Dataset Information

Silencing a Chitinase Gene, PstChia1, Reduces Virulence of Puccinia striiformis f. sp. tritici.

ABSTRACT: Chitin is the main component of fungal cell walls, which can be recognized by pattern recognition receptors (PRRs) as pathogen-associated molecular patterns (PAMP). Chitinase in filamentous fungi has been reported to degrade immunogenic chitin oligomers, thereby preventing chitin-induced immune activation. In this study, we identified the chitinase families in 10 fungal genomes. A total of 131 chitinase genes were identified. Among the chitinase families, 16 chitinase genes from Puccinia striiformis f. sp. tritici (Pst) were identified, and the expression of PstChia1 was the highest during Pst infection. Further studies indicated that PstChia1 is highly induced during the early stages of the interaction of wheat and Pst and has chitinase enzyme activity. The silencing of PstChia1 revealed that PstChia1 limited the growth and reduced the virulence of Pst. The expression level of TaPR1 and TaPR2 was induced in PstChia1 knockdown plants, suggesting that PstChia1 is involved in regulating wheat resistance to Pst. Our data suggest that PstChia1 contributes to pathogenicity by interfering with plant immunity and regulating the growth of Pst.

SUBMITTER: Guo J

PROVIDER: S-EPMC10179651 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Silencing a Chitinase Gene, PstChia1, Reduces Virulence of Puccinia striiformis f. sp. tritici.

Guo Jia J Mou Ying Y Li Yuanxing Y Yang Qing Q Wang Xue X Lin Haocheng H Kang Zhensheng Z Guo Jun J

International journal of molecular sciences 20230504 9

Chitin is the main component of fungal cell walls, which can be recognized by pattern recognition receptors (PRRs) as pathogen-associated molecular patterns (PAMP). Chitinase in filamentous fungi has been reported to degrade immunogenic chitin oligomers, thereby preventing chitin-induced immune activation. In this study, we identified the chitinase families in 10 fungal genomes. A total of 131 chitinase genes were identified. Among the chitinase families, 16 chitinase genes from Puccinia stri ...[more]

PMID: 37175921

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Project description:Puccinia striiformis f. sp. tritici (Pst) is an obligate biotrophic fungus that causes extensive damage in wheat. The pathogen is now known to be a heteroecious fungus with an intricate life cycle containing sexual and asexual stages. Orthologues of the STE12 transcription factor that regulate mating and filamentation in Saccharomyces cerevisiae, as well as virulence in other fungi, have been extensively described. Because reliable transformation and gene disruption methods are lacking for Pst, knowledge about the function of its STE12 orthologue is limited. In this study, we identified a putative orthologue of STE12 from Pst in haustoria-enriched transcripts and designated it as PstSTE12. The gene encodes a protein of 879 amino acids containing three helices in the homeodomain, conserved phenylalanine and tryptophan sites, and two C2 /H2 -Zn2+ finger domains. Real-time reverse transcription-polymerase chain reaction (RT-PCR) analyses revealed that the expression of PstSTE12 was highly induced during the early infection stages and peaked during haustorium formation and the pycniospore stage in the aecial host barberry. Subcellular localization assays indicated that PstSTE12 is localized in the nucleus and functions as a transcriptional activator. Yeast one-hybrid assays revealed that PstSTE12 exhibits transcriptional activity, and that its C-terminus is necessary for the activation of transcription. PstSTE12 complemented the mating defect in an α ste12 mutant of S. cerevisiae. In addition, it partially complemented the defects of the Magnaporthe oryzae mst12 mutant in plant infection. Knocking down PstSTE12 via host-induced gene silencing (HIGS) mediated by Barley stripe mosaic virus (BSMV) resulted in a substantial reduction in the growth and spread of hyphae in Pst and weakened the virulence of Pst on wheat. Our results suggest that PstSTE12 probably acts at an intersection participating in the invasion and mating processes of Pst, and provide new insights into the comprehension of the variation of virulence in cereal rust fungi.

Project description:BackgroundPuccinia striiformis f. sp. tritici is an obligate biotrophic pathogen that causes leaf stripe rust on wheat. Although it is critical to understand molecular mechanisms of pathogenesis in the wheat stripe rust fungus for developing novel disease management strategies, little is known about its genome and gene functions due to difficulties in molecular studies with this important pathogen. To identify genes expressed during early infection stages, in this study we constructed a cDNA library with RNA isolated from urediniospores of P. striiformis f. sp. tritici germinated for 10 h.ResultsA total of 4798 ESTs were sequenced from the germinated urediniospore library and assembled into 315 contigs and 803 singletons. About 23.9% and 13.3% of the resulting 1118 unisequences were homologous to functionally characterized proteins and hypothetical proteins, respectively. The rest 62.8% unisequences had no significant homologs in GenBank. Several of these ESTs shared significant homology with known fungal pathogenicity or virulence factors, such as HESP767 of the flax rust and PMK1, GAS1, and GAS2 of the rice blast fungus. We selected six ESTs (Ps28, Ps85, Ps87, Ps259, Ps261, and Ps159) for assaying their expression patterns during urediniospore germination and wheat infection by quantitative real-time PCR. All of them had the highest transcript level in germinated urediniospores and a much less transcript level in un-germinated urediniospores and infected wheat tissues (1-7 dpi). The transcript level of Ps159 increased at later infection stages (6-7 dpi). Our data indicated that these genes were highly expressed in germinated urediniospores and may play important roles in fungal-plant interactions during early infection stages in the wheat stripe rust fungus.ConclusionGenes expressed in germinated urediniospores of P. striiformis f. sp. tritici were identified by EST analysis. Six of them were confirmed by quantitative real-time PCR assays to be highly expressed in germinated urediniospores.

