A Novel Regulatory Role of HrpD6 in Regulating hrp-hrc-hpa Genes in Xanthomonas oryzae pv. oryzicola

Xanthomonas oryzae pv. oryzicola, the causal agent of bacterial leaf streak in the model plant rice, possesses a hypersensitive response and pathogenicity (hrp), hrp-conserved (hrc), hrp-associated (hpa) cluster (hrp-hrc-hpa) that encodes a type III secretion system (T3SS) through which T3SS effectors are injected into host cells to cause disease or trigger plant defenses. Mutations in this cluster usually abolish the bacterial ability to cause hypersensitive response in nonhost tobacco and pathogenicity in host rice. In Xanthomonas spp., these genes are generally assumed to be regulated by the key master regulators HrpG and HrpX. However, we present evidence that, apart from HrpG and HrpX, HrpD6 is also involved in regulating the expression of hrp genes. Interestingly, the expression of hpa2, hpa1, hpaB, hrcC, and hrcT is positively controlled by HrpD6. Transcriptional expression assays demonstrated that the expression of the hrcC, hrpD5, hrpE, and hpa3 genes was not completely abolished by hrpG and hrpX mutations. As observed in analysis of their corresponding mutants, HrpG and HrpX exhibit contrasting gene regulation, particularly for hpa2 and hrcT. Other two-component system regulators (Zur, LrpX, ColR/S, and Trh) did not completely inhibit the expression of hrcC, hrpD5, hrpE, and hpa3. Immunoblotting assays showed that the secretion of HrpF, which is an HpaB-independent translocator, is not affected by the mutation in hrpD6. However, the mutation in hrpD6 affects the secretion of an HpaB-dependent TAL effector, AvrXa27. These novel findings suggest that, apart from HrpG and HrpX, HrpD6 plays important roles not only in the regulation of hrp genes but also in the secretion of TAL effectors.

Download Full-text

Regulation of the Type III Secretion System in Phytopathogenic Bacteria

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-19-1159 ◽

2006 ◽

Vol 19 (11) ◽

pp. 1159-1166 ◽

Cited By ~ 134

Author(s):

Xiaoyan Tang ◽

Yanmei Xiao ◽

Jian-Min Zhou

Keyword(s):

Signal Transduction ◽

Hypersensitive Response ◽

Type Iii Secretion ◽

Pathogenic Bacteria ◽

Type Iii Secretion System ◽

Secretion System ◽

Host Cells ◽

Plant Pathogenic Bacteria ◽

Type Iii ◽

Hrp Genes

The type III secretion system (TTSS) is a specialized protein secretion machinery used by numerous gram-negative bacterial pathogens of animals and plants to deliver effector proteins directly into the host cells. In plant-pathogenic bacteria, genes encoding the TTSS were discovered as hypersensitive response and pathogenicity (hrp) genes, because mutation of these genes typically disrupts the bacterial ability to cause diseases on host plants and to elicit hypersensitive response on nonhost plants. The hrp genes and the type III effector genes (collectively called TTSS genes hereafter) are repressed in nutrient-rich media but induced when bacteria are infiltrated into plants or incubated in nutrient-deficient inducing media. Multiple regulatory components have been identified in the plant-pathogenic bacteria regulating TTSS genes under various conditions. In Ralstonia solanacearum, several signal transduction components essential for the induction of TTSS genes in plants are dispensable for the induction in inducing medium. In addition to the inducing signals, recent studies indicated the presence of negative signals in the plant regulating the Pseudomonas syringae TTSS genes. Thus, the levels of TTSS gene expression in plants likely are determined by the interactions of multiple signal transduction pathways. Studies of the hrp regulons indicated that TTSS genes are coordinately regulated with a number of non-TTSS genes.

