, Volume 165, Issue 4, pp 455–468

Structural organization of ultrarapidly frozen barley aleurone cells actively involved in protein secretion

  • D. E. Fernandez
  • L. A. Staehelin


The ultrastructural organization of actively secreting barley (Hordeum vulgare L. cv. Himalaya) aleurone cells was examined using ultrarapid-freezing (<-10 000°C s-1) followed by freeze-fracture and freeze-substitution. Our analysis indicates that much of the evidence supporting a direct pathway from the endoplasmic reticulum (ER) to the plasma membrane (i.e. bypassing the Golgi apparatus) for the secretion of α-amylase (EC may not be valid. Cryofixed ER cisternae show no sign of vesiculation during active α-amylase secretion in gibberellic acid (GA3)-treated cells. At the same time, Golgi complexes are abundant and numerous small vesicles are associated with the edges of the cisternae. Vesicles appear to be involved in the delivery of secretory products to the plasma membrane since depressions containing excess membrane material appear there. Treatment with GA3 also induces changes in the composition of Golgi membranes; most notably, the density of intramembrane particles increases from 2700 μm-2 to 3800 μm-2 because of an increase of particles in the 3–8.5-nm size range. A slight decrease in 9–11-nm particles also occurs. These changes in membrane structure appear to occur as the Golgi complex becomes committed to the processing and packaging of secretory proteins. We suggest that secretory proteins in this tissue are synthesized in the abundant rough ER, packaged in the Golgi apparatus, and transported to the plasma membrane via Golgi-derived secretory vesicles. Mobilization of reserves is also accompanied by dynamic membrane events. Our micrographs show that the surface monolayer of the lipid bodies fuses with the outer leaflet of the bilayer of protein-body membranes during the mobilization of lipid reserves. Following the breakdown of the protein reserves, the protein bodies assume a variety of configurations.

