In situlocalization of β‐glucans in the cell wall ofSchizosaccharomyces pombe

Abstract

The chemical composition of the cell wall of Sz. pombe is known as b-1,3-glucan, b-1,6-glucan, a-1,3-glucan and a-galactomannan; however, the three-dimensional interactions of those macromolecules have not yet been clarified. Transmission electron microscopy reveals a three-layered structure: the outer layer is electron-dense, the adjacent layer is less dense, and the third layer bordering the cell membrane is dense. In intact cells of Sz. pombe, the high-resolution scanning electron microscope reveals a surface completely filled with a-galactomannan particles. To better understand the organization of the cell wall and to complement our previous studies, we set out to locate the three different types of b-glucan by immuno-electron microscopy. Our results suggest that the less dense layer of the cell wall contains mainly b-1,6-branched b-1,3-glucan. Occasionally a line of gold particles can be seen, labelling fine filaments radiating from the cell membrane to the a-galactomannan layer, suggesting that some of the radial filaments contain b-1,6-branched b-1,3-glucan. b-1,6-glucan is preferentially located underneath the a-galactomannan layer. Linear b-1,3-glucan is exclusively located in the primary septum of dividing cells. b-1,6-glucan only labels the secondary septum and does not co-localize with linear b-1,3-glucan, while b-1,6-branched b-1,3-glucan is present in both septa. Linear b-1,3-glucan is present from early stages of septum formation and persists until the septum is completely formed; then just before cell division the label disappears. From these results we suggest that linear b-1,3-glucan is involved in septum formation and perhaps the separation of the two daughter cells. In addition, we frequently found b-1,6-glucan label on the Golgi apparatus, on small vesicles and underneath the cell membrane. These results give fresh evidence for the hypothesis that b-1,6-glucan is synthesized in the endoplasmic reticulum-Golgi system and exported to the cell membrane.

Cell Wall β-(1,6)-Glucan of Saccharomyces cerevisiae

“…In comparison, the ␤-(1,6)-glucan molecule that we characterized has a slightly higher DP than the one isolated earlier by Manners et al . 4 The unique recombinant endo-␤-(1,6)-glucanase from T. harzianum used to date releases mainly G 2 and G 3 and some G 4 and G 5 from pustulan (28).…”

“…However, the exact structure of the ␤-(1,6)-glucan and the mode of biosynthesis of this polymer are largely unknown. In S. pombe, immunodetection studies suggested that synthesis of this polymer backbone begins in the endoplasmic reticulum, with extension occurring in the Golgi (4) and final processing at the plasma membrane. In S. cerevisiae, Montijn and co-workers (5), by immunogold labeling, detected ␤-(1,6)-glucan at the plasma membrane, suggesting that the synthesis takes place largely at the cell surface.…”

The Actomyosin Ring Recruits Early Secretory Compartments to the Division Site in Fission Yeast

“…As septum biogenesis proceeds, actin reorganizes to clusters of patches on both sides of the septum (Marks and Hyams, 1985). Secondary septa are then formed on either side of the primary one that is eventually dissolved resulting in cell separation (Humbel et al, 2001).…”

“…As septum biogenesis proceeds, actin reorganizes to clusters of patches on both sides of the septum (Marks and Hyams, 1985). Secondary septa are then formed on either side of the primary one that is eventually dissolved resulting in cell separation (Humbel et al, 2001).Formation and remodeling of the cell wall depend on action of enzymes such as the (1,3)-␤-glucan-synthase and the endo-␤-1,3-glucanase that are involved in organizing the (1,3)-␤-glucan, the major constituent of the cell wall (Ribas et al, 1991;Martin-Cuadrado et al, 2003). These proteins have been shown to localize to the sites of active growth and their localization depends on secretion (Cortes et al, 2002; MartinCuadrado et al, 2003).…”

A Role for the Septation Initiation Network in Septum Assembly Revealed by Genetic Analysis of sid2-250 Suppressors

“…After this, each daughter builds cell wall material on its own side of the primary septum, leading to the assembly of a secondary septum. The primary septum contains mainly 1,3-b-glucan and the secondary septum contains 1,3-a-glucan, 1,6-branched 1,3-b-glucan, and galactomannans (Humbel et al 2001;Sugawara et al 2003). Recently, several enzymes involved in synthesis of 1,3-b-glucan and 1,3-a-glucan have been identified, and all of them are large integral membrane proteins.…”