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Nuclear and chloroplast cytokinin-binding proteins from barley leaves participating in transcription regulation

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Abstract

The 67 cytokinin-binding protein (CBP) previously isolated from the cytosol of mature leaves of 10-day-old barley seedlings has now been purified from nuclei isolated from the same leaves. The procedure of CBP isolation included protein purification by Sephadex G-50, hydrophobic chromatography on phenyl-Sepharose and affinity chromatography on zeatin-Sepharose. Interaction of trans-zeatin with the nuclear protein was demonstrated by the ELISA technique based on cytokinin competition with anti-idiotype antibodies (raised against antibodies to trans-zeatin) for complex formation with the protein immobilized on polystyrene microtiter plates. Nuclear 67 protein in concert with trans-zeatin activated transcription elongation in vitro in systems containing chromatin associated with RNA polymerase I or nuclei isolated from barley leaves. Nuclear 67 protein had no effect on the chloroplast transcription system. A 64 CBP was isolated from chloroplasts of barley leaves by the same procedure as that used for CBP isolation from nuclei. The cytokinin-binding properties of the chloroplast protein were demonstrated by competition of trans-zeatin with Aba-i in complex formation with the protein in ELISA. Chloroplast CBP activated RNA synthesis markedly in chloroplast lysate without any effect on transcription in the chromatin-containing system. Therefore both nuclear and chloroplast transcription machineries are regulated by cytokinins by means of special nuclear and chloroplast cytokinin-binding proteins which can be considered cytokinin receptors with the properties of transfactors regulating the elongation phase of transcription.

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Authors and Affiliations

  1. Timiryazev Institute of Plant Physiology, Russian Academy of Sciences, 127276, Moscow, Russia

    O.N. Kulaeva, N.N. Karavaiko, S.Yu. Selivankina, V.V. Kusnetsov, Ya.V. Zemlyachenko, G.N. Cherepneva, G.G. Maslova & T.V. Lukevich

  2. Institute of Biological Sciences, The University of Wales, Aberystwyth, SY23 3DA, UK

    A.R. Smith & M.A. Hall

Authors
  1. O.N. Kulaeva
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  2. N.N. Karavaiko
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  3. S.Yu. Selivankina
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  4. V.V. Kusnetsov
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  5. Ya.V. Zemlyachenko
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  6. G.N. Cherepneva
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  7. G.G. Maslova
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  8. T.V. Lukevich
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  9. A.R. Smith
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  10. M.A. Hall
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Kulaeva, O., Karavaiko, N., Selivankina, S. et al. Nuclear and chloroplast cytokinin-binding proteins from barley leaves participating in transcription regulation. Plant Growth Regulation 32, 329–335 (2000). https://doi.org/10.1023/A:1010783212459

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