Download Full-text

Concomitant Regulation by a LacI-Type Transcriptional Repressor XylR on Genes Involved in Xylan and Xylose Metabolism and the Type III Secretion System in Rice Pathogen Xanthomonas oryzae pv. oryzae

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-11-17-0277-r ◽

2018 ◽

Vol 31 (6) ◽

pp. 605-613 ◽

Cited By ~ 1

Author(s):

Yumi Ikawa ◽

Sayaka Ohnishi ◽

Akiko Shoji ◽

Ayako Furutani ◽

Seiji Tsuge

Keyword(s):

Gene Expression ◽

Type Iii Secretion ◽

Type Iii Secretion System ◽

Xylose Isomerase ◽

Secretion System ◽

Xanthomonas Oryzae ◽

Upstream Region ◽

Xylose Metabolism ◽

Type Iii ◽

Hrp Genes

The hypersensitive response and pathogenicity (hrp) genes of Xanthomonas oryzae pv. oryzae, the causal agent of bacterial leaf blight of rice, encode components of the type III secretion system and are essential for virulence. Expression of hrp genes is regulated by two key hrp regulators, HrpG and HrpX; HrpG regulates hrpX and hrpA, and HrpX regulates the other hrp genes on hrpB-hrpF operons. We previously reported the sugar-dependent quantitative regulation of HrpX; the regulator highly accumulates in the presence of xylose, followed by high hrp gene expression. Here, we found that, in a mutant lacking the LacI-type transcriptional regulator XylR, HrpX accumulation and hrp gene expression were high even in the medium without xylose, reaching the similar levels present in the wild type incubated in the xylose-containing medium. XylR also negatively regulated one of two xylose isomerase genes (xylA2 but not xylA1) by binding to the motif sequence in the upstream region of the gene. Xylose isomerase is an essential enzyme in xylose metabolism and interconverts between xylose and xylulose. Our results suggest that, in the presence of xylose, inactivation of XylR leads to greater xylan and xylose utilization and, simultaneously, to higher accumulation of HrpX, followed by higher hrp gene expression in the bacterium.

Download Full-text

Presence of Salmonella pathogenicity island 2 genes in seafood-associated Salmonella serovars and the role of the sseC gene in survival of Salmonella enterica serovar Weltevreden in epithelial cells

Microbiology ◽

10.1099/mic.0.043596-0 ◽

2011 ◽

Vol 157 (1) ◽

pp. 160-168 ◽

Cited By ~ 6

Author(s):

Patit P. Bhowmick ◽

Devananda Devegowda ◽

H. A. Darshanee Ruwandeepika ◽

Iddya Karunasagar ◽

Indrani Karunasagar

Keyword(s):

Clinical Isolate ◽

Type Iii Secretion ◽

Crucial Role ◽

Salmonella Enterica ◽

Pcr Amplification ◽

Pathogenicity Island ◽

Host Cells ◽

Serovar Typhimurium ◽

Systemic Infections

The type III secretion system encoded by the Salmonella pathogenicity island 2 (SPI-2) has a central role in the pathogenesis of systemic infections by Salmonella. Sixteen genes (ssaU, ssaB, ssaR, ssaQ, ssaO, ssaS, ssaP, ssaT, sscB, sseF, sseG, sseE, sseD, sseC, ssaD and sscA) of SPI-2 were targeted for PCR amplification in 57 seafood-associated serovars of Salmonella. The sseC gene of SPI-2 was found to be absent in two isolates of Salmonella enterica serovar Weltevreden, SW13 and SW39. Absence of sseC was confirmed by sequencing using flanking primers. SW13 had only 66 bp sequence of the sseC gene and SW39 had 58 bp sequence of this gene. A clinical isolate, S. Weltevreden – SW3, 10 : r : z6 – was used to construct a deletion mutant for the sseC gene. Significant reduction in the survival of SW3, 10 : r : z6 ΔsseC and natural mutants SW13 and SW39 in HeLa cells suggests that sseC has a crucial role in the intracellular survival of S. Weltevreden. Expression of sseC was upregulated during the intracellular phase of both S. enterica serovar Typhimurium and clinical isolate S. Weltevreden SW3, 10 : r : z6, suggesting a crucial role for this gene in the survival of S. Weltevreden inside host cells.

