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Proteinase-inhibitor synthesis in tomato plants: Evidence for extracellular deposition in roots through the secretory pathway

Abstract

The cellular and subcellular localization of proteinase Inhibitors I and II proteins, synthesized in transgenic tomato (Lycopersicon esculentum L.) plants from chimeric genes regulated by the 35S promoter, was investigated by immunocytochemical techniques. Newly synthesized inhibitor proteins were deposited in the cell vacuoles as in wild-type plants, but were also secreted into the cell walls of outer epidermal and secretory cells of the root cap. The Na ionophore monensin increased the levels of proteinase inhibitors found in rough endoplasmic reticulum, Golgi cisternae and in the cell walls of transgenic plants, supporting a role for the secretory pathway in the sorting and targeting of Inhibitor I and II proteins. The two inhibitor proteins were detected by Western-blot analysis in water-washes obtained from roots of transgenic tomato seedlings, confirming their extracellular presence. Wild-type tomato plants exhibited the presence of Inhibitor I and II proteins in the external cell walls, using silver-enhanced immunogold labelling, but not by Western-blot analysis. The extracellular Inhibitor I from transgenic plant roots migrated in electrophoretic gels with a slightly different apparent mass than the Inhibitor I isolated from tomato leaf vacuoles, indicating that specific structural features of this inhibitor protein have been altered during or after extracellular deposition. The presence of extracellular inhibitors in roots may help provide protection for the growing meristems against insects or microorganisms present in the soil.

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Abbreviations

CaMV:

cauliflower mosaic virus

TEM:

transmission electron microscope

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Author information

Correspondence to Clarence A. Ryan.

Additional information

Transmission electron microscopy was performed at the Electron Microscopy Center (EMC) of Washington State University. The authors thank the EMC staff for their technical advice and collaboration. We also thank Greg Wichelns for growing our plants and Greg Pearce, Scott Johnson, and Martha L. Orozco for their advice and technical help. The work was supported in part by the Washington State College of Agriculture and Home Economics Project No. 1791 and National Science Foundation grants Nos. DCB-8702538 and DCB-8608594.

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Narváez-Vásquez, J., Franceschi, V.R. & Ryan, C.A. Proteinase-inhibitor synthesis in tomato plants: Evidence for extracellular deposition in roots through the secretory pathway. Planta 189, 257–266 (1993). https://doi.org/10.1007/BF00195085

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Key words

  • Lycopersicon (proteinase inhibitors)
  • Protein targeting
  • Secretory pathway
  • Transgenic plant