Key words

Aleurone α-Amylase Hordeum (aleurone) Ultrarapid freezing 



endoplasmic reticulum


gibberellic acid






Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Branton, D.S., Bullivant, S., Gilula, N.G., Karnovsky, M.J., Moor, H., Mühlethaler, K., Northcote, D., Packer, L., Satir, P., Speth, V., Staehelin, L.A., Steere, R.L., Weinstein, R.S. (1975) Freeze-etching nomenclature. Science 190, 54–56Google Scholar
  2. Buckhout, T.J., Morré, D.J. (1982) An ultrastructural analysis of aleurone cells: lamellar bodies in aleurone layers of wheat grains during germination. Bot. Gaz. 143, 156–163Google Scholar
  3. Buttrose, M. (1971) Ultrastructure of barley aleurone cells as shown by freeze-etching. Planta 96, 13–26Google Scholar
  4. Chen, R., Jones, R.L. (1974) Studies on the release of barley aleurone cell proteins: autoradiography. Planta 119, 207–220Google Scholar
  5. Chrispeels, M.J. (1976) Biosynthesis, intracellular transport and secretion of extracellular molecules. Annu. Rev. Plant Physiol. 27, 19–38Google Scholar
  6. Chrispeels, M.J. (1983) The Golgi apparatus mediates the transport of phytohemagglutinin to the protein bodies in bean cotyledons. Planta 158, 140–151Google Scholar
  7. Chrispeels, M.J., Varner, J.E. (1967) Gibberellic acid-enhanced synthesis and release of α-amylase and ribonuclease by isolated barley aleurone layers. Plant Physiol. 42, 398–406Google Scholar
  8. Colborne, A.J., Morris, G., Laidman, D.L. (1976) Formation of ER in aleurone cells of germinating wheat: an ultrastructural study. J. Exp. Bot. 27, 759–767Google Scholar
  9. Farquhar, M.G., Palade, G.E. (1981) The Golgi apparatus (complex)-(1954–1981)-from artifact to center stage. J. Cell Biol. 91, 77s-103sGoogle Scholar
  10. Firn, R.D. (1975) On the secretion of α-amylase by barley aleurone layers after incubation in gibberellic acid. Planta 125, 227–233Google Scholar
  11. Gardiner, M., Chrispeels, M.J. (1975) Involvement of the Golgi apparatus in the synthesis and secretion of hydroxyprolinerich cell wall glycoproteins. Plant Physiol. 55, 536–541Google Scholar
  12. Gibson, P.A., Paleg, L.G. (1976) Purification of gibberellic acid-induced lysosomes from wheat aleurone cells. J. Cell Sci. 22, 413–425Google Scholar
  13. Gilkey, J., Staehelin, L.A. (1985) Advances in ultrarapid freezing for the preservation of cellular ultrastructure. J. Electron Microsc. Tech., in pressGoogle Scholar
  14. Gram, N.H. (1982) The ultrastructure of germinating barley seeds. I. Changes in the scutellum and the aleurone layer in Nordal barley. Carlsberg Res. Commun. 47, 143–162Google Scholar
  15. Heuser, J.E., Reese, T.S., Dennis, M.J., Jan, Y., Jan, L., Evans, L. (1979) Synaptic vesicle exocytosis captured by quick freezing and correlated with quantal transmitter release. J. Cell. Biol. 81, 275–300Google Scholar
  16. Hooley, R. (1982) Protoplasts isolated from aleurone layers of wild oat (Avena fatua L.) exhibit the classic response to gibberellic acid. Planta 154, 29–40Google Scholar
  17. Jacobsen, J.V., Higgins, T.J.V. (1982) Characterization of the α-amylases synthesized by aleurone layers of Himalaya barley in response to gibberellic acid. Plant Physiol. 70, 1647–1653Google Scholar
  18. Jacobsen, J.V., Knox, R.B., Pyliotis, N.A. (1971) The structure and composition of aleurone grains in the barley aleurone layer. Planta 101, 189–209Google Scholar
  19. Jacobsen, J.V., Zwar, J.A., Chandler, P.M. (1985) Gibberellic-acid-responsive protoplasts from mature aleurone of Himalaya barley. Planta 163, 430–439Google Scholar
  20. Johnson, K.D., Chrispeels, M.J. (1973) Regulation of pentosan biosynthesis in barley aleurone tissue by gibberellic acid. Planta 111, 353–364Google Scholar
  21. Jones, R.L. (1969a) Gibberellic acid and the fine structure of barley aleurone cells. I. Changes during the lag-phase of α-amylase synthesis. Planta 87, 119–133Google Scholar
  22. Jones, R.L. (1969b) Gibberellic acid and the fine structure of barley aleurone cells. II. Changes during synthesis and secretion of α-amylase. Planta 88, 73–86Google Scholar
  23. Jones, R.L. (1980) Quantitative and qualitative changes in the endoplasmic reticulum of barley aleurone layers. Planta 150, 70–81Google Scholar
