Dataset Information

The virulence factor AvrXa7 of Xanthomonas oryzae pv. oryzae is a type III secretion pathway-dependent nuclear-localized double-stranded DNA-binding protein.

ABSTRACT: AvrXa7 is a member of the avrBs3 avirulence gene family, which encodes proteins targeted to plant cells by a type III secretion apparatus. AvrXa7, the product of avrXa7, is also a virulence factor in strain PXO86 of Xanthomonas oryzae pv. oryzae. Avirulence and virulence specificities are associated with the central repeat domain, which, in avrXa7, consists of 25.5 direct repeat units. Mutations in three C-terminal nuclear localization signal motifs eliminated avirulence and virulence activities in rice and severely reduced nuclear localization in a yeast assay system. Both pathogenicity functions and nuclear localization were restored on the addition of the sequence for the nuclear localization signal motif from SV40 T-antigen. The loss of avirulence activity because of mutations in the acidic transcriptional activation domain was restored by addition of the activation domain from the herpes simplex viral protein VP16. The activation domain was also required for virulence activity. However, the VP16 domain could not substitute for the endogenous domain in virulence assays. In gel shift assays, AvrXa7 bound double-stranded DNA with a preference for dA/dT rich sequences. The results indicate that products of the avrBs3-related genes are virulence factors targeted to host cell nuclei and have the potential to interact with the host DNA and transcriptional machinery as part of their mode of action. The results also suggest that the host defensive recognition mechanisms are targeted to the virulence factor site of action.

SUBMITTER: Yang B

PROVIDER: S-EPMC16946 | biostudies-literature | 2000 Aug

REPOSITORIES: biostudies-literature

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The virulence factor AvrXa7 of Xanthomonas oryzae pv. oryzae is a type III secretion pathway-dependent nuclear-localized double-stranded DNA-binding protein.

Yang B B Zhu W W Johnson L B LB White F F FF

Proceedings of the National Academy of Sciences of the United States of America 20000801 17

AvrXa7 is a member of the avrBs3 avirulence gene family, which encodes proteins targeted to plant cells by a type III secretion apparatus. AvrXa7, the product of avrXa7, is also a virulence factor in strain PXO86 of Xanthomonas oryzae pv. oryzae. Avirulence and virulence specificities are associated with the central repeat domain, which, in avrXa7, consists of 25.5 direct repeat units. Mutations in three C-terminal nuclear localization signal motifs eliminated avirulence and virulence activities ...[more]

PMID: 10931960

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Project description:BackgroundWe previously identified a plant-derived phenolic compound ortho-coumaric acid (OCA) as an inhibitor of type III secretion system (T3SS) of Xanthomonas oryzae pv. oryzae (Xoo), the pathogen causing bacterial leaf blight of rice, one of the most devastating bacterial diseases of this staple crop worldwide. However, the molecular mechanisms by which OCA suppresses T3SS and the transcriptional responses to the OCA treatments in Xoo remains unclear.ResultsThe present study conducted the RNA-seq-based transcriptomic analysis to reveal changes in gene expression in Xoo in response to 30 min, 1 h, 3 h, and 6 h of OCA treatment. Results showed that OCA significantly inhibited the expression of T3SS genes after 30 min, and the inhibition also existed after 1 h, 3 h, and 6 h. After treatment for 30 min, membrane proteins in the functional category of cellular process was the predominant group affected, indicating that Xoo was in the early stress stage. Over time, more differentially-expressed genes (DEGs) gathered in the functional category of biological process. Analysis of common DEGs at all four of time points revealed the core elements of Xoo during the response to OCA treatment. Notable, a multidrug transporter cluster that consisted of a MarR-family protein (PXO_RS13760), a multidrug RND transporter (PXO_RS13755), a multidrug transporter (PXO_RS13750), and an MFS transporter (PXO_RS13745) were significantly up-regulated at all four of the time points. Although these three transporter genes were not upregulated by OCA in the PXO_RS13760 deletion mutant, the deficiency of PXO_RS13760 in Xoo did not affect T3SS transcript, and OCA still had the ability to inhibit the expression of T3SS in the mutant, suggesting that the MarR-family protein was involved in bacterial responses to OCA, but not direct OCA inhibition of T3SS in Xoo.ConclusionsWe analyzed the transcriptome of Xoo during OCA treatment at both early and late stages, which revealed the landscape of Xoo responses to OCA at the whole-genome transcription level. A multidrug transporter cluster was identified to be involved in the response process, but had no direct relation to T3SS in Xoo.

