RESUMO
Transcription factors play a critical regulatory role in development by binding DNA and initiating alterations in gene transcription. The transcript of the putative Magnaporthe oryzae transcription factor-encoding gene TRA1 accumulates during germination and this accumulation was previously found to depend on the transcription factor Con7p. In the current work tra1â» mutants were generated and these strains were found to exhibit a reduced attachment, germination, appressorium formation and virulence. Adhesion to artificial and plant surfaces was affected, and FITC-labelled concanavalin A, a lectin which inhibits attachment of Magnaporthe spores, showed a reduced affinity for mutant spore tip where it normally preferentially binds. We used microarray analysis to identify Tra1p-dependent genes from two different sources: aerial structures and conidia. Mutation of 11 Tra1p-dependent genes showed that the predicted transcription factor encoding gene TDG2 is required for normal adhesion and virulence, that the genes TDG7 and TDG4 are required for normal sporulation and that TDG6 is required for wild-type levels of spore adhesion.
Assuntos
Proteínas Fúngicas/genética , Magnaporthe/genética , Esporos/genética , Fatores de Transcrição/genética , Regulação Fúngica da Expressão Gênica , Proteínas de Fluorescência Verde , Magnaporthe/crescimento & desenvolvimento , Magnaporthe/patogenicidade , Mutação , Oryza/genética , Oryza/crescimento & desenvolvimento , Oryza/microbiologia , Esporos/crescimento & desenvolvimento , Esporos/patogenicidade , Esporos Fúngicos/genética , Esporos Fúngicos/crescimento & desenvolvimento , VirulênciaRESUMO
The plant pathogenic fungus Magnaporthe grisea is able to enter its host via appressorium-mediated penetration. Earlier investigations have shown that these infection structures are rich in the cell wall polysaccharide chitin. Previously, we have described how the transcription of a class VII chitin synthase-encoding gene CHS7 is completely dependent on the putative transcription factor Con7p during the germination of conidia, and how con7(-) mutants are unable to form appressoria under any conditions tested. Because of the pleiotropic effects of the con7(-) mutation, we examined the consequences of the targeted deletion of CHS7. The chs7(-) mutants generated were unable to form appressoria on artificial surfaces, except following the application of the exogenous inducers 1,16-hexadecanediol and cyclic adenosine monophosphate. The appressoria formed had a reduced chitin content and were often found to be smaller and misshapen compared with the wild-type. chs7(-) mutants were significantly reduced in their ability to enter rice plants, but growth in planta was not affected. Reverse transcriptase-polymerase chain reaction analysis demonstrated that CHS7 transcription was strongly induced on germination of spores, and a green fluorescent protein-tagged Chs7p protein was found to be produced abundantly during infection-related morphogenesis. Together, these data suggest that the class VII chitin synthase Chs7p of M. grisea is required for normal appressorium formation and function.
Assuntos
Quitina Sintase/metabolismo , Magnaporthe/enzimologia , Sequência de Bases , Quitina Sintase/genética , Primers do DNA , DNA Complementar , Deleção de Genes , Genes Fúngicos , Proteínas de Fluorescência Verde/genética , Magnaporthe/genética , Microscopia de Fluorescência , Dados de Sequência Molecular , Mutação , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
A strain harbouring an insertion within the promoter of the CON7 gene of Magnaporthe grisea was isolated. This gene was previously shown to be essential for appressorium formation and growth in planta and is predicted to encode a transcription factor. Microarray-based gene expression analysis was used to identify several genes whose transcription during germination depends on Con7p. These include the pathogenicity factor-encoding gene PTH11 and several other genes which like PTH11 are predicted to encode G protein-coupled receptors. Microarray analysis also revealed several Con7p-dependent genes which may encode factors determining cell wall structure or function, either through the synthesis/degradation of cell wall components or by association with the cell exterior. One Con7p-dependent gene predicted to encode a class VII chitin synthase was deleted, leading to dramatic consequences on the pathogenic development of the resultant strain. Within the con7(-) mutant, a 29% reduction in chitin content of germinated spores was found and the mutant was hypersensitive to the chitin synthase inhibitor nikkomycin Z. A green fluorescent protein-tagged Con7p was found to have nuclear localization within spores. Taken together, these observations suggest that Con7p encodes a transcription factor required for the transcription of several genes which participate in disease-related morphogenesis in M. grisea.