Cell Wall

1.2.1 Composition and Structure

The tubular form of the hypha is an ideal structure for forcing entry into living tissues, for extending through soil or for growing erect to produce propagules and disseminate them into the air. For carrying out these functions, the hypha must generate enormous turgor pressure and have strong cell walls to contain it. If the cell wall is digested by microbial cell wall lytic enzymes, round protoplasts are released. This suggests that the tubular shape of the hypha is due to the rigid cell wall. The nature of the cell wall polymers providing rigidity to hypha has been studied by extraction with hot acid or alkali and characterization of the solubilized and insoluble material and by successive digestion with specific polymer-degrading enzymes (chitinase, cellulase, laminarinase) and monitoring the appearance of shadowed preparations of hyphae by electron microscopy (Burnett, 1976). The cell wall is composed of polysaccharides and proteins and the major polysaccharide components are glucan, chitin and chitosan. Some fungi belonging to Straminipila (see Appendix) have cellulose also.

Current findings suggest that the fungal cell wall is a layered structure (Figure 1.11). Most fungi have at least three layers: an outermost amorphous layer that is removed by laminarinase treatment and therefore considered to be principally composed of a-(1^3) and a-(1^6)-linked anhydroglucose units. Polysaccharides occur as interwoven microfibrils embedded in an amorphous matrix. The polymers are apparently cross-linked, consistent with the idea that the hyphal wall can withstand an enormous turgor pressure. The composition and structure of the cell wall may vary with age and environmental conditions.

Fungal Cell Wall
Figure 1.11 Diagram of multilayered fungal cell wall. Adapted from Burnett (1976).
Thermophilic Cell Structure

Figure 1.12 Hydrophobin. (a) Electron micrograph of surface of aerial hyphae of Schizophylum commune after freeze-fracturing and shadowing showing layer of hydrophobin rodlets. (b) Electron micrograph of rodlets after drying solution of extracted hydrophobin. Source: Wessels (1966). With permission of Elsevier.

Figure 1.12 Hydrophobin. (a) Electron micrograph of surface of aerial hyphae of Schizophylum commune after freeze-fracturing and shadowing showing layer of hydrophobin rodlets. (b) Electron micrograph of rodlets after drying solution of extracted hydrophobin. Source: Wessels (1966). With permission of Elsevier.

1.2.2 Hydrophobins

The hyphae of terrestrial fungi grow in tight contact with substratum to perceive microscopic surface signals and orient the direction of their growth or lower the water surface tension and grow into the air for the formation and dissemination of spores (Talbot, 1997). The hydrophobic molecule that determines such growth of the hyphae is a protein called hydrophobin, which is extractable by strong trifluoroacetic acid or formic acid. Hydrophobins have a high proportion of non-polar amino acids that allows them to self-aggregate to form a film that appears as bundles of mosaic rodlets in electron micrographs (Figure 1.12). Hydrophobins were discovered by Joseph Wessels and his associates who estimated that hydrophobins, which are unique to fungi, constitute 10% of total proteins in fungi (Wessels, 1996).

1.3 MYCELIUM FORMATION 1.3.1 Septation

Except in the fungi belonging to Zygomycotina, the hypha is partitioned into cells by transverse walls or septa (singular septum). The septa are visualized by phase contrast microscopy or by fluorescence microscopy following staining with Calcofluor white, a cellulose and chitin binding dye (see Figure 2.1). Septa are spaced evenly—for example, in Aspergillus nidulans the apical cell (distance from the hyphal tip to the closest septum) is variable whereas the intercalary compartments have a uniform length of 38 jam (Wolkow et al., 1996), suggesting that some cellular mechanisms determine the site of placement of the septa. Septa are formed by centripetal growth of the cell wall and have a perforation (see Figure 1.1) through which cytoplasmic organelles, including nuclei, can pass. In fungi, the term "compartment" is often used in lieu of cells to denote that fungal cells have cytoplasmic continuity.

The role of septa is uncertain. The fungi belonging to Zygomycotina lack septa. In Ascomycotina and Basidiomycotina, an electron-dense protein-body called Woronin body—a peroxisome-derived dense-core—is present on either side of the septa and functions as a plug for the septal pore in response to cellular damage. By regulated closing or opening of the septal pores, the movement of protoplasm can be redirected to any region in the mycelium, changing the direction of hyphal extension for productive exploration of nutrient-rich surrounding areas. Septa may compartmentalize hypha to reduce leakage when the hypha is ruptured or provide a reducing environment for redox-sensitive enzymes. Septa, as with the rungs of a ladder, may contribute to the rigidity of hypha. For example, the cross wall less (cwl) mutants of Neurospora crassa are very weak, probably because the contents of a long hyphal cell ooze out as there are no cross walls to prevent "bleeding." In some higher fungi such as the Agarics, the septal pore is of elaborate construction with thickened sidewalls.

