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Regulation of appressorium development in pathogenic fungi (Appressorium…
Regulation of appressorium development in pathogenic
fungi
Appressorium maturation and cuticle rupture
The appressorium pore defines the point at the base of the infection cell from which the penetration hypha emerges.
appressorium pore is the site of
remodelling of the actin cytoskeleton
In M. oryzae and Colletotrichum species, appressorium pore is clearly distinct from the rest of the infection cell
much thinner cell wall and
the absence of melanin
During penetration peg formation, rapid F-actin polymerisation occurs leads to rapid polarised growth of
the penetration hypha
Re-modelling of actin requires
morphogenetic septin GTPases
A septin ring was observed at the appressorium pore which is necessary for scaffolding actin, leads to formation of toroidal F-actin network at the base of appressorium
Also acts as a lateral diffusion barrier, tethering in place proteins implicated in F-actin polymerisation
eg; Las17 component of the arp2/3 complex
Nox2 and its regulatory subunit NoxR required for septin ring formation at the base of the appressorium and a second NAPDH oxidase, encoded by the
NOX1
gene.
necessary for maintenance of the polarised growth and organisation of the toroidal F-actin network at the base of the appressorium during penetration peg formation
Mutation of gene either of the
NOX2
and
NOXR
genes is sufficient to prevent plant infection and the appressorium pore fails to differentiate from the rest of the infection cell
mutation of NOX1 leads to arrest of the penetration process just after differentiation of a stunted penetration peg which fails to
elongate and breach the cuticle
Reactive oxygen
species (ROS)
stimulate cytoskeletal remodelling
act directly on proteins
gelsolin, which are involved in actin severing and formation of free barbed ends that stimulate rapid F-actin polymerisation
acts on signalling components
operate down stream of a turgor sensor (or sensors) that must operate in the appressorium
INTRODUCTION
Plant pathogenic fungi cause many of world's most crop disease.
significant expenditure is require each year to combat the plant disease and ensure the fod security
Up to 30% of global harvest is lost each year
Identifying durable solution to plant disease is important by means to increase plant productivity in sustainable way.
specialized structure called appressorium
single cell structure which can collectively form structure known as infectious cushion.
simple terminal swelling at the tip of germ tube that emerge from the spore on the leave.
melanin pigmented, septate structure initially form at the tip og the germ tube, then differentiate into dome shape infection structure
magnaporthe oryza
e and
Colletotrichum
sp.
Targeting the fundamental morphogenetic process may important in developing the next-generation of anti-penetrant fungicides.
The penetration peg as site of effector delivery
Plant infection by pathogens involves
Deployment of
effector proteins that suppress plant immunity response
Facilitate proliferation of the pathogen within plant tissues
C. higginsianum
effectors within the appressorium pore at the point of plant infection
specialised focal secretion mechanisms for effectors are present in
Colletotrichum orbiculare
and
M. oryzae
Essential pre-requisite for focal secretion of effectors
Means of communication between the extending hyphal tip and the fungal nucleus
In
U. maydis
, a retrograde, early endosome-mediated, long-distance signalling pathway is necessary for transcriptional regulation of effector genes and effector secretion from the hyphal tip
APPRESORIUM TURGOR GENERATION
M.oryzae
appressorium maturation is acoompanied by rapi glycerol and other polyols synthesis.
lead to turgor generation and formation of thich differentiated melanin layer on inner side of the appressorium cell wall.
require to retard efflux of glycerol from rapidly expanding and provide structural rigidity and resilence to nfection cell.
Collectotrichum graminicola
anthracnose pathogen of corn.
turgor accumulate even when the melanin biosynthesis is inhibited and penetration of intact leaves and substrates still occur
melanin is not require for solute accumulation and turgor generation
by using mach-Zehnder interferometry
melanin may not provide the barrier for osmolytes.
Phakopsora pachyrhiz
i
soybean rust fungus
high turgor (5.13MPa) can be observe in non-melanised apprssoria.
using transmitted light double beam inteference Mach-Zehnder microscopy
hyaline (non pigmented appressoria) can generate turgor in absence of melanin in the cell wall.
Turgor generation still require the accumulation of osmotically active polyols but can be retained.
Early appressorium development
development
occurs after a spore lands o the surface of host
three cells conidium germinates within 1 hr after attachment ( adhesive= can adhere tightly to hydrophobic, waxy leaf cuticle)
upon hydration, the conidium germinates rapidly. The germ tube germinates out from the conidium and extends for 10-15 micrometre before flattening at the tips.
