21st Century Guidebook to Fungi, SECOND EDITION, by David Moore, Geoffrey D. Robson and Anthony P. J. Trinci

Table of Contents

1. Chapter 4: Hyphal cell biology and growth on solid substrates
2. Chapter 5: Fungal cell biology
   1. Mechanisms of mycelial growth
   2. The fungus as a model eukaryote
   3. The essentials of cell structure
   4. Subcellular components of eukaryotic cells
   5. The nucleus
   6. The nucleolus and nuclear import and export
   7. Nuclear genetics
   8. Mitotic nuclear division
   9. Meiotic nuclear division
   10. mRNA translation and protein sorting
   11. The endomembrane systems
   12. Cytoskeletal systems
   13. Molecular motors
   14. Plasma membrane and signalling pathways
   15. Fungal cell wall
   16. Cell biology of the hyphal apex
   17. Hyphal fusions and mycelial interconnections
   18. Cytokinesis and septation
   19. Yeast-mycelial dimorphism
   20. Chapter 5 References and further reading about the cell cycle
   21. Buy a PDF of Chapter 5
3. Chapter 6: Structure and synthesis of fungal cell walls

5.15 Fungal cell wall

The fungal wall is a sophisticated cell organelle. It defines the volumetric shape of the cell, provides osmotic and physical protection and, together with the plasma membrane and periplasmic space, influences and regulates the influx of materials into the cell. However, it is also able to control the environment in the immediate external vicinity of the cell membrane, and it represents the interface between the organism and the outside world.

This is an active interface, since the interaction of the organism and the outside world (and the latter will include other cells) is subject to modulation and modification. The fungal cell wall is metabolically active, interactions between its components occur to give rise to the mature cell wall structure. So the wall must be understood to be a dynamic structure which is subject to modification at various times to suit various functions. Besides enclosing and supporting the cytoplasm, those functions include selective permeability, as a support for immobilised enzymes and cell–cell recognition and adhesion.

The wall is a multilayered complex of polysaccharides, glycoproteins and proteins. The polysaccharides are glucans and mannans and include some very complex polysaccharides (like gluco-galacto-mannans). In hyphae the major component of the wall, and certainly the most important for its structural integrity, is chitin (a linear polymer of N-acetylglucosamine) though this is frequently cross linked to other wall constituents, particularly a β(1→3)-glucan, the terminal reducing residue of a chitin chain being attached to the non-reducing end of a β(1→3)-glucan chain by a (1→4) linkage.

Removal of the outer wall layers with lytic enzymes has revealed the architecture of the inner chitin wall to be composed of microfibrils formed by the aggregation of the chitin polymers by hydrogen bonding. The chitin inner wall is cross-linked to the outer β-glucan components and forms a major structural component of the walls of most true fungi. Synthesis of the cell wall occurs at the outer surface of the plasma membrane of the growing hyphal tip. Chitin synthase is the enzyme that catalyses formation of chitin from the precursor UDP-N-acetylglucosamine. Chitin synthase adds two molecules of UDP-N-acetylglucosamine (UDPGlcNAc) to the existing chitin chain in the reaction:

(GlcNAc)_n + 2UDPGlcNAc → (GlcNAc)_n+2 + 2UDP

It appears as an inactive zymogen requiring activation by cleavage of a peptide by an endogenous protease to generate the active enzyme (Robson, 1999).

Phylogenetic analysis has revealed seven classes of chitin synthases divided into two families. Class I, II and IV genes make up the first family and are present in all fungi, whereas classes III, V, VI and VII (the second family) are specific to filamentous fungi and certain dimorphic species. The number of putative chitin synthase genes (as opposed to classes of chitin synthase) within each species also varies, with three in Saccharomyces cerevisiae, four in Candida albicans, seven in Wangiella dermatitidis and Neurospora crassa and eight each in Aspergillus nidulans, Aspergillus fumigatus and Cryptococcus neoformans (Rogg et al., 2012). For the most part, the precise functions of the individual genes remain to be determined; however, three chitin synthase genes, CHSI, CHSII and CHSIII that have been cloned from budding yeast, Saccharomyces cerevisiae, serve different functions in the cell:

Three chitin synthase genes, CHSI, CHSII and CHSIII have been cloned from budding yeast, Saccharomyces cerevisiae, and shown to serve different functions in the cell:

• CHSI acts as a repair enzyme and is involved in synthesising chitin at the point where the daughter and mother cells separate;
• CHS2 is involved in septum formation;
• CHS3 is involved in chitin synthesis of the main cell wall.