Download Full-text

Enteropathogenic Escherichia coli (EPEC) Tir Receptor Molecule Does Not Undergo Full Modification When Introduced into Host Cells by EPEC-Independent Mechanisms

Infection and Immunity ◽

10.1128/iai.69.3.1444-1453.2001 ◽

2001 ◽

Vol 69 (3) ◽

pp. 1444-1453 ◽

Cited By ~ 29

Author(s):

Brendan Kenny ◽

Jonathan Warawa

Keyword(s):

Escherichia Coli ◽

Molecular Mass ◽

Type Iii Secretion ◽

Efficiency Factor ◽

Host Cells ◽

Sufficient Information ◽

Phosphorylation Event ◽

Associated Proteins ◽

Phosphate Groups

ABSTRACT Enteropathogenic Escherichia coli (EPEC), like many other gram-negative pathogens, encodes a type III secretion apparatus dedicated to the release of virulence-associated proteins. One such protein, Tir, is translocated into host cells, where it is modified by the addition of phosphate groups, resulting in a number of species with distinct molecular mass. One phosphorylation event, on tyrosine residue 474 of Tir, does not contribute to shifts in molecular mass but is essential for its actin-nucleating function. The role of the nonphosphotyrosine related modifications is unknown. In this paper, we demonstrate, using three different approaches, that Tir does not encode sufficient information to facilitate its complete modification when introduced into host cells in EPEC-independent mechanisms. Each system revealed that Tir is a substrate for a host kinase whose action results in its partial modification to a form similar to one evident in EPEC-infected host cells. Further Tir modification could not be induced by infecting cells with EPEC, suggesting that Tir must be coexpressed with other EPEC factors to enable its full modification within host cells. One approach usedYersinia spp. to deliver Tir into host cells, and this system revealed that Tir secretion and translocation can occur in the absence of the Tir chaperone molecule, CesT (formerly known as OrfU). CesT was found to be an efficiency factor which was not required, unlike in EPEC, for Tir stability, indicating that it may function to guide Tir to the translocation apparatus or maintain it in a secretion-competent form.

Download Full-text

Mutagenesis of 18 Type III Effectors Reveals Virulence Function of XopZPXO99 in Xanthomonas oryzae pv. oryzae

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-23-7-0893 ◽

2010 ◽

Vol 23 (7) ◽

pp. 893-902 ◽

Cited By ~ 68

Author(s):

Congfeng Song ◽

Bing Yang

Keyword(s):

Bacterial Blight ◽

Xanthomonas Oryzae ◽

Host Cells ◽

Lesion Length ◽

Transcription Activator ◽

Tal Effectors ◽

Type Iii ◽

Effector Genes ◽

Acid Protein ◽

Bacterial Blight Of Rice

Xanthomonas oryzae pv. oryzae depends on a type III secretion system (T3SS) to translocate effectors into host cells for its ability to cause bacterial blight of rice. All type III (T3) effectors with known function in X. oryzae pv. oryzae belong to a family of transcription activator-like (TAL) effectors. However, other, non–TAL-related effector genes are present in the genome, although their role in virulence and their mode of action have yet to be elucidated. Here, we report the generation of mutants for 18 non-TAL T3 effector genes and the identification of one that contributes to the virulence of strain PXO99A. XopZPXO99 encodes a predicted 1,414-amino-acid protein of unknown function. PXO99A contains two identical copies of the gene due to a duplication of 212 kb in the genome. Strains with knockout mutations of one copy of XopZPXO99 did not exhibit any visible virulence defect. However, strains with mutations in both copies of XopZPXO99 displayed reduced virulence in terms of lesion length and bacterial multiplication compared with PXO99A. The introduction of one genomic copy of XopZPXO99 restores the mutant to full virulence. Transient expression of XopZPXO99 in Nicotiana benthamiana leaves suppresses host basal defense, which is otherwise induced by a T3SS mutant of PXO99A, suggesting a role for XopZPXO99 in interfering with host innate immunity during X. oryzae pv. oryzae infection. XopZPXO99-related genes are found in all Xanthomonas spp. whose genomic sequences have been determined, suggesting a conserved role for this type of effector gene in pathogenesis of Xanthomonas spp. Our results indicate that XopZPXO99 encodes a novel T3 effector and contributes virulence to X. oryzae pv. oryzae strains for bacterial blight of rice.