The germ tube hooking and start to differentiate into unicellular appressorium.
Initiation of appressorium development
based on hydrophobicity
surface need to have >90 degree water contact angles and surface hardness
fungus able to respond to wax monomers (eg. 1,16-hexadecanediol , powerful inducers of appressorium development at the leaf surface)
conidium
three-celled, each cell contains nucleus
germ tube emerges undergoes a single round of nuclear division before appressorium development
S-phase necessary for initiation of appressorium development
hydroxyurea or by generation of a temperature-sensitive nim1 mutant inhibit DNA replication
undergoes aberrant mitosis in the absence of DNA replication
completely prevents the ability of germ tubes to differen-
tiate at their tips
appressorium maturation and melanisation
controlled by entry of the
nucleus into G2 and mitosis
If mitosis occurs does
the appressorium become
fully functional
cytokinesis occurs
contractile actomyosin ring forms
at the neck of the appressorium
Differentiates the
cell from the rest of the pre-penetration structures
Autophagy
stimulated within the conidium during the development of appressorium
all of the intracellular contents of the three-celled spore are degraded before being trafficked to the incipient appressorium
culmination of this process is turgor generation within the appressorium of up to 6–8 MPa
sufficient to breach the underlying rice
cuticle
blocking autophagy by targeted mutation of any of the 16 genetic components of the non-selective macroautophagy pathway
Cell cycle control
During the appressorium development, the cell cycle is likely to be a conserved process
e.g
U. maydis
cycle arrest is necessary for an infective filament of the
fungus to be able to penetrate plant tissue
undergoes a self-/non-self-recognition process on
the corn leaf surface in which two monokaryotic sporidia fuse to form an infectious dikaryotic filament
forms an appressorium, which is necessary to breach the
corn leaf surface
Recent evidence
cell cycle arrest is required for plant infection
because of cooperation of at least two
distinct underlying mechanisms
activation of the DNA damage response cascade
the other relies on transcriptional regulation of a gene called HSL1
encodes a protein kinase that modulates
the G2 to M transition
Thus, the control of nuclear division and its coordination with morphogenesis at the leaf surface appear to be processes which are fundamental to penetration of the cuticle by diverse plant pathogens
physical and biochemical cues at the
leaf surface
Mac1 adenylate
cyclase interacts with Cap1
Cap1 = cyclase-associated protein
that activates adenylate cyclase
potentially involved
in re-modelling the actin cytoskeleton
appears to strongly associate based on its locali-
zation pattern in appressoria
Pmk1 MAPK pathway
necessary for appressorium
development
Upstream of Pmk1 a number of potential receptors are involved in perception of surface signals
e.g
PTH11
CFEM domain Gprotein
coupled receptor,
necessary for perception of
the hydrophobic leaf surface
its
absence, appressoria do not form
RAS signalling
likely to act upstream of the Pmk1 and cAMP response
pathways
because generation of a dominant-active allele
of Ras2 (RAS2G17V)
leads to abnormal appressorium
formation in the absence of a surface
appressorium-
like structures can be formed by aerial hyphae
Expression of the dominant M. oryzae RAS2G17V
allele in Colletotrichum graminicola and C. gloeosporioides
led to aerial appressoria
suggesting conservation of
the surface perception signalling mechanism
Pmk1 kinase cascade
composed of three protein kinases
Mst7
Pmk1
Mst11
scaffolded by a protein called Mst50 which interacts
with Mst11
upon activation and phosphorylation of
its components
a phosphor-relay is triggered leading to
the detachment of Pmk1
its traversal to the nucleus
during appressorium maturation
Recent tran-
scriptional profiling results and interaction studies suggesting
several transcription factors operate
downstream of Pmk1, including Mst12 and Mcm1,
which likely activate a large set of gene products involved
in cell wall differentiation
the physiological
changes associated with appressorium maturation
turgor generation, in addition to the control of autophagy
programmed cell death of the conidium that precedes
appressorium maturation
Pmk1 pathway also appears to regulate microconidia formation
by M. oryzae
because Pmk1 and Mst12 mutants show
reduced microconidia production
while Mcm1 is essen-
tial for their development
Microconidia may represent
an alternative means of propagation by the pathogen
to
facilitate rapid spread within plant tissue
Pmk1 pathway is widely conserved in other plant
pathogenic fungi
likely to be required for infectionrelated
morphogenesis