Project description:BackgroundPathogenic Gram-negative bacteria interact with their eukaryotic hosts by deploying the type III translocon to inject effector proteins into the cytosol of eukaryotic cells. The translocon compositions, the number and biochemical characteristics of type III translocators in animal-pathogenic bacteria have been well elucidated, but information is lacking for plant-pathogenic bacteria. With extensive studies on biological functions of the Hpa1 protein secreted by the type III secretion system in Xanthomonas oryzae pv. oryzae (Xoo), we show here that Hpa1 is a type III translocator based on measurements of two proteins categorized as transcription activator-like (TAL) effector.ResultsHpa1 was functionally associated with the TAL effector PthXo1 or AvrXa10 by genetic analysis of the wild-type Xoo strain and related mutants or recombinant strains. Inoculation experiments suggested that Hpa1 is required not only for the virulent role of PthXo1 in the susceptible rice variety Nipponbare, but also for the avirulent function of AvrXa10 on the resistant rice variety IRBB10. Hpa1 is unrelated to the secretion of PthXo1 and AvrXa10 out of bacterial cells. However, Hpa1 is critical for both TAL effectors to be translocated from bacterial cells into the cytosol of rice cells based on replicate experiments performed on the susceptible and resistant varieties, respectively. Hpa1-mediated translocation of PthXo1 is coincident with induced expression of rice SWEET11 gene, which is the regulatory target of PthXo1, resulting in the occurrence of the bacterial blight disease in the susceptible rice variety. By contrast, the immune hypersensitive response is induced in agreement with induced expression of rice Xa10 gene, which is the target of AvrXa10, only when AvrXa10 is translocated from bacteria into cells of the resistant rice variety. All the virulent or avirulent performances of the TAL effectors are nullified by directed mutation that removes the α-helix motif from the Hpa1 sequence.ConclusionsThe genetic and biochemical data demonstrate that Hap1 is a type III translocator at least for TAL effectors PthXo1 and AvrXa10. The effect of the directed mutation suggests that Hpa1 depends on its α-helical motif to fulfil the translocator function.

Project description:In Xanthomonas oryzae pv. oryzae, the bacterial blight pathogen of rice, there are over 20 genes encoding GGDEF, EAL, and HD-GYP domains, which are potentially involved in the metabolism of second messenger c-di-GMP. In this study, we focused on the characterization of an EAL domain protein, EdpX1. Deletion of the edpX1 gene resulted in a 2-fold increase in the intracellular c-di-GMP levels, which were restored to the wild-type levels in the complemented ΔedpX1(pB-edpX1) strain, demonstrating that EdpX1 is an active phosphodiesterase (PDE) in X. oryzae pv. oryzae. In addition, colorimetric assays further confirmed the PDE activity of EdpX1 by showing that the E153A mutation at the EAL motif strongly reduced its activity. Virulence assays on the leaves of susceptible rice showed that the ΔedpX1 mutant was severely impaired in causing disease symptoms. In trans expression of wild-type edpX1, but not edpX1 E153A, was able to complement the weakened virulence phenotype. These results indicated that an active EAL domain is required for EdpX1 to regulate the virulence of X. oryzae pv. oryzae. We then demonstrated that the ΔedpX1 mutant was defective in secreting exopolysaccharide (EPS) and forming biofilms. The expression of edpX1 in the ΔedpX1 mutant, but not edpX1 E153A, restored the defective phenotypes to near-wild-type levels. In addition, we observed that EdpX1-green fluorescent protein (EdpX1-GFP) exhibited multiple subcellular localization foci, and this pattern was dependent on its transmembrane (TM) region, which did not seem to directly contribute to the regulatory function of EdpX1. Thus, we concluded that EdpX1 exhibits PDE activity to control c-di-GMP levels, and its EAL domain is necessary and sufficient for its regulation of virulence in X. oryzae pv. oryzae.IMPORTANCE Bacteria utilize c-di-GMP as a second messenger to regulate various biological functions. The synthesis and degradation of c-di-GMP are catalyzed by GGDEF domains and an EAL or HD-GYP domain, respectively. Multiple genes encoding these domains are often found in one bacterial strain. For example, in the genome of X. oryzae pv. oryzae PXO99A, 26 genes encoding proteins containing these domains were identified. Therefore, to fully appreciate the complexity and specificity of c-di-GMP signaling in X. oryzae pv. oryzae, the enzymatic activities and regulatory functions of each GGDEF, EAL, and HD-GYP domain protein need to be elucidated. In this study, we showed that the EAL domain protein EdpX1 is a major PDE to regulate diverse virulence phenotypes through the c-di-GMP signaling pathway.

Dataset Information

The virulence factor AvrXa7 of Xanthomonas oryzae pv. oryzae is a type III secretion pathway-dependent nuclear-localized double-stranded DNA-binding protein.

Publications

The virulence factor AvrXa7 of Xanthomonas oryzae pv. oryzae is a type III secretion pathway-dependent nuclear-localized double-stranded DNA-binding protein.

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