Project description:Stripe rust [caused by Puccinia striiformis Westend. f. sp. tritici Eriks. (Pst)] is a destructive disease of wheat (Triticum aestivum L.) worldwide. Genetic resistance is the preferred method for control and the Yr5 gene, originally identified in Triticum spelta var. album, represents a major resistance (R) gene that confers all-stage resistance to all currently known races of Pst in the United States. To identify transcripts associated with the Yr5-mediated incompatible interaction and the yr5-compatible interaction, the Wheat GeneChip was used to profile the changes occurring in wheat isolines that differed for the presence of the Yr5 gene after inoculation with Pst. This time-course study (6, 12, 24 and 48 h post-inoculation) identified 115 transcripts that were induced during the R-gene-mediated incompatible interaction, and 73 induced during the compatible interaction. Fifty-four transcripts were induced in both interactions and were considered as basal defence transcripts, whilst 61 transcripts were specific to the incompatible interaction [hypersensitive response (HR)-specific transcripts] and 19 were specific to the compatible interaction (biotrophic interaction-specific transcripts). The temporal pattern of transcript accumulation showed a peak at 24 h after infection that may reflect haustorial penetration by Pst at ~16 h. An additional 12 constitutive transcript differences were attributed to the presence of Yr5 after eliminating those considered as incomplete isogenicity. Annotation of the induced transcripts revealed that the presence of Yr5 resulted in a rapid and amplified resistance response involving signalling pathways and defence-related transcripts known to occur during R-gene-mediated responses, including protein kinase signalling and the production of reactive oxygen species leading to a hypersensitive response. Basal defence also involved substantial induction of many defence-related transcripts but the lack of R-gene signalling resulted in weaker response.

Project description:BACKGROUND:The stripe rust pathogen, Puccinia striiformis f. sp. tritici (Pst), threats world wheat production. Resistance to Pst is often overcome by pathogen virulence changes, but the mechanisms of variation are not clearly understood. To determine the role of mutation in Pst virulence changes, in previous studies 30 mutant isolates were developed from a least virulent isolate using ethyl methanesulfonate (EMS) mutagenesis and phenotyped for virulence changes. The progenitor isolate was sequenced, assembled and annotated for establishing a high-quality reference genome. In the present study, the 30 mutant isolates were sequenced and compared to the wide-type isolate to determine the genomic variation and identify candidates for avirulence (Avr) genes. RESULTS:The sequence reads of the 30 mutant isolates were mapped to the wild-type reference genome to identify genomic changes. After selecting EMS preferred mutations, 264,630 and 118,913 single nucleotide polymorphism (SNP) sites and 89,078 and 72,513 Indels (Insertion/deletion) were detected among the 30 mutant isolates compared to the primary scaffolds and haplotigs of the wild-type isolate, respectively. Deleterious variants including SNPs and Indels occurred in 1866 genes. Genome wide association analysis identified 754 genes associated with avirulence phenotypes. A total of 62 genes were found significantly associated to 16 avirulence genes after selection through six criteria for putative effectors and degree of association, including 48 genes encoding secreted proteins (SPs) and 14 non-SP genes but with high levels of association (P ≤ 0.001) to avirulence phenotypes. Eight of the SP genes were identified as avirulence-associated effectors with high-confidence as they met five or six criteria used to determine effectors. CONCLUSIONS:Genome sequence comparison of the mutant isolates with the progenitor isolate unraveled a large number of mutation sites along the genome and identified high-confidence effector genes as candidates for avirulence genes in Pst. Since the avirulence gene candidates were identified from associated SNPs and Indels caused by artificial mutagenesis, these avirulence gene candidates are valuable resources for elucidating the mechanisms of the pathogen pathogenicity, and will be studied to determine their functions in the interactions between the wheat host and the Pst pathogen.

Dataset Information

Silencing a Chitinase Gene, PstChia1, Reduces Virulence of Puccinia striiformis f. sp. tritici.

Publications

Silencing a Chitinase Gene, <i>PstChia1</i>, Reduces Virulence of <i>Puccinia striiformis</i> f. sp. <i>tritici</i>.

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