Download Full-text

Elucidation of the hrp Clusters of Xanthomonas oryzae pv. oryzicola That Control the Hypersensitive Response in Nonhost Tobacco and Pathogenicity in Susceptible Host Rice

Applied and Environmental Microbiology ◽

10.1128/aem.00511-06 ◽

2006 ◽

Vol 72 (9) ◽

pp. 6212-6224 ◽

Cited By ~ 79

Author(s):

Li-fang Zou ◽

Xing-ping Wang ◽

Yong Xiang ◽

Bing Zhang ◽

Yu-Rong Li ◽

...

Keyword(s):

Hypersensitive Response ◽

Inducible Promoter ◽

Xanthomonas Oryzae ◽

Complementation Test ◽

Bacterial Leaf Streak ◽

Rice Seedlings ◽

Susceptible Host ◽

The Core ◽

Cluster A ◽

Regulatory Cluster

ABSTRACT Xanthomonas oryzae pv. oryzicola, the cause of bacterial leaf streak in rice, possesses clusters of hrp genes that determine its ability to elicit a hypersensitive response (HR) in nonhost tobacco and pathogenicity in host rice. A 27-kb region of the genome of X. oryzae pv. oryzicola (RS105) was identified and sequenced, revealing 10 hrp, 9 hrc (hrp conserved), and 8 hpa (hrp-associated) genes and 7 regulatory plant-inducible promoter boxes. While the region from hpa2 to hpaB and the hrpF operon resembled the corresponding genes of other xanthomonads, the hpaB-hrpF region incorporated an hrpE3 gene that was not present in X. oryzae pv. oryzae. We found that an hrpF mutant had lost the ability to elicit the HR in tobacco and pathogenicity in adult rice plants but still caused water-soaking symptoms in rice seedlings and that Hpa1 is an HR elicitor in nonhost tobacco whose expression is controlled by an hrp regulator, HrpX. Using an Hrp phenotype complementation test, we identified a small hrp cluster containing the hrpG and hrpX regulatory genes, which is separated from the core hrp cluster. In addition, we identified a gene, prhA (plant-regulated hrp), that played a key role in the Hrp phenotype of X. oryzae pv. oryzicola but was neither in the core hrp cluster nor in the hrp regulatory cluster. A prhA mutant failed to reduce the HR in tobacco and pathogenicity in rice but caused water-soaking symptoms in rice. This is the first report that X. oryzae pv. oryzicola possesses three separate DNA regions for HR induction in nonhost tobacco and pathogenicity in host rice, which will provide a fundamental base to understand pathogenicity determinants of X. oryzae pv. oryzicola compared with those of X. oryzae pv. oryzae.

Download Full-text

Role of the Salmonella Pathogenicity Island 1 (SPI-1) Protein InvB in Type III Secretion of SopE and SopE2, Two Salmonella Effector Proteins Encoded Outside of SPI-1

Journal of Bacteriology ◽

10.1128/jb.185.23.6950-6967.2003 ◽

2003 ◽

Vol 185 (23) ◽

pp. 6950-6967 ◽

Cited By ~ 62

Author(s):

Kristin Ehrbar ◽

Andrea Friebel ◽

Samuel I. Miller ◽

Wolf-Dietrich Hardt

Keyword(s):

