scholarly journals Chemical Targeting and Manipulation of Type III Secretion in the Phytopathogen Xanthomonas campestris for Control of Disease

2019 ◽  
Vol 86 (3) ◽  
Author(s):  
Le Zhou ◽  
Cheng Wang ◽  
Guo-Hua Wang ◽  
Zai-Wa Wei ◽  
Qiu-Xia Fu ◽  
...  
Keyword(s):  
Disease Control ◽  
Small Molecules ◽  
Type Iii Secretion ◽  
Xanthomonas Campestris ◽  
Black Rot ◽  
Content Type ◽  
Type Iii ◽  
Rot Disease ◽  
Antivirulence Agents ◽  
Further Development

ABSTRACT Xanthomonas campestris pv. campestris is the causative agent of black rot disease in crucifer plants. This Gram-negative bacterium utilizes the type III secretion system (T3SS), encoded by the hrp gene cluster, to aid in its resistance to host defenses and the ability to cause disease. The T3SS injects a set of proteins known as effectors into host cells that come into contact with the bacterium. The T3SS is essential for the virulence and hypersensitive response (HR) of X. campestris pv. campestris, making it a potential target for disease control strategies. Using a unique and straightforward high-throughput screening method, we examined a large collection of diverse small molecules for their potential to modulate the T3SS without affecting the growth of X. campestris pv. campestris. Screening of 13,129 different compounds identified 10 small molecules that had a significant inhibitory influence on T3SS. Moreover, reverse transcription-quantitative PCR (qRT-PCR) assays demonstrated that all 10 compounds repress the expression of the hrp genes. Interestingly, the effect of these small molecules on hrp genes may be through the HpaS and ColS sensor kinase proteins that are key to the regulation of the T3SS in planta. Five of the compounds were also capable of inhibiting X. campestris pv. campestris virulence in a Chinese radish leaf-clipping assay. Furthermore, seven of the small molecules significantly weakened the HR in nonhost pepper plants challenged with X. campestris pv. campestris. Taken together, these small molecules may provide potential tool compounds for the further development of antivirulence agents that could be used in disease control of the plant pathogen X. campestris pv. campestris. IMPORTANCE The bacterium Xanthomonas campestris pv. campestris is known to cause black rot disease in many socioeconomically important vegetable crops worldwide. The management and control of black rot disease have been tackled with chemical and host resistance methods with variable success. This has motivated the development of alternative methods for preventing this disease. Here, we identify a set of novel small molecules capable of inhibiting X. campestris pv. campestris virulence, which may represent leading compounds for the further development of antivirulence agents that could be used in the control of black rot disease.

scholarly journals The Predicted Lytic Transglycosylase HpaH from Xanthomonas campestris pv. vesicatoria Associates with the Type III Secretion System and Promotes Effector Protein Translocation

2016 ◽  
Vol 85 (2) ◽  
Author(s):  
Jens Hausner ◽  
Nadine Hartmann ◽  
Michael Jordan ◽  
Daniela Büttner
Keyword(s):  
Type Iii Secretion ◽  
Protein Function ◽  
Xanthomonas Campestris ◽  
Effector Proteins ◽  
Transport Pathway ◽  
Content Type ◽  
Bacterial Membranes ◽  
Type Iii ◽  
Lytic Transglycosylase

ABSTRACT The pathogenicity of the Gram-negative plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria depends on a type III secretion (T3S) system, which spans both bacterial membranes and translocates effector proteins into plant cells. The assembly of the T3S system presumably involves the predicted lytic transglycosylase (LT) HpaH, which is encoded adjacent to the T3S gene cluster. Bacterial LTs degrade peptidoglycan and often promote the formation of membrane-spanning macromolecular protein complexes. In the present study, we show that HpaH localizes to the bacterial periplasm and binds to peptidoglycan as well as to components of the T3S system, including the predicted periplasmic inner rod proteins HrpB1 and HrpB2 as well as the pilus protein HrpE. In vivo translocation assays revealed that HpaH promotes the translocation of various effector proteins and of early substrates of the T3S system, suggesting a general contribution of HpaH to type III-dependent protein export. Mutant studies and the analysis of reporter fusions showed that the N-terminal region of HpaH contributes to protein function and is proteolytically cleaved. The N-terminally truncated HpaH cleavage product is secreted into the extracellular milieu by a yet-unknown transport pathway, which is independent of the T3S system.

