Growth, Differentiation and Sexuality

In agreement with an essential role of the
chitin–glucan complex in hyphal morphogenesis,
mutants of A. nidulans and Neurospora crassa
affected either in chitin or β-glucan synthesis show
osmotic sensitivity and abnormal morphology
(Leal-Morales and Ruiz-Herrera 1985; Martinez
et al. 1989; Borgia and Dodge 1992). Because chitin
and (1-3)-β-glucan are synthesised by different
enzymes at the plasma membrane (see below),
linkage of these two polymers to each other can
occur only outside the plasma membrane within
the wall domain.
A. Chitin Synthesis
1. Regulation of Chitin Synthase Activity
It is now generally accepted that chitin is synthesised
by a trans-membrane protein, accepting its
substrate, uridine-diphospho-N-acetyl-glucosamine
(UDP-GlcNac) at the cytoplasmic site while
the (1-4)-β-linked N-acetyl-glucosamine polymer
is extruded to the outside (Duran et al. 1975;
Vermeulen et al. 1979; Cabib et al. 1983). This
topic has been reviewed by Roncero (2002, in The
Mycota, Vol. III, 2nd edn., Chap. 14), and by Latgé
and Calderone (Chap. 5, this volume).
In fungi, chitin synthases are encoded by multigene
families containing from three members in
Saccharomyces cerevisiae to eight (e.g. for Benjaminiella
poitrasii) or even ten members (e.g. Phycomyces
blakesleeanus; Bulawa 1993; Miyazaki and
Ootaki 1997; Chitnis et al. 2002). Based on sequence
homology, chitin synthase genes are divided into
five classes. Studies on disruptions of these genes
provide more data for the function of these genes,
indicating that different functions can be assigned
to members of different classes. When disrupted,
class I genes show hardly any phenotypical effect,
a repair function during cytokinesis having been
assigned only to the yeast member of this class
(Cabib et al. 1992). Disruption of class II genes had
an effect on septum synthesis and conidiogenesis
(Fujiwara et al. 2000; Munro et al. 2001). Class IV
contains chitin synthase genes coding for enzymes
responsible for the synthesis of the bulk of chitin
present in yeast or hyphal cell wall. Although disruption
of these genes causes a considerable reduction
in the amount of chitin in the wall, it does
not produce aberrant hyphal morphologies (Din
et al. 1996; Specht et al. 1996). Genes belonging to
classes III and V are present only in mycelial fungi
and absent in yeasts (Weber et al. 2003). Disruption
Apical Wall Biogenesis 57
of these genes shows in several cases abnormal hyphal
growth (Yarden and Yanofsky 1991; Mellado
et al. 1996). Significantly, genes belonging to class V
code for fusion proteins between myosin and chitin
synthase, the myosin part possibly playing a major
role in directing chitin synthase to the site of action,
the hyphal tip (Aufauvre-Brown et al. 1997; Roncero
2002). In S. cerevisiae,evidenceexiststhatamyosin
motor molecule (Myo2) transports chitin synthase
to its site of action (Santos and Snyder 1997). However,
Myo2 is not involved in chitin synthase transport
in mycelial fungi (Weber et al. 2003).
The aforementioned autoradiographic studies
indeed show chitin synthase to be particularly active
at growing hyphal apices and at developing
septa. This suggests precise localisation of chitin
synthase and/or its precise local activation.
Chitin synthase, like other membrane proteins,
may arise at the ER far behind the hyphal tip and
then be transported by vesicles to the apex where
it is inserted into the plasma membrane by vesicle
fusion. Docking SNARE and vesicle SNARE
molecules are found to be present at hyphal tips
and are thought to play a role in the precise localisation
of vesicle fusion to the cytoplasmic membrane
(Gupta et al. 2003). Vesicle-like particles called chitosomes,
containing inactive chitin synthase, have
been isolated from a variety of fungi (Bartnicki-
Garcia et al. 1978). They show a variety of proteins
and lipids, which seem to be essential for the integrity
and functioning of the chitosomes (Flores-
Martinez et al. 1990); upon activation with proteolytic
enzymes, they produce crystalline chitin in
vitro. However, these chitosomes are much smaller
than the usual secretory vesicles present at the hyphal
apex, and do not seem to be delineated by
a unit membrane (Bracker et al. 1976). Therefore,
it is questionable whether these structures can be
called true vesicles. Chitosomes may be unique assemblages
of lipids and proteins, the latter possibly
synthesised on free ribosomes. Significantly, the
chitin synthase genes cloned thus far do not indicate
the presence of canonical signal sequences for
secretion (Silverman 1989).
Proteolytic activation in vitro of chitin
synthases has been generally observed (Bartnicki-
Garcia et al. 1978; Cabib et al. 1982). Cabib et al.
(1982) have implied local proteolytic activation
of chitin synthase at the site of septum formation
in S. cerevisiae, assuming a zymogenic form of
the enzyme uniformly present in the plasma
membrane. However, there is no direct evidence
that proteolytic activation does occur in vivo. In