Another plausible role of septa is to allow spatial regulation of branch sites and development of reproductive structures by redistribution of nutrients. By the sealing of septal pores, the translocation of nutrients can be redirected toward a developing fruit body. For example, the spores of Coprinus sterquilinus that fall on vegetation are taken in by herbivores and voided with dung where they germinate. The hyphae subsequently fuse and form a single three-dimensional interconnected mycelial network that operationally acts as a unit for maximizing extraction of nutrients from the substrate and translocating these to a developing basidiocarp (fruiting body). "Cooperation by fusing mycelia, rather than competition by individual colonies, is the general feature of fungal growth in nature" (Burnett, 1976).

1.3.2 Branching

Branches arise in acropetal succession in proximity, generally sub-apically to the septum. Hyphae tend to avoid their neighbors and grow outwardly from the center. The pattern of branching can be compared to the pattern of branching in a fir tree with a main hypha and a series of branches borne alternately in two dimensions, suggesting a marked apical dominance. As the leading hyphae diverge from one another, apical dominance becomes weaker. Branching allows the hyphae to effectively colonize and exploit its surroundings.

The mechanisms involved in the cellular organization of hypha are beginning to be studied by genetic methods using conditional mutants (see Chapter 2). Mutants with defects in branching are easily recognized because, in contrast to the wild types, they form compact (colonial) colonies. The colonial temperature sensitive (cot ) mutant was identified as having a temperature-sensitive defect in hyphal growth: it produces colonial (tight, button-shaped) colonies at 32°C but normal wild-type colonies at 25°C. In a nuclear distribution mutant of Aspergillus nidulans, branching intensity—which was determined as the hyphal growth unit length (the ratio between the total hyphal length and the number of tips) —was higher in nutrient-rich media than on nutrient-limiting media (Dynesen and Nielsen, 2003). The cytoplasm volume per nucleus is unaffected by substrate availability, suggesting that branching and nuclear division are coordinated to maintain this ratio. An ecological implication of this is that in nature the fungus can explore its surrounding with thin, sparsely branched hyphae (minimum formation of biomass).

Aspergillus Cellwall Nature
Figure 1.13 Hyphal fusion. Tracings from Hickey (2002).

1.3.3 Hyphal Fusion

As different parts of a mycelium extend, neighboring hyphae may become physically interconnected by cell fusion, bringing all hyphae into a cytoplasmic continuity. The process of hyphal fusion was investigated in living hyphae by time-lapse imaging (Hickey et al., 2002). Short lateral branches arise and redirect their growth toward each other by a remote sensing mechanism to facilitate their contact. This process involves signaling and response of two hyphal tips but the nature of the signaling molecules is not yet known. Since the Spitzenkörper is observed where hyphae meet, this structure presumably delivers enzymes and cell adhesion molecules required for the dissolution and fusion, respectively, of cell wall at the point of contact of hyphal tips to allow for cytoplasmic continuity (Figure 1.13). Formation of interconnected hyphae allows the mycelium to forage in space and in time. Because of hyphal fusion and nuclear intermingling, a mycelium is a three-dimensional mosaic of hyphae of different genotypes (Brasier, 1984).

1.3.4 Multihyphal Structures

In hyphae the cell walls are laid down transversely. Even so, without cell divisions in vertical and anticlinal planes, some fungi form large macroscopic structures (tissue) such as basidiocarps, a fruiting body that produces the sexual spores of the mushroom or the bracket fungi1 (Figure 1.14). These structures are formed by the synchronized growth of thousands of hyphae towards each other with their branching, interweaving, thickening and gluing together by ß1—>6, ß 1 —>3 linked glucan. A feature of fungal morphogenesis is the synchronization of the activities of hyphae but how this occurs is not known.

1A basidiocarp of Rigidiocarpus ulmarius (over 5>2feet, 284 kg in 1996) in a corner of the Royal Botanic Gardens, Kew, Surrey, England is mentioned in the Guinness Book of World Records (http://tolweb.org/tree?group=Fungi&contgroup=Eukaryotes).

Bracket Fungus Oak Tree

Figure 1.14 Basidiocarps of a bracket fungus on the trunk of a mature oak tree. Development of these leathery structures of enormous strength requires the synchronized growth of hyphae toward one another, their localized branching, interweaving, binding and thickening. Courtesy of Heather Angel/Natural Visions. (See color insert following page 140.)

Figure 1.14 Basidiocarps of a bracket fungus on the trunk of a mature oak tree. Development of these leathery structures of enormous strength requires the synchronized growth of hyphae toward one another, their localized branching, interweaving, binding and thickening. Courtesy of Heather Angel/Natural Visions. (See color insert following page 140.)

0 0

Post a comment