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RNAi suppression of RPN12a decreases the expression of type-A ARRs, negative regulators of cytokinin signaling pathway, in Arabidopsis

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Molecules and Cells

Abstract

The 26S proteasome is a 2-MDa complex with a central role in protein turn over. The 26S proteasome is comprised of one 20S core particle and two 19S regulatory particles (RPs). The RPN12a protein, a non-ATPase subunit of the 19S RP, was previously shown to be involved in cytokinin signaling in Arabidopsis. To further investigate cellular roles of RPN12a, RNAi transgenic plants of RPN12a were constructed. As expected, the 35S:RNAi-RPN12a plants showed cytokinin signaling defective phenotypes, including abnormal formation of leaves and inflorescences. Furthermore, RNAi knock-down transgenic plants exhibited additional unique phenotypes, including concave and heart-shape cotyledons, triple cotyledons, irregular and clustered guard cells, and defects in phyllotaxy, all of which are typical for defective cytokinin signaling. We next examined the mRNA level of cytokinin signaling components, including type-A ARRs, type-B ARRs, and CRFs. The expression of type-A ARRs, encoding negative regulators of cytokinin signaling, was markedly reduced in 35S:RNAi-RPN12a transgenic plants relative to that in wild type plants, while type-B ARRs and CRFs were unaffected. Our results also indicate that in vivo stability of the ARR5 protein, a negative regulator of cytokinin signaling, is mediated by the 26S proteasome complex. These results suggest that RPN12a participates in feedback inhibitory mechanism of cytokinin signaling through modulation of the abundance of ARR5 protein in Arabidopsis.

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References

  • Bae, H., Choi, S.M., Yang, S.W., Pai, H.-S., and Kim, W.T. (2009). Suppression of the ER-localized AAA ATPase NgCDC48 inhibits tobacco growth and development. Mol. Cells 28, 57–65.

    Article  CAS  PubMed  Google Scholar 

  • Bergmann, D.C., and Sack, F.D. (2007). Stomatal development. Annu. Rev. Plant Biol. 58, 163–181.

    Article  CAS  PubMed  Google Scholar 

  • Bergmann, D.C., Lukowitz, W., and Somerville, C.R. (2004). Stomatal development and pattern controlled by a MAPKK kinase. Science 304, 1494–1497.

    Article  CAS  PubMed  Google Scholar 

  • Brenner, W.G., Romanov, G.A., Köllmer, I., Bürkle, L., and Schmülling, T. (2005). Immediate-early and delayed cytokinin response genes of Arabidopsis thaliana identified by genome-wide expression profiling reveal novel cytokinin-sensitive processes and suggest cytokinin action through transcriptional cascades. Plant J. 44, 314–333.

    Article  CAS  PubMed  Google Scholar 

  • Brukhin, V., Gheyselinck, J., Gagliardini, V., Genschik, G., and Grossniklausa, U. (2005). The RPN1 subunit of the 26S proteasome in Arabidopsis is essential for embryogenesis. Plant Cell 17, 2723–2737.

    Article  CAS  PubMed  Google Scholar 

  • Cho, Y-.H., Yoo, S-.D., and Sheen, J. (2006a). Regulatory functions of nuclear hexokinase1 complexin glucose signaling. Cell 127, 579–589.

    Article  CAS  PubMed  Google Scholar 

  • Cho, S.K., Chung, H.S., Ryu, M.Y., Park, M.J., Lee, M.M., Bahk, Y.-Y., Kim, J., Pai, H.S., and Kim, W.T. (2006b). Heterologous expression and molecular and cellular characterization of CaPUB1 encoding a hot pepper U-box E3 ubiquitin ligase homolog. Plant Physiol. 142, 1664–1682.

    Article  CAS  PubMed  Google Scholar 

  • Cho, S.K., Ryu, M.Y., Song, C., Kwak, J.M., and Kim, W.T. (2008). Arabidopsis PUB22 and PUB23 are homologous U-Box E3 ubiquitin ligases that play combinatory roles in response to drought stress. Plant Cell 20, 1899–1914.

    Article  CAS  PubMed  Google Scholar 

  • Choi, J., and Hwang, I. (2007). Cytokinin: Perception, signal transduction, and role in plant growth and development. J. Plant Biol. 50, 98–108.

    Article  CAS  Google Scholar 

  • Dreher, K., and Callis, J. (2007). Ubiquitin, hormones and biotic stress in plants. Ann. Bot. 99, 787–822.