Type Iii Secretion ◽

Inflammatory Responses ◽

Pathogenicity Island ◽

Effector Proteins ◽

Host Cells ◽

Effector Protein ◽

Dependent Manner ◽

Type Iii ◽

Serovar Typhimurium

ABSTRACT Salmonella enterica subspecies 1 serovar Typhimurium encodes a type III secretion system (TTSS) within Salmonella pathogenicity island 1 (SPI-1). This TTSS injects effector proteins into host cells to trigger invasion and inflammatory responses. Effector proteins are recognized by the TTSS via signals encoded in their N termini. Specific chaperones can be involved in this process. The chaperones InvB, SicA, and SicP are encoded in SPI-1 and are required for transport of SPI-1-encoded effectors. Several key effector proteins, like SopE and SopE2, are located outside of SPI-1 but are secreted in an SPI-1-dependent manner. It has not been clear how these effector proteins are recognized by the SPI-1 TTSS. Using pull-down and coimmunoprecipitation assays, we found that SopE is copurified with InvB, the known chaperone for the SPI-1-encoded effector protein Sip/SspA. We also found that InvB is required for secretion and translocation of SopE and SopE2 and for stabilization of SopE2 in the bacterial cytosol. Our data demonstrate that effector proteins encoded within and outside of SPI-1 use the same chaperone for secretion via the SPI-1 TTSS.

Download Full-text

Essential Role of the SycP Chaperone in Type III Secretion of the YspP Effector

Journal of Bacteriology ◽

10.1128/jb.01021-08 ◽

2008 ◽

Vol 191 (5) ◽

pp. 1703-1715 ◽

Cited By ~ 5

Author(s):

Hiroyuki Matsumoto ◽

Glenn M. Young

Keyword(s):

Amino Acids ◽

Type Iii Secretion ◽

Mammalian Cells ◽

System Analysis ◽

Tyrosine Phosphatase ◽

Host Cells ◽

Type Iii ◽

System A ◽

Minimal Region

ABSTRACT The Ysa type III secretion (T3S) system enhances gastrointestinal infection by Yersinia enterocolitica bv. 1B. One effector protein targeted into host cells is YspP, a protein tyrosine phosphatase. It was determined in this study that the secretion of YspP requires a chaperone, SycP. Genetic analysis showed that deletion of sycP completely abolished the secretion of YspP without affecting the secretion of other Ysps by the Ysa T3S system. Analysis of the secretion and translocation signals of YspP defined the first 73 amino acids to form the minimal region of YspP necessary to promote secretion and translocation by the Ysa T3S system. Function of the YspP secretion/translocation signals was dependent on SycP. Curiously, when YspP was constitutively expressed in Y. enterocolitica bv. 1B, it was recognized and secreted by the Ysc T3S system and the flagellar T3S system. In these cases, the first 21 amino acids were sufficient to promote secretion, and while SycP did enhance secretion, it was not essential. However, neither the Ysc T3S system nor the flagellar T3S system translocated YspP into mammalian cells. This supports a model where SycP confers secretion/translocation specificities for YspP by the Ysa T3S system. A series of biochemical approaches further established that SycP specifically interacts with YspP and protected YspP degradation in the cell prior to secretion. Collectively, the evidence suggests that YspP secretion by the Ysa T3S system is a posttranslational event.

Download Full-text

Glycan-Glycan Interaction Determines Shigella Tropism toward Human T Lymphocytes

mBio ◽

10.1128/mbio.02309-17 ◽

2018 ◽

Vol 9 (1) ◽

Cited By ~ 14

Author(s):

Ilia Belotserkovsky ◽

Katja Brunner ◽

Laurie Pinaud ◽

Alexander Rouvinski ◽

Mariano Dellarole ◽

...

Keyword(s):

