https://mmbr.asm.org/content/mmbr/43/2/117.full.pdf

Biosynthesis of cell walls of fungi.

Cell wall synthesis in yeast protoplasts.

In the presence of MgSO4 as osmotic stabilizer, nucleated protoplasts of Schizophyllum commune developed a large vacuole and could be isolated on the basis of their low buoyant density. All these protoplasts were capable of wall regeneration and about 50 percent reverted to the hyphal mode of growth in liquid medium. The kinetics of the formation of three main cell-wall components, S-glucan (α-1,3-glucan), R-glucan (β-1,3, β-1,6-glucan) and chitin were studied from the onset of regeneration. S-glucan and chitin accumulation as well as RNA and protein synthesis started simultaneously after a short lag, but R-glucan formation was delayed. The reversion to hyphal tubes only began after several hours of rapid R-glucan synthesis. Cycloheximide (0.5 μg/ml), inhibiting protein synthesis by 98% inhibited the formation of R-glucan and the reversion to hyphal growth but the formation of chitin and S-glucan did start and continued seemingly unimpaired for several hours. This indicates that the enzymes responsible for the synthesis of S-glucan and chitin remained intact during protoplast preparation. Polyoxin D inhibited both the synthesis of chitin and R-glucan and also the reversion to hyphal growth. However, the synthesis of S-glucan was not suppressed. These inhibitor studies as well as the kinetics of R-glucan formation during normal regeneration suggest that the synthesis of R-glucan is required for the initiation of hyphal morphogenesis.

Chemical analysis of cell wall regeneration and reversion of protoplasts from Schizophyllum commune.

Publisher Summary Cytological studies of the cells of fungi have revealed the presence of a cell wall surrounding the fungal protoplasm Recent electron photomicrographs indicate that the wall is composed of an outer dense layer about 005μ thick and a less dense layer of about 03μ The inner part of the wall may in turn be subdivided into about three layers Cell walls are composed of polymers of glucose and mannose, more rarely galactose, with smaller amounts of protein, lipid, and chitin A number of workers have isolated the cell walls by mechanical disintegration of the fungal cells and subsequent centrifugation and washing Selective digestion of the cell wall by enzyme systems has been effected in several species of yeasts and filamentous fungi, resulting in the liberation of the fungal protoplasm in an intact and spherical form Electron microscopy observations on ultrathin sections of these bodies did not show any structure corresponding to the wall of the cell The occurrence of forms of fungi resembling protoplasts under particular cultural conditions, or under abnormal conditions, is well known From current studies, it would appear that unbalanced growth can occur Under many conditions, suppression of wall synthesis may be effected without concomitant inhibition of protoplasmic growth A variety of lytic organisms, mainly bacteria and actinomycetes of the soil, are able to excrete all kinds of cell wall digestive enzymes into the surrounding media

Chapter XXIV Production and Use of Fungal Protoplasts

Host Parasite Specificity and Pathogenesis R. Gupta, K.G. Mukerji. Yeast Species for Biocontrol of Apple Postharvest Diseases: An Encouraging Case of Study for Practical Use M. Haissam Jijakli, et al. Biological Control Agents of Canola and Rapeseed Diseases - Status and Practical Approaches S.M. Boyetchko. Innovative Applications of Microbial Agents for Biological Weed Control S.M. Boyetchko. Bacteria as Biocontrol Agents of Insects S. Kaur, K.G. Mukerji. Biocontrol-Plant Growth Promoting Rhizobacteria: Mechanism of Action K.V. B.R. Tilak, et al. Mycorrhiza in Control of Plant Pathogens: Molecular Approaches K.G. Mukerji. Biological Control of Bacterial Plant Diseases S. Kaur, K.G. Mukerji. Protoplast Fusion in Disease Control S. Mukerji, K.G. Mukerji. Role of Tissue Culture in Plant Disease Control P.S. Srivastava, et al. Genetic Manipulation of Antagonistic Fusarium spp Q. Migheli, et al. The Application of lux-Gene Technology in the Control of Soil-Borne Diseases D. White, et al. Index.

Biotechnological approaches in biocontrol of plant pathogens.

SUMMARY: The process of regeneration of Fusarium culmorum protoplasts, obtained by means of strepzyme RA, has been followed. Protoplast regeneration started with the formation of cellular aggregates originating a mass of globular forms which later gave rise to the formation of a pseudomycelial form. Regeneration of protoplasts was formed in three different ways: (a) by means of a chain of yeast-like forms and later originating a germ tube; (b) by direct formation of the germ tube from the protoplast; (c) through a complicated process with the formation of various globular forms and giving rise to the formation of the germ tube at the end. Maximum regeneration, about 80 % of the protoplasts, was found in the presence of 2% (w/v) sucrose (or 1% (w/v) glucose + 1% (w/v) fructose) + 10% (w/v) sorbose + 0·2% agar + the mineral salts of the Czapek medium. No other sugars were able to substitute for sucrose and sorbose. Agar was the best substance for regeneration; gelatin produced an inhibition. Regeneration was greater under acid conditions, alkaline pH values interfering with the phenomenon. The number of nuclei/protoplast varied from 1 to 4, the lack of them in some spherical forms perhaps being the cause of failure to regenerate. No differences were found in the regenerative process as between protoplasts obtained by the use of snail or by microbial lytic enzymes.

/pdf/regeneration-of-mycelial-protoplasts-of-fusarium-culmorum-28xf3ptm4l.pdf

J. R. Villanueva

Papers

Regeneration of Mycelial Protoplasts of Fusarium culmorum