Like chitin, β(1→3)-glucan is synthesised by a membrane-associated β(1→3)-glucan synthase which utilises UDP-glucose as its monomeric substrate, inserting glucose into the β-glucan chains. The β(1→3)-glucan synthase activity is found both in the membrane and cytoplasmic fractions of fungal mycelia and is stimulated by GTP. Although genes encoding a β-glucan synthase have been isolated from a number of fungi, it is not clear whether a gene family for β(1→3)-glucan synthase exists in fungi as it does for chitin synthase (Teparić & Mrša, 2013).

The fungal cell wall represents an attractive target for pharmacologic inhibition, as many of the components are fungal-specific. Though targeted inhibition of β-glucan synthesis has been used to treat some fungal infections, for example Pneumocystis pneumonia (Schmatz et al. 1990), the ability of the cell wall to dynamically compensate via the cell wall integrity pathway limits their usefulness. To date, chitin synthesis inhibitors have not been successfully deployed in the clinical setting.

Two classes of nucleoside peptides, the polyoxins and the nikkomycins, act as potent and specific inhibitors of chitin synthesis, competing as analogues of UDP-N-acetylglucosamine. However, their toxicity to animals (in which they act as analogues of other UDP-linked metabolites) has prevented them from being exploited as clinically-useful antifungal agents, and their use as agrochemical fungicides has been hampered by the rapid emergence of resistant fungal strains. Another class of naturally occurring antibiotics, the echinocandins, are specific inhibitors of β-glucan biosynthesis. Semisynthetic echinocandins which have a broad antifungal spectrum and high potency and can be taken orally show promise in the treatment of human fungal infections.

Although during normal apical extension of the hypha the incorporation of newly synthesised chitin is limited to the hyphal apex, there is evidence that inactivated chitin synthases are widely distributed in the plasma membrane, indeed inactivated chitin synthase activity appears to be an intrinsic property of the plasma membrane, the enzyme being activated in some way specifically at the hyphal apex and at sites where branch formation is initiated.

Chitin is important at particular sites in yeast walls, although the major structural component in these organisms is a fibrillar inner layer of β-glucan. The glucan has secreted mannoproteins attached to it. Mannoproteins play an essential role in cell wall organisation, and there is evidence for the formation of covalent bonds between these molecules and the structural polymers (glucans and chitin) outside the plasma membrane. This makes the point that the protein components of fungal walls are of considerable importance, too.

Some of the proteins identified in walls have enzymic activities associated with them; these include a-glucosidases and β-glucosidases, enolase, and alkaline phosphatase. Other obvious components are proteins involved in cell-to-cell recognition like the products of mating type factors; cell-to-cell adhesion during yeast mating depends on interaction of two glycoproteins inserted into the outer coat of the cell wall, which are the gene products of components of the mating type factors and are first located in the plasma membranes with GPI anchors.

The outer surface of many fungal walls is usually layered with proteins that modify the biophysical properties of the wall surface appropriately to the environment. Hydrophobic surfaces (like spores and aerial mycelium) have rodlets of the protein hydrophobin as the outermost rodlet layer (see Section 6.8; CLICK HERE to view now). Some GPI-anchored wall proteins span the wall with extended glycosylated polypeptides protruding into the surrounding medium and providing the wall with hydrophilic and even adhesive surfaces (Bayry et al., 2012).

Fungal walls are remarkably variable in their detailed aspects of structure. Different strains of the same species may exhibit differences in overall composition of the wall and in the polysaccharide and polypeptide structures the wall contains. Indeed, a single colony may have different wall structures in its different regions (aerial structures, surface mycelium, submerged mycelium, etc.). In the face of such differences it is not feasible to give an exact description of a typical fungal wall (there’s no such thing), but it is possible to describe the conceptual framework of the fungal wall. In brief, the fungal wall concept is this:

• the main structural substance of the wall is provided by polysaccharides, mostly glucan, and in filamentous forms the shape-determining component is chitin;
• various polysaccharide components are linked together by hydrogen bonding and by covalent bonds;
• a variety of proteins/glycoproteins contribute to wall function, some of these are structural, some are enzymic, and some vary the biological and biophysical characteristics of the outer surface of the wall;
• proteins may be anchored in the plasma membrane, covalently bonded to wall polysaccharides or more loosely associated with the wall;
• the wall is a dynamic structure which is modified (a) as it matures, and/or (b) as part of hyphal differentiation, and/or (c) on a short-term basis to react to changes in physical and physiological conditions.

Finally, by definition, the wall is extracellular, its entire structure lies outside the plasma membrane, so all additions to its structure must be externalised through the membrane before the wall can be restructured and some of the chemical reactions which link wall components together are extracellular reactions. Fungal wall synthesis will be dealt with in detail in Chapter 6.

Updated July, 2019