are self-assembling aggregates of the enzyme chitin
synthase, each particle containing sufficient enzyme
molecules to produce, after proteolytic activation, a
chitin microfibril composed of several chitin chains. So,
this elegant collaborative study by biochemists and elec-
tron microscopists led to the identification of one of
the types of vesicle often seen in the hyphal apex. One
of the questions still to be resolved is how these par-
ticles reach the hyphal tip after they have been assem-
bled further back in the hyphae. One suggested
mechanism is that they are packaged into membrane-
bound multivesicular bodiessuch as those some-
times seen in electron micrographs (Fig. 3.20).
Vacuoles
The vacuolar system of fungi has several functions,
including the storage and recycling of cellular meta-
bolites. For example the vacuoles of several fungi,
including mycorrhizal species (Chapter 13), accumulate
phosphates in the form of polyphosphate. Vacuoles also
seem to be major sites for storage of calcium which can
be released into the cytoplasm as part of the intra-
cellular signalling system (Chapter 5). Vacuoles contain
proteases for breaking down cellular proteins and recycl-
ing of the amino acids, and vacuoles also have a role
in the regulation of cellular pH. All these important
physiological roles are in addition to the potential role
of vacuoles in cell expansion and (possibly) in driving
the protoplasm forwards as hyphae elongate at the tips.
Vacuoles often are seen as conspicuous, rounded
structures in the older regions of hyphae, but recent
work has shown that there is also a tubular vacuolar
system extending into the tip cells. It is an extremely
dynamic system, consisting of narrow tubules which
can dilate and contract, as inflated elements travel
along them in a peristaltic manner (Fig. 3.14). This was
demonstrated by studying the movement of a fluores-
cent compound, carboxyfluorescein (CF), in living
hyphae of a range of fungi. The hyphae were treated
with a nonfluorescent precursor, carboxyfluorescein
diacetate (CFDA), which is lipid-soluble so it enters
the plasma membrane and is then rapidly absorbed
from the cytosol into the vacuoles. There the acetate
groups are cleaved by esterases to yield CF, which is
membrane-impermeable so it remains in the vacuolar
system. Rees et al. (1994) showed that the tubular
vacuoles which contain CF form a more or less
continuous system which passes through even the
dolipore septa of Basidiomycota, transporting mater-
ials backwards and forwards in the hyphae. This has
important implications for the bidirectional trans-
location of materials within single hyphae (Chapter 7),
and perhaps especially for mycorrhizal systems (Chap-
ter 13).Endocytosis and vesicle traffickingStudies on plant cells, animal cells, and budding yeast
suggest that the uptake of substances through the60 CHAPTER 3
Fig. 3.13Ribbon-like aggregates of
chitin microfibrils (R) produced in vitro
from chitosomes (C) isolated from
Mucor rouxii, when incubated with
substrate (N-acetylglucosamine) and a
proteolytic activator. (Courtesy of C.E.
Bracker; from Bartnicki-Garcia et al.
1978.)