A comparative structural analysis of direct and indirect shoot ...

58 M. Volgger et al.
Fig. 6 a–h After 24 h of incubation: adaptation of root hairs to
plasmolytic conditions. The left column shows whole roots and root
hairs in increasing concentrations of mannitol, dark field mode; the
right column shows single root hairs under these conditions in higher
magnification, bright field mode. Bar left column 30 μm; bar right
column 10 μm
Organisation of the cytoplasm and deposition
of wall material
In root hairs of Tradescantia fluminensis, Schröter and
Sievers (1971) observed differences in cellular organisation
after 3 h in 0.05 molar glucose solutions and a cessation of
growth in existing hairs.
Our results of wheat root hairs show that the cytoarchitecture
is not influenced dramatically during osmotic
changes; even in plasmolysed root hairs, the clear zone
and polar organisation of the cells are maintained for a long
time independent of the osmotic value of the medium. In
addition, the deposition of new cell wall is continuous
during all stages of plasmolysis. It is secreted into the
emerging plasmolytic space between the cell wall and the
protoplast. Obviously, the tip-focussed gradient of exocytotic
vesicles and the continuing organelle motility allow
for a constant supply of wall components and for continued
cell wall synthesis. In plants, disrupted cellulose synthase
may form callose (Brett 2000). Depending of the speed of
plasmolysis, wall depositions appear dispersed or as solid
rings. They remain in place and become especially visible
after renewed plasmolysis.
This new cell wall can be observed by autofluorescence
or after callose staining with aniline blue just exterior of the
plasma membrane, and it appears brighter than within the
plasmolytic space. Electron micrographs of cell walls built
under osmotic stress show a rather loose consistency and
confirm the deposition of mainly pectins and hemicelluloses
in moss protonemata (Schnepf et al. 1986) and in root
hairs of T. fluminensis (Schröter and Sievers 1971). In root
hairs of Zea mays and T. aestivum, also callose depositions
have been observed after turgor reduction (Lerch 1960). In
Tradescantia virginiana leaf epidermal cells, we also found
striking callose depositions at the plasmolysed protoplast,
the inner side of the cell wall (Lang et al. 2004).
Plasmolysis seems to be a membrane-driven process which
still takes place after disruption of actin microfilaments with
cytochalasin B (M Volgger, data not shown) and, in onion
epidermis cells, plasmolysis and Hechtian strand formation was
still observed after cytoskeleton elements had been depolymerized
(Lang-Pauluzzi and Gunning 2000). In osmotically
stressed cells, additional osmocytotic vesicles are formed to
reduce membrane surface area of shrinking protoplasts
(Oparka et al. 1990; Oparkaetal.1996). Interestingly, these
authors demonstrated that osmocytotic vesicles were not
reused in deplasmolysis of onion epidermal cells.
Formation and attachment of Hechtian strands
In strong plasmolysis, Hechtian strands form between the
plasma membrane and the cell wall of plant tissue (Hecht 1912;
Attree and Sheffield 1985;Oparka1994; Lang-Pauluzzi 2000;
Lang et al. 2004), and we show the strands here after FM1-34
and DiOC 6 (3) staining in a tip growing system, i.e. root hairs.
Hechtian strands reached right towards the newly built wall at
the clear zone, suggesting that the anchoring sites are readily
incorporated at the very tip. The strands clearly form without
plasmodesmata (Pont-Lezica et al. 1993) and thus, the
initiation sites of Hechtian strands within the cell wall have