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Regulation of appressorium development in pathogenic fungi (Early…
Regulation of appressorium development in pathogenic fungi
Appressorium maturation and cuticle rupture
appressorium pore distinct from the infection cell
thinner cell wall and the absence of melanin.
During penetration peg formation:
rapid F-actin polymerisation occur
rapid growth of the penetration hypha
Re-modelling of actin requires morphogenetic septin GTPases
Septin ring:
-observed at the appressorium
pore
-composed of four core septins,
Sep3, Sep4, Sep5 and Sep6
-necessary for scaffolding actin
-leading to formation of a toroidal Factin network at the base of the appressorium
-acts as a lateral diffusion barrier
-tethering in place proteins implicated in F-actin polymerisation (Las17 component of the arp2/3 complex)
Ezrin, radixin, moesin (ERM) domain proteins:
-required for actin membrane interactions at the cortex of
cells
-located within the septin ring at the appressorium pore
BAR domain proteins implicated in the control of membrane curvature generation
-Eukaryotic cells undergo membrane curvature generation
-to generate invaginations associated
-with endocytosis and also cellular protrusions, such as lamellipodia found in epithelial cells
Cellular protrusions:
-require membrane curvature to be stimulated
-followed by rapid membrane biogenesis and F-actin polymerisation
-must be spatially regulated to the point of plant infection
reactive oxygen species burst catalysed by the
Nox2
-NADPH oxidase is necessary for septin-mediated appressorium repolarisation
-.
Nox2
and its regulatory subunit
NoxR
are required for septin ring formation
-second NAPDH oxidase, encoded by
NOX1
gene, is necessary for maintenance of thepolarised growth and organisation of the toroidal F-actin during penetration peg formation
-
NOX2
and
NOXR
genes is sufficient to prevent plant infection
-the appressorium pore fails to differentiate from
the rest of the infection cell.
-mutation of
NOX1
leads to arrest of the penetration process
Reactive oxygen species (ROS):
-generated within the appressorium
-act directly on proteins(gelsolin)
-involved in actin severing
-formation of free barbed ends that stimulate rapid F-actin
polymerisation
-action of latrunculin, an actin depolymerising agent, could be competitively inhibited by the presence of ROS leading to
penetration peg formation
ROS:
-acts on signalling components that operate downstream of a turgor sensor (or sensors) that must operate in
the appressorium
-re-polarisation needs to be triggered
formation of the hetero-oligomeric septin ring
involved in this process likely include
Chm1, a protein kinase implicated in septin phosphorylation
The penetration peg as site of effector delivery
Plant infection:
-involves deployment of
effector proteins that suppress plant immunity responses
-facilitate proliferation of the pathogen within plant
tissues
Ultra-structural analysis of
C. higginsianum
:
-has detected effectors within the appressorium pore
-specialised focal secretion mechanisms for effectors are likely to be present in both
Colletotrichum orbiculare
and
M. oryzae
U. maydis
During plant tissue
colonisation:
-retrograde, early endosome-mediated
-long-distance signalling pathway is necessary for transcriptional regulation of effector genes
-effector secretion from the hyphal tip
Early Approssoria Developement
Occurance
: Spore lands on surface of host
Adhere tightly to the hydrophobic, waxy leaf cuticle - respond to wax monomers (1,6-hexadecanediol)
germ tube extends- flatterning tps- hooked- differentiate unicell. appressorium
single round of nuclear divisione
DNA replication (S-phase) - initiation appressorium development.
G2 & mitosis - appressorium maturation and melanisation
Cytokinesis - contractile actomyosin ring forms @ neck of appressorium- differentiate cell from the rest of the pre-penetration structures.
Autophagy - the intracellular content of 3-celled conidium are degraded. Blockage autophagy - sufficient to render the fungus non-pathogenicity
Cell cycle arrest - cooperation at least two dinstinct underlying mechanism (HSL1 - protein kinase- modulates G2 & M transition)
Formation- Physical & biochemical cues @ leaf surface
Pmk 1 pathway -infection related morphogenesis
Appressorium turgor generation
maturation of appressorium by rapid synthesis of glycerol and other polynols
formation of thick differentiated melanin layer on the inner side of the appressorium cell wall
- required to retard efflux of glycerol from rapid expanding appressorium and to provide structural rigidity and resilience to the infection cell
melanin in the appressorium
- to maintain turgor pressure due to lowering the porosity of the appressorium cell wall
cell collapse assay (cytorrhysis) analysis of the appressorium osmolyte content using method called
Mach-Zehnder interferometry
- shows melanin is not required for solute accumulation and turgor generation
melanin
plays structural role -because albino mutants, lacking the CgPKSI polyketide synthase gene involved in 1,3,6,8-tetrahydroxy-naphthalene biosynthesis-which prone to rupture and impaired in their ability to cause disease
high turgor could be observed in non-melanised appressorium
turgor generation still requires accumulation of asmotically active polyols but can apparently be retained in the absence of melanin
other non-melanised fungi still undertake mechanical appressorium-mediated infection
Introduction
Appressorium
Can be single-celled structures or compound (consist of numerous cells collectively form structures known as infection cushions).
Simple terminal swellings at the tip of germ tubes that emerged from spores on the leaf surface.
Melanin-pigmented, septate structures then fully differentiated into dome-shaped in model organisms (
M. oryzae
and
Colletotrichum
sp.)
Appressorium formation
is a highly orchestrated development process required
(1)
perception of physical and chemical cues from plant leaf surface and
(2)
control of both nuclear and cell division
Future prospects
How isotropic expansion of appressoria is translated into the generation of invasive forces to break plant cuticle.
Appressorium pore as key signaling hub during plant infection for site of rapid fungal growth and initial secretion of fungal effectors and regulatory components.
Future works include
(1) How focal secretion is regulated and the conservation of early endosome-mediated signaling.
(2) Precise mechanism of how appressorium monitors turgor because it determined the optimal point for repolarization and host cell penetration
(3) To identify turgor sensing mechanism and understand how it trigger regulated synthesis of ROS leading to septin-mediated cytoskeletal re-organization.
High throughput targeted genome editing like CRISPR-Cas9 and silencing