Growth, Differentiation and Sexuality

60 J.H. Sietsma and J.G.H. Wessels
toxin (Hutchings and Bussey 1983). Bussey and coworkers
have used killer-toxin-resistant (KRE)mutants
to identify several genes required for the synthesis
of (1-6)-β-glucan. The analyses of numerous
mutants suggest that the polymer is synthesised in
a sequential manner involving several gene products,
some of which are in the secretory pathway,
others being cytoplasmic or membrane proteins
(Brown et al. 1993). It appears that both in yeast
and mycelial fungi, glyco-proteins, synthesised on
a GPI-anchor, are involved in linking a (1-6)-βglucan
protein complex to the cell wall matrix. The
protein part often plays an active role in fungal
wall synthesis and modification outside the plasma
membrane (Mouyna et al. 2000; Klis et al. 2002).
V. Wall Modifications During Wall
Expansion
A. Steady-State Model for Apical Wall Growth
The simplest interpretation of the events occurring
at the hyphal tip, based on what is known
of the structure of the mature wall and the synthesis
of individual wall components described in
the preceding section, is that the major polysaccharides,
(1-3)-β-glucan and chitin, are extruded
into the apical wall as individual chains. Together
with water, they probably constitute a plastic, hydrated,
gel-like wall, easily deformable by pressure
exerted by the cytoplasm. However, as soon as the
two primary components are extruded into the wall
domain, they become subject to enzymic modifications,
become covalently linked to each other,
and intra-molecular hydrogen bonds are formed.
It is plausible that the total of these changes results
in a gradual change in the mechanical properties
of the wall, going from plastic to more rigid. Because
cross-linking of the primary wall polymers
is a time-dependent process and the newly formed
wall continuously falls behind the expanding apical
wall, this would imply the presence of a steady-state
amount of plastic wall material at the growing hyphal
tip, and the presence of hardened wall at the
base of the extension zone. This mechanism has
therefore been called the “steady-state” model of
apical growth (Wessels 1986). In accordance with
this model, it was found that the structure of the
growing apical wall is indeed unique. Early studies
(reviewed by Wessels 1986) and a recent study (Momany
et al. 2004) have shown that the hyphal apex
differentially binds the dyes calcofluor white and
congo red, the lectin wheat agglutinin and some antibodies
raised against cell walls. More specifically
it was found that, in contrast to the subapical wall,
the wall at the growing apex contains non-fibrillar
chitin and no alkali-insoluble glucan, and that cessation
of growth transforms the apical wall into
a structure indistinguishable from that of the mature
subapical wall (Wessels et al. 1983; Vermeulen
and Wessels 1984; Sietsma et al. 1985).
Direct experimental evidence, obtained with
spore germlings of S. commune (Sietsma et al. 1985)
supporting the steady-state growth theory of apical
wall growth, is summarised in Fig. 4.1.
– Pulse-labelling with [ 3H]acetylglucosamine resulted
in the labelling of chitin, nearly all alkaliinsoluble,
maximally at the extreme apex and
rapidly decreasing subapically. Labelling with
[ 3H]glucose resulted in the labelling of total
glucan in the wall in a similar pattern; after
the short labelling period, no label was detectable
in chitin. However, at the apex very
little label appeared in alkali-insoluble glucan;
Fig. 4.1. Experimental evidence supporting the steady-state
growth theory of apical wall expansion. Patterns of labelling
and tabulated results are from Wessels et al. (1983), Vermeulen
and Wessels (1984), and Sietsma et al. (1985). NA
Not applicable. See text for more explanation (from Wessels
et al. 1990, with permission)