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Suppression of the ER-localized AAA ATPase NgCDC48 inhibits tobacco growth and development

  • Published:
Molecules and Cells

Abstract

CDC48 is a member of the AAA ATPase superfamily. Yeast CDC48 and its mammalian homolog p97 are implicated in diverse cellular processes, including mitosis, membrane fusion, and ubiquitin-dependent protein degradation. However, the cellular functions of plant CDC48 proteins are largely unknown. In the present study, we performed virus-induced gene silencing (VIGS) screening and found that silencing of a gene encoding a tobacco CDC48 homolog, NgCDC48, resulted in severe abnormalities in leaf and shoot development in tobacco. Furthermore, transgenic tobacco plants (35S:anti-NgCDC48), in which the NgCDC48 gene was suppressed using the antisense RNA method, exhibited severely aberrant development of both vegetative and reproductive organs, resulting in arrested shoot and leaf growth and sterile flowers. Approximately 57–83% of 35S:anti-NgCDC48 plants failed to develop mature organs and died at early stage of development. Scanning electron microscopy showed that both adaxial and abaxial epidermal pavement cells in antisense transgenic leaves were significantly smaller and more numerous than those in wild type leaves. These results indicate that NgCDC48 is critically involved in cell growth and development of tobacco plants. An in vivo targeting experiment revealed that NgCDC48 resides in the endoplasmic reticulum (ER) in tobacco protoplasts. We consider the tantalizing possibility that CDC48-mediated degradation of an as-yet unidentified protein(s) in the ER might be a critical step for cell growth and expansion in tobacco leaves.

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References

  • Aker, J., Hesselink, R., Engel, R., Karlova, R., Borst J.W., Visser, A.J.W.G., and de Vries, S.C. (2007). In vivo hexamerization and characterization of the Arabidopsis AAA ATPase CDC48A complex using Forster resonance energy transfer-fluorescence lifetime imaging microscopy and fluorescence correlation spectroscopy. Plant Physiol. 145, 339–350.

    Article  PubMed  CAS  Google Scholar 

  • Bays, N.W., and Hampton, R.Y. (2002). Cdc48-Ufd1-Npl4: Stuck in the middle with Ub, Curr. Biol. 12, R366–371.

    Article  PubMed  CAS  Google Scholar 

  • Beyer, A. (1997). Sequence analysis of the AAA protein family. Protein Sci. 6, 2043–2058.

    Article  PubMed  CAS  Google Scholar 

  • Buaboocha, T., Liao, B., and Zielinski, R.E. (2001). Isolation of cDNA and genomic DNA clones encoding a calmodulin-binding protein related to a family of ATPases involved in cell division and vesicle fusion. Planta 212, 774–781.

    Article  PubMed  CAS  Google Scholar 

  • Cao, K., and Zheng, Y. (2004). The Cdc48/p97-Ufd1-Npl4 complex: its potential role in coordinating cellular morphogenesis during the M-G1 transition. Cell Cycle 3, 422–424.

    PubMed  CAS  Google Scholar 

  • Cheeseman, I.M., and Desai, A (2004). Cell division: AAAtacking the mitotic spindle. Curr. Biol. 14, R70–72.

    Article  PubMed  CAS  Google Scholar 

  • Cho, H.-K., Park, J.-A., and Pai, H.-S. (2008a). Physiological function of NbRanBP1 in Nicotiana benthamiana. Mol. Cells 26, 270–277.

    PubMed  CAS  Google Scholar 

  • Cho, S.K, Ryu, M.Y., Song, C., Kwak, J.M., and Kim, W.T. (2008b). 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  PubMed  CAS  Google Scholar 

  • Decottignies, A., Evain, A., and Ghislain, M. (2004). Binding of Cdc48p to a ubiquitin-related UBX domain from novel yeast proteins involved in intracellular proteolysis and sporulation, Yeast 21, 127–139.

    Article  PubMed  CAS  Google Scholar 

  • Deng, X.W., Caspar, T., and Quail, P.H. (1991). cop1: a regulatory locus involved in light-controlled development and gene expression in Arabidopsis. Genes Dev. 5, 1172–1182.

    Article  PubMed  CAS  Google Scholar 

  • Feiler, H.S., Desprez, T., Santoni, V., Kronenberger, J., Caboche, M., and Traas, J. (1995). The higher plant Arabidopsis thaliana encodes a functional CDC48 homologue which is highly expressed in dividing and expanding cells. EMBO J. 14, 5626–5637.

    PubMed  CAS  Google Scholar 

  • Fu, X., Ng, C., Feng, D., and Liang, C. (2003). Cdc48p is required for the cell cycle commitment point at start via degradation of the G1-CDK inhibitor Far1p. J. Cell Biol. 163, 21–26.