  24. Jones, R.L., Carbonell, J. (1984) Regulation of the synthesis of barley aleurone α-amylase by gibberellic acid and calcium ions. Plant Physiol. 76, 213–218Google Scholar
  25. Jones, R.L., Chen, P. (1976) Immunohistochemical localization of α-amylase in barley aleurone cells. J. Cell Sci. 20, 183–198Google Scholar
  26. Jones, R.L., Jacobsen, J.V. (1982) The role of the endoplasmic reticulum in the synthesis and transport of α-amylase in barley aleurone layers. Planta 156, 421–432Google Scholar
  27. Jones, R.L., Jacobsen, J.V. (1983) Calcium regulation of the secretion of α-amylase isoenzymes and other proteins from barley aleurone layers. Planta 158, 1–9Google Scholar
  28. Jones, R.L., Price, J.M. (1970) Gibberellic acid and the fine structure of barley aleurone cells. III. Vacuolation of aleurone cells during the phase of ribonuclease release. Planta 94, 191–202Google Scholar
  29. Jones, R.L., Varner, J.E. (1967) Bioassay of gibberellins. Planta 72, 155–161Google Scholar
  30. Kawasaki, S., Sato, S. (1979) Isolation of the Golgi apparatus from suspension cultured tobacco cells and preliminary observations on the intracellular transport of extensin-precursor. Bot. Mag. (Tokyo) 92, 305–314Google Scholar
  31. Locy, R., Kende, H. (1978) The mode of secretion of α-amylase in barley aleurone layers. Planta 143, 89–99Google Scholar
  32. Mersey, B., McCully, M.E. (1978) Monitoring of the course of fixation of plant cells. J. Microsc. 114, 49–76Google Scholar
  33. Mollenhauer, H.H., Morré, D.J. (1980) The Golgi apparatus. In: The biochemistry of plants, vol. I: The plant cell, pp. 438–489, Tolbert, N.E., ed. Academic Press, New YorkGoogle Scholar
  34. Morré, D.J., Kartenbeck, J., Franke, W.W. (1979) Membrane flow and interconversions among endomembranes. Biochim. Biophys. Acta 559, 71–152Google Scholar
  35. Paleg, L., Hyde, B. (1964) Physiological effects of gibberellic acid. VII. Electron microscopy of barley aleurone cells. Plant Physiol. 39, 673–680Google Scholar
  36. Platt-Aloia, K.A., Thomson, W.W. (1982) Freeze-fracture of intact plant tissues. Stain Technol. 57, 327–334Google Scholar
  37. Plattner, H., Bachmann, L. (1982) Cryofixation: a tool in biological ultrastructural research. Int. Rev. Cytol. 97, 237–304Google Scholar
  38. Roberts, K., Gurney-Smith, M., Hills, G.J. (1972) Structure, composition and morphogenesis of the cell wall of Chlamydomonas reinhardi. I. Ultrastructure and preliminary chemical analysis. J. Ultrastruct. Res. 40, 599–613Google Scholar
  39. Rothman, J.E. (1981) The Golgi apparatus: two organelles in tandem. Science 213, 1212–1219Google Scholar
  40. Sato, T. (1968) A modified method for lead staining of thin sections. J. Electron Microsc. 17, 158Google Scholar
  41. Spurr, A.R. (1969) A low-viscosity epoxy resin embedding medium for electron microscopy. J. Ultrastruct. Res. 26, 31–43Google Scholar
  42. Staehelin, L.A. (1976) Reversible particle movements associated with unstacking and restacking of chloroplast membranes. J. Cell Biol. 71, 136–158Google Scholar
  43. Staehelin, L.A., Kiermayer, O. (1970) Membrane differentiation in the Golgi complex of Micrasterias denticulata Breb. visualized by freeze-etching. J. Cell Sci. 7, 787–792Google Scholar
  44. Swift, J.G., O'Brien, T.P. (1972) The fine structure of the wheat scutellum during germination. Aust. J. Biol. Sci. 25, 469–486Google Scholar
  45. Varner, J.E., Ho, D.T. (1976) The role of hormones in the integration of seedling growth. In: The moleular biology of hormone action, pp. 173–194, Papaconstantinou, J., ed. Academic Press, New York LondonGoogle Scholar
  46. Vigil, E.L., Ruddat, M. (1973) Effect of giberellic acid and actinomycin D on the formation and distribution of rough endoplasmic reticulum in barley aleurone cells. Plant Physiol. 51, 549–558Google Scholar
  47. Volkmann, D. (1983) A freeze-fracture study on the differentiation of Golgi and plasma membranes in plant cells. Eur. J. Cell Biol. 30, 258–265Google Scholar
  48. Yatsu, L.Y., Jacks, T.J. (1972) Spherosome membranes: half unit-membranes. Plant Physiol. 49, 937–943Google Scholar

Copyright information

© Springer-Verlag 1985

Authors and Affiliations

  • D. E. Fernandez
    • 1
  • L. A. Staehelin
    • 1
  1. 1.Department of Molecular, Cellular and Developmental BiologyUniversity of ColoradoBoulderUSA

Personalised recommendations