribosyl-2-methylthiozeatin or 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthio-9-f-D-ribofuranosylpurine; CTAB: cetyltrimethylammonium bromide; KE: kinetin equivalent, defined as the ,ug of kinetin (6-furfurylaminopurine) required to give the same activity as the test sample under the specified conditions. tRNA preparations. Hall et al. (13) have reported c-io6A and i6 A as constituents of plant tRNA, namely from corn kernels, spinach, and frozen peas. Wheat germ tRNA contains four cytokinins, i6A, io6A, and their methylthio derivatives (5). Pea root tRNA hydrolysates have been shown to contain mainly io6A and i6A (25). Vreman et al. (30,31) have reported the presence of i6A, ms2i6A, c-io6A, c-ms2io6A, and t-ms2io06A in tRNA hydrolysates from pea shoots. Thus, the cytokinins show greater diversity in plant tRNAs than in animal tRNAs or than generally found in tRNAs of microorganisms. The presence of different cytokinins in tRNAs of a single organism raises questions as to their origin and pos...

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Unique presence of 2-methylthio-ribosylzeatin in the transfer ribonucleic acid of the bacterium Pseudomonas aeruginosa

Cited by 32 publications

References 17 publications

Cytokinins in Corynebacterium fascians Cultures

Cytokinins in Corynebacterium fascians Cultures

Cytokinins in tRNA Obtained from Spinacia oleracea L. Leaves and Isolated Chloroplasts

Subcellular Localization of Cytokinins in Transfer Ribonucleic Acid

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