T Cells ◽

Plasma Membrane ◽

T Lymphocytes ◽

Immune Cells ◽

Type Iii Secretion ◽

Type Iii Secretion System ◽

Secretion System ◽

Host Cells ◽

Gram Negative

ABSTRACT Direct interactions between bacterial and host glycans have been recently reported to be involved in the binding of pathogenic bacteria to host cells. In the case of Shigella, the Gram-negative enteroinvasive bacterium responsible for acute rectocolitis, such interactions contribute to bacterial adherence to epithelial cells. However, the role of glycans in the tropism of Shigella for immune cells whose glycosylation pattern varies depending on their activation state is unknown. We previously reported that Shigella targets activated, but not nonactivated, human CD4+ T lymphocytes. Here, we show that nonactivated CD4+ T lymphocytes can be turned into Shigella-targetable cells upon loading of their plasma membrane with sialylated glycosphingolipids (also termed gangliosides). The Shigella targeting profile of ganglioside-loaded nonactivated T cells is similar to that of activated T cells, with a predominance of injection of effectors from the type III secretion system (T3SS) not resulting in cell invasion. We demonstrate that gangliosides interact with the O-antigen polysaccharide moiety of lipopolysaccharide (LPS), the major bacterial surface antigen, thus promoting Shigella binding to CD4+ T cells. This binding step is critical for the subsequent injection of T3SS effectors, a step which we univocally demonstrate to be dependent on actin polymerization. Altogether, these findings highlight the critical role of glycan-glycan interactions in Shigella pathogenesis. IMPORTANCE Glycosylation of host cell surface varies with species and location in the body, thus contributing to species specificity and tropism of microorganisms. Cross talk by Shigella, the Gram-negative enteroinvasive bacterium responsible for bacillary dysentery, with its exclusively human host has been extensively studied. However, the molecular determinants of the step of binding to host cells are poorly defined. Taking advantage of the observation that human-activated CD4+ T lymphocytes, but not nonactivated cells, are targets of Shigella, we succeeded in rendering the refractory cells susceptible to targeting upon loading of their plasma membrane with sialylated glycosphingolipids (gangliosides) that are abundantly present on activated cells. We show that interactions between the sugar polar part of gangliosides and the polysaccharide moiety of Shigella lipopolysaccharide (LPS) promote bacterial binding, which results in the injection of effectors via the type III secretion system. Whereas LPS interaction with gangliosides was proposed long ago and recently extended to a large variety of glycans, our findings reveal that such glycan-glycan interactions are critical for Shigella pathogenesis by driving selective interactions with host cells, including immune cells.

Download Full-text

Small Molecule Inhibitors Specifically Targeting the Type III Secretion System of Xanthomonas oryzae on Rice

International Journal of Molecular Sciences ◽

10.3390/ijms20040971 ◽

2019 ◽

Vol 20 (4) ◽

pp. 971 ◽

Cited By ~ 3

Author(s):

Hui Tao ◽

Su-Su Fan ◽

Shan Jiang ◽

Xuwen Xiang ◽

Xiaojing Yan ◽

...

Keyword(s):

Virulence Factors ◽

Bacterial Growth ◽

Hypersensitive Response ◽

Type Iii Secretion ◽

Type Iii Secretion System ◽

Secretion System ◽

Xanthomonas Oryzae ◽

Mrna Levels ◽

Growth And Survival ◽

Type Iii

The initiative strategy for the development of novel anti-microbial agents usually uses the virulence factors of bacteria as a target, without affecting their growth and survival. The type III secretion system (T3SS), one of the essential virulence factors in most Gram-negative pathogenic bacteria because of its highly conserved construct, has been regarded as an effective target that developed new anti-microbial drugs. Xanthomonas oryzae pv. oryzae (Xoo) causes leaf blight diseases and is one of the most important pathogens on rice. To find potential anti-virulence agents against this pathogen, a number of natural compounds were screened for their effects on the T3SS of Xoo. Three of 34 compounds significantly inhibited the promoter activity of the harpin gene, hpa1, and were further checked for their impact on bacterial growth and on the hypersensitive response (HR) caused by Xoo on non-host tobacco plants. The results indicated that treatment of Xoo with CZ-1, CZ-4 and CZ-9 resulted in an obviously attenuated HR without affecting bacterial growth and survival. Moreover, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that the expression of the Xoo T3SS was suppressed by treatment with the three inhibitors. The mRNA levels of representative genes in the hypersensitive response and pathogenicity (hrp) cluster, as well as the regulatory genes hrpG and hrpX, were reduced. Finally, the in vivo test demonstrated that the compounds could reduce the disease symptoms of Xoo on the rice cultivar (Oryza sativa) IR24.

Download Full-text