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DIFFERENTIATION AND DEVELOPMENT 91

displaced towards the surface on which the fungus
is growing, facilitating contact-sensing. Consistent
with this, transmission electron micrographs show
that parallel arrays of microtubules are abundant just
beneath the plasma membrane of the lower surface
of the germ-tube, where they could be involved in
transducing the contact signals. All of this depends
on close adhesion to the surface, mediated by the
extracellular matrix, and in the rust Uromyces appen-
diculatusthe digestion of this matrix by applying a pro-
tease, Pronase E, prevents the topographical signaling.
By preparing protoplasts of this fungus and using
the patch-clamp technique it has been shown that
the plasma membrane at the germ-tube tip contains
stretch-activated ion channels. So it is believed that
stretching of the membrane when the germ-tube tip

encounters a ridge or groove leads to an ion flux (pos-

sibly Ca^2 +) through these channels, that coordinates the

orientation response.

The alignment of a germ-tube is the first stage in an

intricate developmental sequence which ensures that

infection from uredospores of rust fungi always occurs

through the stomata of a leaf surface. This sequence is

shown in Fig. 5.7, for the bean rust fungus Uromyces

appendiculatus.

When a germ-tube locates a stomatal ridge, it stops

growing after about 4 minutes, and the apex swells

to form an appressorium. The original two nuclei in

the germ-tube migrate into the appressorium, then

divide, and a septum develops to isolate the appres-

sorium from the germ-tube. About 120 minutes after

contacting the stoma, an infection peg grows into

Fig. 5.5Scanning electron micrograph
showing directional growth of hyphae
arising from uredospores of Puccinia
graminison an inert replica of the
lower surface of a wheat leaf. Note the
perpendicular alignment of the hyphae
to the contours of the leaf replica, and
the short lateral branches that arise in
the grooves. The hyphae arising from
two spores have located the “stomata”
on the leaf replica and produced
appressoria directly over the stomatal
pores. (Courtesy of N.D. Read; from
Read et al. 1992.)

Fig. 5.6Scanning electron micro-
graph of two hyphae of Puccinia
graminisgrowing perpendicularly over
the ridges and grooves of a polystyrene
replica of a microfabricated silicon
wafer. The germ-tube tips have a
“nose-down” orientation which might
facilitate topographical sensing. Arrow-
heads indicate the dried remains of
mucilage that adhered the germ-tubes
to the surface. Although it is not clearly
shown in this image, the hyphae
also form projections that grow into
the grooves (arrows), equivalent to the
short lateral branches shown in Fig. 4.6.
(Courtesy of N.D. Read; from Read
et al. 1992.)