scholarly journals Functional Characterization of the Type III Secretion Substrate Specificity Switch Protein HpaC from Xanthomonas campestris pv. vesicatoria

2011 ◽  
Vol 79 (8) ◽  
pp. 2998-3011 ◽  
Author(s):  
Steve Schulz ◽  
Daniela Büttner
Keyword(s):  
Amino Acids ◽  
Substrate Specificity ◽  
Type Iii Secretion ◽  
Protein Function ◽  
Xanthomonas Campestris ◽  
Functional Characterization ◽  
Effector Proteins ◽  
Content Type ◽  
Type Iii ◽  
In The Wild

ABSTRACTPathogenicity ofXanthomonas campestrispv.vesicatoriadepends on a type III secretion (T3S) system which translocates effector proteins into eukaryotic cells and is associated with an extracellular pilus and a translocon in the host plasma membrane. T3S substrate specificity is controlled by the cytoplasmic switch protein HpaC, which interacts with the C-terminal domain of the inner membrane protein HrcU (HrcUC). HpaC promotes the secretion of translocon and effector proteins but prevents the efficient secretion of the early T3S substrate HrpB2, which is required for pilus assembly. In this study, complementation assays with serial 10-amino-acid HpaC deletion derivatives revealed that the T3S substrate specificity switch depends on N- and C-terminal regions of HpaC, whereas amino acids 42 to 101 appear to be dispensable for the contribution of HpaC to the secretion of late substrates. However, deletions in the central region of HpaC affect the secretion of HrpB2, suggesting that the mechanisms underlying HpaC-dependent control of early and late substrates can be uncoupled. The results of interaction and expression studies with HpaC deletion derivatives showed that amino acids 112 to 212 of HpaC provide the binding site for HrcUCand severely reduce T3S when expressed ectopically in the wild-type strain. We identified a conserved phenylalanine residue at position 175 of HpaC that is required for both protein function and the binding of HpaC to HrcUC. Taking these findings together, we concluded that the interaction between HpaC and HrcUCis essential but not sufficient for T3S substrate specificity switching.

scholarly journals The Periplasmic HrpB1 Protein from Xanthomonas spp. Binds to Peptidoglycan and to Components of the Type III Secretion System

2013 ◽  
Vol 79 (20) ◽  
pp. 6312-6324 ◽  
Author(s):  
Jens Hausner ◽  
Nadine Hartmann ◽  
Christian Lorenz ◽  
Daniela Büttner
Keyword(s):  
Type Iii Secretion ◽  
Protein Function ◽  
Xanthomonas Campestris ◽  
Protein Complexes ◽  
Structural Similarity ◽  
Effector Proteins ◽  
Host Cells ◽  
Content Type ◽  
Type Iii ◽  
Inner Membrane Protein

ABSTRACTThe plant-pathogenic bacteriumXanthomonas campestrispv. vesicatoria employs a type III secretion (T3S) system to translocate bacterial effector proteins into eukaryotic host cells. The membrane-spanning secretion apparatus consists of 11 core components and several associated proteins with yet unknown functions. In this study, we analyzed the role of HrpB1, which was previously shown to be essential for T3S and the formation of the extracellular T3S pilus. We provide experimental evidence that HrpB1 localizes to the bacterial periplasm and binds to peptidoglycan, which is in agreement with its predicted structural similarity to the putative peptidoglycan-binding domain of the lytic transglycosylase Slt70 fromEscherichia coli. Interaction studies revealed that HrpB1 forms protein complexes and binds to T3S system components, including the inner membrane protein HrcD, the secretin HrcC, the pilus protein HrpE, and the putative inner rod protein HrpB2. The analysis of deletion and point mutant derivatives of HrpB1 led to the identification of amino acid residues that contribute to the interaction of HrpB1 with itself and HrcD and/or to protein function. The finding that HrpB1 and HrpB2 colocalize to the periplasm and both interact with HrcD suggests that they are part of a periplasmic substructure of the T3S system.

scholarly journals The host exocyst complex is targeted by a conserved bacterial type III effector protein that promotes virulence

2020 ◽  
Author(s):  
Vassiliki A. Michalopoulou ◽  
Konstantinos Kotsaridis ◽  
Glykeria Mermigka ◽  
Dina Kotsifaki ◽  
Michael Kokkinidis ◽  
...  
Keyword(s):  
Type Iii Secretion ◽  
Xanthomonas Campestris ◽  
Plant Immunity ◽  
Gram Negative Bacteria ◽  
Callose Deposition ◽  
Type Iii ◽  
Exocyst Complex ◽  
Successful Infection ◽  
Rot Disease ◽  
Bacterial Type