    Article  CAS  PubMed  Google Scholar 

  • Ferdous, A., Gonzalez, F., Sun, L., Kodadek, T., and Johnston, S.A. (2001). The 19S regulatory particle of the proteasome is required for efficient transcription elongation by RNA polymerase II. Mol. Cell 7, 981–991.

    Article  CAS  PubMed  Google Scholar 

  • Ferreira, F.J., and Kieber, J.J. (2005). Cytokinin signaling. Curr. Opin. Plant Biol. 8, 518–525.

    Article  CAS  PubMed  Google Scholar 

  • Fu, H., Reis, N., Lee, Y., Glickman, M.H., and Vierstra, R.D. (2001). Subunit interaction maps for the regulatory particle of the 26S proteasome and the COP9 signalosome. EMBO J. 20, 7096–7107.

    Article  CAS  PubMed  Google Scholar 

  • Giulini, A., Wang, J., and Jackson, D. (2004). Control of phyllotaxy by the cytokinin-inducible response regulator homologue ABPHYL1. Nature 430, 1031–1034.

    Article  CAS  PubMed  Google Scholar 

  • Glickman, M.H., and Ciechanover, A. (2002). The ubiquitin-proteasome proteolytic pathway: destruction for the sake of construction. Physiol. Rev. 82, 373–428.

    CAS  PubMed  Google Scholar 

  • Huang, W., Pi, L., Liang, W., Xu, B., Wang, H., Dai, R., and Huang, H. (2006). The proteolytic function of the Arabidopsis 26S proteasome is required for specifying leaf adaxial identity. Plant Cell 18, 2479–2492.

    Article  CAS  PubMed  Google Scholar 

  • Hwang, I., and Sheen, J. (2001). Two-component circuitry in Arabidopsis cytokinin signal transduction. Nature 413, 383–389.

    Article  CAS  PubMed  Google Scholar 

  • Ichimura, Y., Kirisako, T., Takao, T., Satomi, Y., Shimonishi, Y., and Ishihara, N. (2000). A ubiquitin-like system mediates protein lipidation. Nature 408, 488–492.

    Article  CAS  PubMed  Google Scholar 

  • Jean, R. (1994). Phyllotaxis: a systemic study in plant morphogenesis. (Cambridge Univ. Press).

  • Jin, H., Li, S., and Villegas, A. Jr. (2006). Down-regulation of the 26S proteasome subunit RPN9 inhibits viral systemic transport and alters plant vascular development. Plant Physiol. 142, 651–661.

    Article  CAS  PubMed  Google Scholar 

  • Joo, S., Seo, Y.S., Kim, S.M., Hong, D.K., Park, K.Y., and Kim, W.T. (2006). Brassinosteroid-induction of AtACS4 encoding an auxin-responsive 1-aminocyclopropane-1-carboxylate synthase 4 in Arabidopsis seedlings. Physiol. Plant. 126, 592–604.

    CAS  Google Scholar 

  • Kraft, E., Stone, S.L., Ma, L., Su, N., Gao, Y., Lau, O.-S., Deng, X.-W., and Callis, J. (2005). Genome analysis and functional characterization of the E2 and RING-type E3 ligase ubiquitination enzymes of Arabidopsis. Plant Physiol. 139, 1597–1611.

    Article  CAS  PubMed  Google Scholar 

  • Kuhlemeier, C., and Reinhardt, D. (2001). Auxin and phyllotaxis. Trends Plant Sci. 6, 187–189.

    Article  CAS  PubMed  Google Scholar 

  • Kurepa, J., Toh-E, A., and Smalle, J.A. (2008). 26S proteasome regulatory particle mutants have increased oxidative stress tolerance. Plant J. 53, 102–114.

    Article  CAS  PubMed  Google Scholar 

  • Lee, D., Ezhkova, E., Li, B., Pattenden, S.G., Tansey, W.P., and Workman, J.L. (2005). The proteasome regulatory particle alters the SAGA coactivator to enhance its interactions with transcriptional activators. Cell 123, 423–436.

    Article  CAS  PubMed  Google Scholar 

  • Lee, J.-H., Deng, X.W., and Kim, W.T. (2006a). Possible role of light in the maintenance of EIN3/EIL1 stability in Arabidopsis seedlings. Biochem. Biophys. Res. Commun. 350, 484–491.

