Paramecium trichocysts isolated with their membranes are stable in the presence of millimolar Ca2+

Author:

LIMA OSCAR1,GULIK-KRZYWICKI TADEUSZ2,SPERLING LINDA2

Affiliation:

1. Centre de Génétique Moléculaire, Associated with the Université Pierre et Marie Curie, CNRS, Gif-sur-Yvette, 91190 France; Laboratoire de Génétique Physiologique, Université Paris XI, Batîment 400, 91405 Orsay Cedex, France

2. Centre de Génétique Moléculaire, Associated with the Université Pierre et Marie Curie, CNRS, Gif-sur-Yvette, 91190 France

Abstract

We have developed a simple and rapid procedure for the isolation of a pure fraction of Paramecium trichocysts (mature secretory vesicles) with their membranes. Since in wild-type Paramecium cells essentially all trichocysts are docked at pre-formed cortical sites, trichocysts were isolated from cells in which functional trichocysts remain free in the cytoplasm owing to a mutation, tam6, that affects the docking site. Examination of the preparations by freeze-fracture electron microscopy confirms the presence of the membranes. The distribution of particles in the membranes of the isolated trichocysts and in the membranes of wild-type trichocysts in situ are nearly identical and this argues against any rearrangement of the membranes during the isolation procedure. Although the trichocyst matrix undergoes a dramatic structural transition in the presence of Ca2+ and water (matrix expansion), the isolated vesicles with intact membranes are perfectly stable in the presence of millimolar free Ca2+. This result supports a chronology in which the first step in exocytosis is membrane fusion, the swelling of vesicle contents occurring only afterwards, once the contents come into contact with the water and Ca2+ of the external medium. The role of swelling would then be to help disperse, propel or otherwise empty the contents of the vesicle outside the cell.

Publisher

The Company of Biologists

Subject

Cell Biology

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