Distribution of chitin in the yeast cell wall. An ultrastructural and chemical study.

Abstract

The distribution of chitin in S. cerevisiae primary septa and cell walls was studied with 3 methods: EM of colloidal gold particles coated either with wheat germ agglutinin or with 1 of 2 different chitinases, fluorescence microscopy with fluorescein isothiocyanate derivatives of the same markers, and enzymatic treatments of [14C]glucosamine-labeled cells. The septa were uniformly and heavily labeled with the gold-attached markers, an indication that chitin was evenly distributed throughout. To study the localization of chitin in lateral walls, alkali-extracted cell ghosts were used. Observations by electron and fluorescence microscopy suggest that lectin-binding material is uniformly distributed over the whole cell ghost wall. This material also appears to be chitin, on the basis of the analysis of the products obtained after treatment of 14C-labeled cell ghosts with lytic enzymes. The chitin of lateral walls can be specifically removed by treatment with .beta.-(1 .fwdarw. 6)-glucanase containing a slight amount of chitinase. During this incubation, .apprx. 7% of the total radioactivity is solubilized, about the same amount liberated when lateral walls of cell ghosts are completely digested with snail glucanase to yield primary septa. The remaining chitin, i.e., > 90% of the total, is in the septa. The facilitation of chitin removal from the cell wall by .beta.-(1 .fwdarw. 6)-glucanase indicates a strong association between chitin and .beta.-(1 .fwdarw. 6)-glucan. Covalent linkages between the 2 polysaccharides were not detected but cannot be excluded.