    Article  CAS  PubMed  Google Scholar 

  • Lee, B.-J., Kwon, S.J., Kim, S.-K., Kim, K.-J., Park, C.-J., Kim, Y.-J., Park, O.K., and Peak, K.-H. (2006b). Functional study of hot pepper 26S proteasome subunit RPN7 induced by Tobacco mosaic virus from nuclear proteome analysis. Biochem. Biophys. Res. Commun. 351, 405–411.

    Article  CAS  PubMed  Google Scholar 

  • Lee, D.J., Park, J.Y., Ku, S.J., Ha, Y.M., Kim, S., Kim, M.D., Oh, M.H., and Kim, J. (2007a) Genome-wide expression profiling of ARABIDOPSIS RESPONSE REGULATOR 7 (ARR7) overexpression in cytokinin response. Mol. Genet. Genomics 277, 115–137.

    Article  CAS  PubMed  Google Scholar 

  • Lee, M.O., Hwang, J.H., Lee, D.H., and Hong, C.B. (2007b). Gene expression profile for Nicotiana tabacum in the early phase of flooding stress. J. Plant Biol. 50, 496–503.

    Article  CAS  Google Scholar 

  • Lee, D.J., Kim, S., Ha, Y.-M., and Kim, J. (2008). Phosphorylation of Arabidopsis response regulator 7 (ARR7) at the putative phosphor-accepting site is required for ARR7 to act as a negative regulator of cytokinin signaling. Planta 227, 577–587.

    Article  CAS  PubMed  Google Scholar 

  • Lee, H.K., Cho, S.K., Son, O., Xu, Z., Hwang, I., and Kim, W.T. (2009). Drought stress-induced Rma1H1, a RING membraneanchor E3 ubiquitin ligase homolog, regulates aquaporin levels via ubiquitination in transgenic Arabidopsis plants. Plant Cell 21, 622–641.

    Article  CAS  PubMed  Google Scholar 

  • Leibfried, A., To, J.P.C., Busch, W., Stehling, S., Kehle, A., Demar, M., Kieber, J.J., and Lohmann, J.U. (2005). WUSCHEL controls meristem function by direct regulation of cytokinin-inducible response regulators. Nature 438, 1172–1175.

    Article  CAS  PubMed  Google Scholar 

  • Moon, J., Parry, G., and Estelle, M. (2004). The Ubiquitin-proteasome pathway and plant development. Plant Cell 16, 3181–3195.

    Article  CAS  PubMed  Google Scholar 

  • Mukhopadhyay, D., and Riezman, H. (2007). Proteasome-independent functions of ubiquitin in endocytosis and signaling. Science 315, 201–205

    Article  CAS  PubMed  Google Scholar 

  • Oh, M., Lee, H., Kim, Y.-K., Nam, J.-W., Rhee, J.-K., Zhang, B.-T., Kim, V.N., and Lee, I. (2007). Identification and characterization of small RNAs from vernalized Arabidopsis thaliana. J. Plant Biol. 50, 562–572.

    Article  CAS  Google Scholar 

  • Pickart, C.M. (2001). Ubiquitin enters the new millennium. Mol. Cell 8, 499–504.

    Article  CAS  PubMed  Google Scholar 

  • Pickart, C.M., and Eddins, M.J. (2004). Ubiquitin: structures, functions, mechanisms. Biochim. Biophys. Acta 1695, 55–72.

    Article  CAS  PubMed  Google Scholar 

  • Rashotte, A.M., Mason, M.G., Hutchison, C.E., Ferreira, F.J., Schaller, G.E., and Kieber, J.J. (2006). A subset of Arabidopsis AP2 transcription factors mediates cytokinin responses in concert with a two-component pathway. Proc. Natl. Acad. Sci. USA 103, 11081–11085.

    Article  CAS  PubMed  Google Scholar 

  • Seo, Y.S., Kim, E.Y., Mang, H.G., and Kim, W.T. (2008). Heterologous expression and biochemical and cellular characterization of CaPLA1 encoding a hot pepper phospholipase A1 homolog. Plant J. 53, 895–908.

    Article  CAS  PubMed  Google Scholar 

  • Seo, Y.S., Kim, E.Y., Kim, J.H., and Kim, W.T. (2009). Enzymatic characterization of class I DAD1-like acylhydrolase members targeted to chloroplast in Arabidopsis. FEBS Lett. 583, 2301–2307.

    Article  CAS  PubMed  Google Scholar 

  • Sheen, J. (2002). Phosphorelay and transcription control in cytokinin signal transduction. Science 296, 1650–1652.