ABSTRACTFor most Gram-negative bacteria, pathogenicity largely depends on the type-III secretion system that delivers virulence effectors into eukaryotic cells. The subcellular targets for the majority of these effectors remain unknown. Here, we show that Xanthomonas campestris, the causal agent of black rot disease, delivers the highly conserved effector XopP that interacts with host’s EXO70 protein. EXO70 is an essential component of the exocyst complex with a role in plant immunity. The XopP/EXO70 interaction is specific and inhibits exocyst-dependent exocytosis without activating a specific plant NLR receptor that guards EXO70. In this way, Xanthomonas efficiently inhibits the host’s PAMP-triggered immunity (PTI) by blocking exocytosis of PR1, callose deposition and the FLS2 immunity-receptor translocation to the plasma membrane, promoting successful infection.

scholarly journals Natural Genetic Variation of Xanthomonas campestris pv. campestris Pathogenicity on Arabidopsis Revealed by Association and Reverse Genetics

mBio ◽  
2013 ◽  
Vol 4 (3) ◽  
Author(s):  
Endrick Guy ◽  
Anne Genissel ◽  
Ahmed Hajri ◽  
Matthieu Chabannes ◽  
Perrine David ◽  
...  
Keyword(s):  
Xanthomonas Campestris ◽  
Genetic Basis ◽  
Reverse Genetics ◽  
Causal Agent ◽  
Aflp Markers ◽  
Avirulence Gene ◽  
Black Rot ◽  
Virulence Determinants ◽  
Content Type ◽  
Type Iii

ABSTRACTThe pathogenic bacteriumXanthomonas campestrispv. campestris, the causal agent of black rot of Brassicaceae, manipulates the physiology and the innate immunity of its hosts. Association genetic and reverse-genetic analyses of a world panel of 45X. campestrispv. campestris strains were used to gain understanding of the genetic basis of the bacterium’s pathogenicity toArabidopsis thaliana. We found that the compositions of the minimal predicted type III secretome varied extensively, with 18 to 28 proteins per strain. There were clear differences in aggressiveness of thoseX. campestrispv. campestris strains on twoArabidopsisnatural accessions. We identified 3 effector genes (xopAC,xopJ5, andxopAL2) and 67 amplified fragment length polymorphism (AFLP) markers that were associated with variations in disease symptoms. The nature and distribution of the AFLP markers remain to be determined, but we observed a low linkage disequilibrium level between predicted effectors and other significant markers, suggesting that additional genetic factors make a meaningful contribution to pathogenicity. Mutagenesis of type III effectors inX. campestrispv. campestris confirmed thatxopACfunctions as both a virulence and an avirulence gene inArabidopsisand thatxopAMfunctions as a second avirulence gene on plants of the Col-0 ecotype. However, we did not detect the effect of any other effector in theX. campestrispv. campestris 8004 strain, likely due to other genetic background effects. These results highlight the complex genetic basis of pathogenicity at the pathovar level and encourage us to challenge the agronomical relevance of some virulence determinants identified solely in model strains.IMPORTANCEThe identification and understanding of the genetic determinants of bacterial virulence are essential to be able to design efficient protection strategies for infected plants. The recent availability of genomic resources for a limited number of pathogen isolates and host genotypes has strongly biased our research toward genotype-specific approaches. Indeed, these do not consider the natural variation in both pathogens and hosts, so their applied relevance should be challenged. In our study, we exploited the genetic diversity ofXanthomonas campestrispv. campestris, the causal agent of black rot on Brassicaceae (e.g., cabbage), to mine for pathogenicity determinants. This work evidenced the contribution of known and unknown loci to pathogenicity relevant at the pathovar level and identified these virulence determinants as prime targets for breeding resistance toX. campestrispv. campestris in Brassicaceae.

scholarly journals Two Xanthomonas Extracellular Polygalacturonases, PghAxc and PghBxc, Are Regulated by Type III Secretion Regulators HrpX and HrpG and Are Required for Virulence

