Unknown,Transcriptomics,Genomics,Proteomics

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Transcription factor AbVf19 induces hydrolytic enzyme genes during pathogenesis in Alternaria brassicicola

ABSTRACT: Alternaria brassicicola is a successful saprophyte and necrotrophic pathogen with a broad host range. It produces secondary metabolites that marginally affect virulence, in contrast to many A. alternata strains that produce secondary metabolites as host-specific pathogenicity factors. Cell wall-degrading enzymes (CWDEs) have been considered important for pathogenesis, but no CWDEs have been identified as significant virulence factors in A. brassicicola. In this study, we discovered mutants of a gene, AbVf19, which consistently produced smaller lesions than the wild type. The mutants grew slower than the wild type on an axenic medium with pectin as a major carbon source. Gene expression comparisons identified several hydrolytic enzyme-coding genes being down-regulated in the mutant during a late stage of infection. These down-regulated genes comprised a small fraction of genes within each family. Three of these genes had mutants that showed no or little change in virulence. This suggested that each down regulated gene only made a small contribution to virulence, or that their functions were redundant. This study demonstrated the existence and importance of a transcription factor that regulates a suite of genes that are probably important for decomposing and utilizing plant material during the late stage of plant infection. gene expression profile comparisons between wild type and a transcription factor mutant during host infection

ORGANISM(S): Brassica oleracea

SUBMITTER: Yangrae Cho

PROVIDER: E-GEOD-33148 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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A zinc-finger-family transcription factor, AbVf19, is required for the induction of a gene subset important for virulence in Alternaria brassicicola.

Srivastava Akhil A Ohm Robin A RA Oxiles Lindsay L Brooks Fred F Lawrence Christopher B CB Grigoriev Igor V IV Cho Yangrae Y

Molecular plant-microbe interactions : MPMI 20120401 4

Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen with a broad host range within the family Brassicaceae. It produces secondary metabolites that marginally affect virulence. Cell wall-degrading enzymes (CDWE) have been considered important for pathogenesis but none of them individually have been identified as significant virulence factors in A. brassicicola. In this study, knockout mutants of a gene, AbVf19, were created and produced considerably smaller lesions ...[more]

PMID: 22185468

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Project description:Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Several A. brassicicola genes have been characterized as virulence factors affecting pathogenesis of Brassica species. To study regulatory mechanisms of pathogenesis, we mined 421 genes in silico encoding putative transcription factors in a machine-annotated, draft genome sequence of A. brassicicola. Targeted gene disruption mutants for 114 of the genes with proteins predicted to contain at least one putative zinc-finger domain were produced and functionally analyzed. Six of these genes were associated with pathogenesis. Disruption mutants corresponding to five of the genes were ≥50% less virulent than the wild type. Unexpectedly, the mutants of one gene (designated Amr1) were 100% more virulent. Amr1 is a homolog of Cmr1, a transcription factor previously found to regulate melanin biosynthesis in several fungi. Gene deletion mutants (Δamr1) were created and their phenotypes characterized. The Δamr1 mutants utilized pectin as a carbon source more efficiently than the wild type, were melanin-deficient, and more sensitive to UV light and glucanase digestion. The Amr1 protein was localized in the nuclei of hyphae and in highly melanized conidia during the late stage of plant pathogenesis. RNA-seq analysis revealed that three genes in the melanin biosynthesis pathway, along with the deleted Amr1 gene, were not expressed in the mutants. In contrast, many hydrolytic enzyme-coding genes were expressed at much higher levels in the mutants than in the wild type during pathogenesis. The results of this study suggested that only a small number of transcription factors with zinc finger domains are needed to maintain strong virulence. Furthermore, a gene important for survival in nature negatively affected virulence, probably by less efficient use of pectin. We speculate that the functions of the Amr1 gene are important to the success of A. brassicicola as a competitive saprophyte and plant parasite gene expression profile comparisons between wild type and a transcription factor mutant during the late stage of host infection

Project description:Alternaria brassicicola is a successful saprophyte and necrotrophic plant pathogen. Molecular determinants of its life style shift between saprophyte and pathogen, however, are unknown. To identify these determinants we studied nonpathogenic mutants of a transcription factor-coding gene, AbPf2. Frequency and timing of germination and appressorium formation on host plants were similar between the nonpathogenic Δabpf2 mutants and wild-type A. brassicicola. The mutants were also similar in vitro to wild-type A. brassicicola in vegetative growth, conidium production, and responses to chemical stressors, such as a phytoalexin, reactive oxygen species, and osmolites. The mutants, however, did not penetrate host plant tissues, though their hyphae continued to grow on the plant surface. Transcripts of the AbPf2 gene increased exponentially soon after wild-type conidia encountered their host plants. A small amount of AbPf2 protein, monitored by fused green fluorescent protein, was located in both the cytoplasm and nuclei of young, mature conidia. The protein level decreased during saprophytic growth but increased several-fold during pathogenesis. Levels of both the proteins and transcripts sharply declined following colonization of host tissues beyond the initial infection site. When the transcription factor was expressed at an induced level in the wild type during early pathogenesis, the expression of 106 fungal genes was down-regulated in the Δabpf2 mutants. Notably, 33 of the 106 genes encoded secreted proteins, including eight putative effector proteins. Plants inoculated with Δabpf2 mutants expressed higher levels of genes associated with photosynthesis, the pentose phosphate pathway, and primary metabolism, but lower levels of defense-related genes. Our results suggest that conidia of A. brassicicola are programmed as saprophytes, but become parasites upon contact with their hosts. AbPf2 coordinates this transformation by expressing pathogenesis-associated genes, including those coding for effectors.