    Article  CAS  PubMed  Google Scholar 

  • Smalle, J., and Vierstra, R.D. (2004) The ubiquitin 26S proteasome proteolytic pathway. Annu. Rev. Plant Biol. 55, 555–590.

    Article  CAS  PubMed  Google Scholar 

  • Smalle, J., Kurepa, J., Yang, P., Babiychuk, E., Kushnir, S., Durski, A., and Vierstra, R.D. (2002). Cytokinin growth responses in Arabidopsis involve the 26S proteasome subunit RPN12. Plant Cell 14, 17–32.

    Article  CAS  PubMed  Google Scholar 

  • Smalle, J., Kurepa, J., Yang, P., Emborg, T.J., Babiychuk, E., Kushnir, S., Vierstra, R.D. (2003). The pleiotropic role of the 26S proteasome subunit RPN10 in Arabidopsis growth and development supports a substrate-specific function in abscisic acid signaling. Plant Cell 15, 965–980.

    Article  CAS  PubMed  Google Scholar 

  • Stone, S.L., Hauksdottir, H., Troy, A., Herschleb, J., Kraft, E., and Callis, J. (2005) Functional analysis of the RING-type ubiquitin ligase family of Arabidopsis. Plant Physiol. 137, 13–30.

    Article  CAS  PubMed  Google Scholar 

  • To, J.P.C., and Kieber, J.J. (2008). Cytokinin signaling: two-components and more. Trends Plant Sci. 13, 85–92.

    Article  CAS  PubMed  Google Scholar 

  • To, J.P.C., Deruère, J., Maxwell, B.B., Morris, V.F., Hutchison, C.E., Ferreira, F.J, Schaller, G.E., and Kieber, J.J. (2007). Cytokinin regulates type-A Arabidopsis Response Regulator activity and protein stability via two-component phosphorelay. Plant Cell 19, 3901–3914.

    Article  CAS  PubMed  Google Scholar 

  • Tran, L.-S.P., Urao, T., Qin, F., Maruyama, K., Kakimoto, T., Shinozaki, K., and Yamaguchi-Shinozaki, K. (2007). Functional analysis of AHK1/ATHK1 and cytokinin receptor histidine kinases in response to abscisic acid, drought, and salt stress in Arabidopsis. Proc. Natl. Acad. Sci. USA 104, 20623–20628.

    Article  CAS  PubMed  Google Scholar 

  • Urao, T., Yamaguchi-Shinozaki, K., and Shinozaki, K. (2001). Plant histidine kinases: An emerging picture of two-component signal transduction in hormone and environmental responses. Sci. STKE. 2001, re18.

    Article  CAS  PubMed  Google Scholar 

  • Vierstra, R.D. (2003). The ubiquitin/26S proteasome pathway, the complex last chapter in the life of many plant proteins. Trends Plant Sci. 8, 135–142.

    Article  CAS  PubMed  Google Scholar 

  • Voges, D., Zwickl, P., and Baumeister, W. (1999). The 26S proteasome: a molecular machine designed for controlled proteolysis. Annu. Rev. Biochem. 68, 1015–1068.

    Article  CAS  PubMed  Google Scholar 

  • Wohlbach, D.J., Quirino, B.F., and Sussman, M.R. (2008). Analysis of the Arabidopsis histidine kinase ATHK1 reveals a connection between vegetative osmotic stress sensing and seed maturation. Plant Cell 20, 1101–1117.

    Article  CAS  PubMed  Google Scholar 

  • Wojcik, C. (2001). Ubiquitin-more than just a signal for protein degradation. Trends Cell Biol. 11, 397–399.

    Article  CAS  PubMed  Google Scholar 

  • Yee, D., and Goring, D.R. (2009). The diversity of plant U-box E3 ubiquitin ligases: from upstream activators to downstream target substrates. J. Exp. Bot. 60, 1109–1121.

    Article  CAS  PubMed  Google Scholar 

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Correspondence to Woo Taek Kim.

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Ryu, M.Y., Cho, S.K. & Kim, W.T. RNAi suppression of RPN12a decreases the expression of type-A ARRs, negative regulators of cytokinin signaling pathway, in Arabidopsis . Mol Cells 28, 375–382 (2009). https://doi.org/10.1007/s10059-009-0132-x

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  • DOI: https://doi.org/10.1007/s10059-009-0132-x

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