2008 ◽  
Vol 21 (5) ◽  
pp. 555-563 ◽  
Author(s):  
Lifeng Wang ◽  
Wei Rong ◽  
Chaozu He
Keyword(s):  
Type Iii Secretion ◽  
Xanthomonas Campestris ◽  
Type Ii Secretion ◽  
Dependent Manner ◽  
In Planta ◽  
Cell Wall Degrading Enzymes ◽  
Type Iii ◽  
Type Ii Secretion System ◽  
Rot Disease ◽  
Xanthomonas Campestris Pv Campestris

Xanthomonas campestris pv. campestris, the causal agent of black rot disease, produces a suite of extracellular cell-wall degrading enzymes (CWDE) that are involved in bacterial virulence. Polygalacturonase (PG) is an important CWDE and functions to degrade the pectic layers of plant cell walls. Although previous studies have documented the virulence functions of PG in Erwinia and Ralstonia species, the regulation of PG genes still needs to be elucidated. In this study, we identified two novel PG genes (pghAxc and pghBxc) encoding functional PG from X. campestris pv. campestris 8004. The expressions of these two PG genes are regulated by the type III secretion regulators HrpX and HrpG and the global regulator Clp. These PG genes could be efficiently induced in planta and were required for the full virulence of X. campestris pv. campestris to Arabidopsis. In addition, these PG were confirmed to be secreted via the type II secretion system in an Xps-dependent manner.

scholarly journals A Salmonella Virulence Factor Activates the NOD1/NOD2 Signaling Pathway

mBio ◽  
2011 ◽  
Vol 2 (6) ◽  
Author(s):  
A. Marijke Keestra ◽  
Maria G. Winter ◽  
Daisy Klein-Douwel ◽  
Mariana N. Xavier ◽  
Sebastian E. Winter ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Type Iii Secretion ◽  
Salmonella Enterica ◽  
Intestinal Inflammation ◽  
Inflammatory Responses ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Content Type ◽  
Type Iii

ABSTRACTThe invasion-associated type III secretion system (T3SS-1) ofSalmonella entericaserotype Typhimurium (S. Typhimurium) activates the transcription factor NF-κB in tissue culture cells and induces inflammatory responses in animal models through unknown mechanisms. Here we show that bacterial delivery or ectopic expression of SipA, a T3SS-1-translocated protein, led to the activation of the NOD1/NOD2 signaling pathway and consequent RIP2-mediated induction of NF-κB-dependent inflammatory responses. SipA-mediated activation of NOD1/NOD2 signaling was independent of bacterial invasionin vitrobut required an intact T3SS-1. In the mouse colitis model, SipA triggered mucosal inflammation in wild-type mice but not in NOD1/NOD2-deficient mice. These findings implicate SipA-driven activation of the NOD1/NOD2 signaling pathway as a mechanism by which the T3SS-1 induces inflammatory responsesin vitroandin vivo.IMPORTANCESalmonella entericaserotype Typhimurium (S. Typhimurium) deploys a type III secretion system (T3SS-1) to induce intestinal inflammation and benefits from the ensuing host response, which enhances growth of the pathogen in the intestinal lumen. However, the mechanisms by which the T3SS-1 triggers inflammatory responses have not been resolved. Here we show that the T3SS-1 effector protein SipA induces NF-κB activation and intestinal inflammation by activating the NOD1/NOD2 signaling pathway. These data suggest that the T3SS-1 escalates innate responses through a SipA-mediated activation of pattern recognition receptors in the host cell cytosol.

scholarly journals Genetic Analysis of the Salmonella FliE Protein That Forms the Base of the Flagellar Axial Structure

mBio ◽  
2021 ◽  
Author(s):  
Jordan J. Hendriksen ◽  
Hee Jung Lee ◽  
Alexander J. Bradshaw ◽  
Keiichi Namba ◽  
Fabienne F. V. Chevance ◽  
...  
Keyword(s):  
Self Assembly ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Adaptor Protein ◽  
Ring Structure ◽  
Secretion System ◽  
Dual Roles ◽  
Content Type ◽  
Bacterial Flagellum ◽  
Type Iii

The FliE component of the bacterial flagellum is the first protein secreted through the flagellar type III secretion system (fT3SS) that is capable of self-assembly into the growing bacterial organelle. The FliE protein plays dual roles in the assembly of the Salmonella flagellum as the final component of the flagellar type III secretion system (fT3SS) and as an adaptor protein that anchors the rod (drive shaft) of the flagellar motor to the membrane-imbedded MS-ring structure.

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