    Article  PubMed  CAS  Google Scholar 

  • Hetzer, M., Meyer, H.H., Walther, T.C., Biobao-Cortes, D., Warren, G., and Mattaj, I.W. (2001). Distinct AAA-ATPase p97 complexes function in discrete steps of nuclear assembly. Nat. Cell Biol. 3, 1086–1091.

    Article  PubMed  CAS  Google Scholar 

  • Hong, J.K., Hwang, J.E., Chung, W.S., Lee, K.O., Choi, Y.J., Gal, S.W., Park, B.-S., and Lim, C.O. (2008). Expression of a chinese cabbage cysteine protease inhibitor, BrCYS1, retards seed germination and plant growth in transgenic tobacco plants. J. Plant Biol. 51, 347–353.

    Article  CAS  Google Scholar 

  • Jentsch, S., and Rumpf, S. (2007). Cdc48 (p97): a “molecular gearbox” in the ubiquitin pathway? Trends Biochem. Sci. 32, 6–11.

    Article  PubMed  CAS  Google Scholar 

  • Kim, M., Lim, J.-H., Ahn, C.S., Park, K., Kim, G.T., Kim, W.T., and Pai, H.-S. (2006). Mitochondria-associated hexokinase play a role in the control of programmed cell death in Nicotiana benthamiana. Plant Cell 18, 2341–2355.

    Article  PubMed  CAS  Google Scholar 

  • Lee, J.-H., and Kim, W.T. (2003). Molecular and biochemical characterization of VR-EILs encoding mung bean ETHYLENE INSENSITIVE3-LIKE proteins. Plant Physiol. 132, 1475–1488.

    Article  PubMed  CAS  Google Scholar 

  • Lee, J.-H., Hong, J.-P., Oh, S.-K., Lee, S., Choi, D., and Kim, W.T. (2004). The ethylene-responsive factor like protein 1 (CaERFLP1) of hot pepper (Capsicum annuum L.) interacts in vitro with both GCC and DRE/CRT sequences with different binding affinities: possible biological roles of CaERFLP1 in response to pathogen infection and high salinity conditions in transgenic tobacco plants. Plant Mol. Biol. 55, 61–81.

    Article  PubMed  CAS  Google Scholar 

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

    Article  CAS  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  PubMed  CAS  Google Scholar 

  • Neuwald, A.F., Aravind, L., Spouge, J.L., and Koonin, E.V. (1999). AAA+: a class of chaperone-like ATPases associated with the assembly, operation, and disassembly of protein complexes. Genome Res. 9, 27–43.

    PubMed  CAS  Google Scholar 

  • Park, J.A., Cho, S.K., Kim, J.E., Chung, H.S., Hong, J.P., Hwang B., Hong, C.B., and Kim, W.T. (2003). Isolation of cDNAs differentially expressed in response to drought stress and characterization of the Ca-LEAL1 gene encoding a new family of atypical LEA-LIKE protein homologue in hot pepper (Capsicum annuum L. cv. Pukang). Plant Sci. 165, 471–481.

    Article  CAS  Google Scholar 

  • Park, J.-A., Kim, T.-W., Kim, S.-K., Kim, W.T., and Pai, H.-S. (2005a). Silencing of NbECR encoding a putative enoyl-CoA reductase results in disorganized membrane structure and epidermal cell ablation in Nicotiana benthamiana. FEBS Lett. 579, 4459–4464.

    Article  PubMed  CAS  Google Scholar 

  • Park, J.-A., Ahn, J.-W., Kim, Y.-K., Kim, S.J., Kim, J.-K., Kim, W.T., and Pai, H.-S. (2005b). Retinoblastoma protein regulates cell proliferation, differentiation, and endoreduplication in plants. Plant J. 42, 153–163.

    Article  PubMed  CAS  Google Scholar 

  • Park, S., Rancour, D.M., and Bednarek, S.Y. (2007). Protein domain-domain interactions and requirements for the negative regulation of Arabidopsis CDC48/p97 by the plant ubiquitin regulatory X (UBX) domain-containing protein, PUX1. J. Biol. Chem. 282, 5217–5224.

    Article  PubMed  CAS  Google Scholar 

  • Park, S., Rancour, D.M., and Bednarek, S.Y. (2008). In planta analysis of the cell cycle-dependent localization of AtCDC48A and its critical roles in cell division, expansion and differentiation. Plant Physiol. 148, 246–258.

