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Host–microbe interactions: fungi
activity of some of these genes that are differentially expressedduring plant colonization could be followed through thevarious stages of the infection cycle [7,10,11,12
•
,13
••
,14
•
].In the following paragraphs, we have divided the genesidentified into groups corresponding to events occurring onthe plant surface and during growth within the plant (Table 1).
The early phase: attachment, germination,appressorium development
Plant surface parameters determine initial developmentand can provide signaling cues, as exemplified by genesuniquely expressed at this stage in
Colletotrichum gloeosporioides
(Table 1) [5]. The function of these genes is unknown;however, they are selectively induced by waxes on thehost surface [5], and CAP20 is essential for appressoriumfunction [15]. For
Colletotrichum
species, it has also beendemonstrated that the plant hormone ethylene inducesboth germination and appressorium formation and, in doingso, probably links infection to the time of fruit ripening [5].The response to host wax and ethylene requires contact of conidia with a hard surface [16] and evidence exists, thathard surface contact induces Ca
2+
calmodulin signaling inthis fungus[17]. In extending these studies, a number of
C.gloeosorioides
genes (
CHIP
) that are expressed upon hardsurface contact were identified [18,19].
CHIP1
encodes afunctional ubiquitin-conjugating enzyme.
CHIP2
may be atranscription factor, and CHIP3 protein has been predictedto be membrane-localized.Hydrophobins are small, secreted fungal proteins thatassemble at hydrophobic/hydrophilic interfaces and thusmay be involved in adherence of fungi to the hydrophobicsurface of plants [20].Such a role may be fulfilled byMPG1, a hydrophobin of
M. grisea
that is expressed duringappressorium formation ([21] and references therein). Inthe same fungus,
PTH11
was identified as a genuine path-ogenicity gene that is required for efficient appressoriumformation in response to cutin monomers as well as contactsurface hydrophobicity [13
••
].
PTH11
encodes a noveltransmembrane protein, Pth11p, which is only weaklydetectable in the cell cortex and is redistributed to vacuolarmembranes upon appressorium induction, which mayindicate downregulation. Both appressorium formationand pathogenicity could be restored by cAMP treatment of two independent REM1 mutants lacking PTH11, suggestingthat Pth11p is an upstream activator of cAMP-mediatedsignaling or functions in an overlapping signaling pathway.On poorly inductive surfaces, Pth11p appears to repressappressorium differentiation and may thus direct thefungus to the appropriate site of entry [13
••
].Melanization of appressoria is necessary for their functionand key genes of melanin biosynthesis have been identifiedin
M. grisea
and
Colletotrichum lagenarium
([22] and referencestherein; [23,24]). In
C. lagenarium
, the expression of threeof these genes is developmentally regulated [22].Successful penetration of the plant may not only dependon infection structure formation but may also be supportedby enzymes capable of degrading components of the plantcell wall. Cutinase genes that are induced during the earlyinteraction stage have been identified in several pathosys-tems (Table 1) [5,25,26]. In
Colletotrichum lindemuthianum
,an endopolygalacturonase encoding gene,
clpg2,
was tran-siently induced by pectin and showed expression ingerminating conidia, in appressoria, as well as in penetratinghyphae [12
•
,27]. In
Fusarium solani
f. sp.
pisi
, the pectate
Figure 1
Strategies to identify regulatory componentsof plant-induced genes.
(a)
Identification ofa negative regulator (R). The promoter (P) ofa negatively regulated gene (
mig1
) is fusedto a selectable marker gene (
SM
) and isectopically introduced into the fungalgenome. After UV-mutagenesis, strains areselected that express
SM
. Candidates arethen analyzed for increased expression ofthe endogenous
mig1
gene to exclude cismutations.
(b)
Identification of positivelyacting cues. A strain is constructed thatcontains two reporter genes (for SM andGFP) each, fused to the regulated promoterof
mig1
. Three alternative methods foridentifying positive regulators of thepromoter are shown. (1) An expressionlibrary is introduced and strainsconstitutively expressing a positive regulator(Ac) are selected via SM. (2) Plantcompounds (PS) are screened for elevatedGFP expression levels. (3) AfterUV-mutagenesis, strains are isolated thatexpress
SM
. In these mutants, componentsof the signaling cascade are constitutivelyactivated. The second reporter gene(forGFP) allows monitoring of the strengthof induction conferred.
AcAc
mig1mig1P SM mig1P
GFP
mig1P SM mig1P
PS
Current Opinion in Microbiology
UVExpressionlibraryUVR123
mig1mig1P SM mig1P
R
(a)(b)
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