    Article  PubMed  CAS  Google Scholar 

  • Partridge, J.J., Lopreiato, J.O., Latterich, M., and Indig, F.E. (2003). DNA damage modulates nucleolar interaction of the Werner protein with the AAA ATPase p97/VCP. Mol. Biol. Cell 14, 4221–4229.

    Article  PubMed  CAS  Google Scholar 

  • Rancour, D.M., Dickey, C.E., Park, S., and Bednarek, S.Y. (2002). Characterization of AtCDC48. Evidence for multiple membrane fusion mechanisms at the plane of cell division in plants. Plant Physiol. 130, 1241–1253.

    Article  PubMed  CAS  Google Scholar 

  • Rancour, D.M., Park, S., Knight, S.D., and Bednarek, S.Y. (2004). Plant UBX domain-containing protein 1, PUX1, regulates the oligomeric structure and activity of Arabidopsis CDC48. J. Biol. Chem. 279, 54264–54274.

    Article  PubMed  CAS  Google Scholar 

  • Ratcliff, F., Martin-Hernandez, A.M., and Baulcombe, D.C. (2001). Tobacco rattle virus as a vector for analysis of gene function by silencing. Plant J. 25, 237–245.

    Article  PubMed  CAS  Google Scholar 

  • Richly, H., Rape, M., Braun, S., Rumpf, S., Hoege, C., and Jentsch, S. (2005). A series of ubiquitin binding factors connects CDC48/p97 to substrate multiubiquitination and proteasomal targeting. Cell 120, 73–84.

    Article  PubMed  CAS  Google Scholar 

  • Rienties, I.M., Vink, J., Borst, J.W., Russinova, E., and de Vries, S.C. (2005). The Arabidopsis SERK1 protein interacts with the AAA-ATPase AtCDC48, the 14-3-3 protein GF14λ and the PP2C phosphatase KAPP. Planta 221, 394–405.

    Article  PubMed  CAS  Google Scholar 

  • Romisch, K. (2006). Cdc48 is UBX-linked to ER ubiquitin ligases. Trends Biochem. Sci. 31, 24–25.

    Article  PubMed  Google Scholar 

  • Sarowar, S., Lee, J.-Y., Ahn, E.-R., and Pai, H.-S. (2008). A role of hexokinases in plant resistance to oxidative stress and pathogen infection. J. Plant Biol. 51, 341–346.

    Article  CAS  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  PubMed  CAS  Google Scholar 

  • Waterhouse, P.M., Wang, M.-B., and Lough, T. (2001). Gene silencing as an adaptive defense against viruses. Nature 411, 834–842.

    Article  PubMed  CAS  Google Scholar 

  • Yang, S.W., Kim, D.H., Lee J.J., Chun, Y.J., Lee, J.-H., Kim, Y.J., Chung, I.K., and Kim, W.T. (2003). Expression of the telomeric repeat binding factor gene NgTRF1 is closely coordinated with the cell division program in tobacco BY-2 suspension culture cells. J. Biol. Chem. 278, 21395–21407.

    Article  PubMed  CAS  Google Scholar 

  • Yang, S.W., Kim, S.K., and Kim, W.T. (2004). Perturbation of NgTRF1 expression induces apoptosis-like cell death in tobacco BY-2 cells and implicates NgTRF1 in the control of telomere length and stability. Plant Cell 16, 3370–3385.

    Article  PubMed  CAS  Google Scholar 

  • Ye, Y., Meyer, H.H., and Rapoport, T.A. (2001). The AAA ATPase Cdc48/p97 and its partners transport proteins from the ER into the cytosol. Nature 414, 652–656.

    Article  PubMed  CAS  Google Scholar 

  • Ye, Y., Meyer, H.H., and Rapoport, T.A. (2003). Function of the p97-Ufd1-Npl4 complex in retrotranslocation from the ER to the cytosol: dual recognition of nonubiquitinated polypeptide segments and polyubiquitin chains. J. Cell Biol. 162, 71–84.

    Article  PubMed  CAS  Google Scholar 

  • Yuan, X., Shaw, A., Zhang, X., Kondo, H., Lally, J., Freemont, P.S., and Matthews, S. (2001). Solution structure and interaction surface of the C-terminal domain from p47: a major p97-cofactor involved in SNARE disassembly. J. Mol. Biol. 311, 255–263.

    Article  PubMed  CAS  Google Scholar 

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Correspondence to Woo Taek Kim.

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Bae, H., Choi, S.M., Yang, S.W. et al. Suppression of the ER-localized AAA ATPase NgCDC48 inhibits tobacco growth and development. Mol Cells 28, 57–65 (2009). https://doi.org/10.1007/s10059-009-0101-4

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  • DOI: https://doi.org/10.1007/s10059-009-0101-4

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