Genetic Diversity of a Brazilian Strain Collection of Xanthomonas citri subsp. citri Based on the Type III Effector Protein Genes

Exclusion and eradication or management based on an integrated approach with less susceptible varieties, copper-based bactericides, and windbreaks are the two main strategies used to prevent or control citrus canker. Field tolerance or resistance to citrus canker is not found in the most important commercial sweet orange cultivars, and pathogen-derived resistance has been developed and applied in different crops to obtain resistant genotypes to plant pathogens. We describe the development of DNA primers and probes based on the type III effector genes avrXacE1, avrXacE2, avrXacE3, avrBs2, pthA4, hpaF, and XAC3090 (leucine rich protein), and their application in the evaluation of the genetic diversity of the pathogen. A total of 49 haplotypes were identified in 157 strains by Southern blot analysis. No genetic polymorphism was detected by BOX elements - and enterobacterial repetitive intergenic consensus–polymerase chain reaction (ERIC-PCR) analysis, nor with the genes avrBs2, XAC3090, and hpaF. Nei's genetic diversity indexes varied from 0.65 to 0.96 for subcollections of the pathogen. One or few haplotypes were most frequent in the strain collection, but several haplotypes were represented by solely one or few strains. The PthA4 probe resulted in the higher number of haplotypes identified in the Brazilian subcollections. Greater variation in the frequency of haplotypes occurred within subcollections (93.7%) than among subcollections. Only some haplotypes were genetically distant from all others, especially those originated from Rio Grande do Sul and Santa Catarina states. These bacterial effectors are widely spread in the collections and are useful for a better understanding of the host–pathogen interaction and the search for resistance genes in host and nonhost plants.

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Crystal Structure-Based Exploration of Arginine-Containing Peptide Binding in the ADP-Ribosyltransferase Domain of the Type III Effector XopAI Protein

International Journal of Molecular Sciences ◽

10.3390/ijms20205085 ◽

2019 ◽

Vol 20 (20) ◽

pp. 5085 ◽

Cited By ~ 1

Author(s):

Jyung-Hurng Liu ◽

Jun-Yi Yang ◽

Duen-Wei Hsu ◽

Yi-Hua Lai ◽

Yun-Pei Li ◽

...

Keyword(s):

Crystal Structure ◽

Drug Targets ◽

Plant Pathogens ◽

Peptide Binding ◽

Target Protein ◽

Dynamics Simulation ◽

Type Iii Effector ◽

Type Iii ◽

Pathogen Invasion ◽

Adp Ribosyltransferase

Plant pathogens secrete proteins called effectors into the cells of their host to modulate the host immune response against colonization. Effectors can either modify or arrest host target proteins to sabotage the signaling pathway, and therefore are considered potential drug targets for crop disease control. In earlier research, the Xanthomonas type III effector XopAI was predicted to be a member of the arginine-specific mono-ADP-ribosyltransferase family. However, the crystal structure of XopAI revealed an altered active site that is unsuitable to bind the cofactor NAD+, but with the capability to capture an arginine-containing peptide from XopAI itself. The arginine peptide consists of residues 60 through 69 of XopAI, and residue 62 (R62) is key to determining the protein–peptide interaction. The crystal structure and the molecular dynamics simulation results indicate that specific arginine recognition is mediated by hydrogen bonds provided by the backbone oxygen atoms from residues W154, T155, and T156, and a salt bridge provided by the E265 sidechain. In addition, a protruding loop of XopAI adopts dynamic conformations in response to arginine peptide binding and is probably involved in target protein recognition. These data suggest that XopAI binds to its target protein by the peptide-binding ability, and therefore, it promotes disease progression. Our findings reveal an unexpected and intriguing function of XopAI and pave the way for further investigation on the role of XopAI in pathogen invasion.

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Pseudomonas syringae type III effector HopAF1 suppresses plant immunity by targeting methionine recycling to block ethylene induction

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1606322113 ◽

2016 ◽

Vol 113 (25) ◽

pp. E3577-E3586 ◽

Cited By ~ 22

Author(s):

Erica J. Washington ◽

M. Shahid Mukhtar ◽

Omri M. Finkel ◽

Li Wan ◽

Mark J. Banfield ◽

...

Keyword(s):

Pseudomonas Syringae ◽

Plant Pathogens ◽

Plant Immunity ◽

Ethylene Biosynthesis ◽

Host Protein ◽

Loss Of Function ◽

Type Iii Effector ◽

Type Iii ◽

Host Defense Response ◽

Molecular Patterns

HopAF1 is a type III effector protein of unknown function encoded in the genomes of several strains of Pseudomonas syringae and other plant pathogens. Structural modeling predicted that HopAF1 is closely related to deamidase proteins. Deamidation is the irreversible substitution of an amide group with a carboxylate group. Several bacterial virulence factors are deamidases that manipulate the activity of specific host protein substrates. We identified Arabidopsis methylthioadenosine nucleosidase proteins MTN1 and MTN2 as putative targets of HopAF1 deamidation. MTNs are enzymes in the Yang cycle, which is essential for the high levels of ethylene biosynthesis in Arabidopsis. We hypothesized that HopAF1 inhibits the host defense response by manipulating MTN activity and consequently ethylene levels. We determined that bacterially delivered HopAF1 inhibits ethylene biosynthesis induced by pathogen-associated molecular patterns and that Arabidopsis mtn1 mtn2 mutant plants phenocopy the effect of HopAF1. Furthermore, we identified two conserved asparagines in MTN1 and MTN2 from Arabidopsis that confer loss of function phenotypes when deamidated via site-specific mutation. These residues are potential targets of HopAF1 deamidation. HopAF1-mediated manipulation of Yang cycle MTN proteins is likely an evolutionarily conserved mechanism whereby HopAF1 orthologs from multiple plant pathogens contribute to disease in a large variety of plant hosts.

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Cutting the line: manipulation of plant immunity by bacterial type III effector proteases

Journal of Experimental Botany ◽

10.1093/jxb/erab095 ◽

2021 ◽

Vol 72 (9) ◽

pp. 3395-3409

Author(s):

Brian C Mooney ◽

Melissa Mantz ◽

Emmanuelle Graciet ◽

Pitter F Huesgen

Keyword(s):

Immune Response ◽

Plant Immunity ◽

Host Cells ◽

Secretion Systems ◽

Peptide Bonds ◽

Type Iii Effector ◽

Type Iii ◽

Protease Cleavage ◽

Type Iii Secretion Systems ◽

Bacterial Effectors

Abstract Pathogens and their hosts are engaged in an evolutionary arms race. Pathogen-derived effectors promote virulence by targeting components of a host’s innate immune system, while hosts have evolved proteins that sense effectors and trigger a pathogen-specific immune response. Many bacterial effectors are translocated into host cells using type III secretion systems. Type III effector proteases irreversibly modify host proteins by cleavage of peptide bonds and are prevalent among both plant and animal bacterial pathogens. In plants, the study of model effector proteases has yielded important insights into the virulence mechanisms employed by pathogens to overcome their host’s immune response, as well as into the mechanisms deployed by their hosts to detect these effector proteases and counteract their effects. In recent years, the study of a larger number of effector proteases, across a wider range of pathogens, has yielded novel insights into their functions and recognition. One key limitation that remains is the lack of methods to detect protease cleavage at the proteome-wide level. We review known substrates and mechanisms of plant pathogen type III effector proteases and compare their functions with those of known type III effector proteases of mammalian pathogens. Finally, we discuss approaches to uncover their function on a system-wide level.

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Identification and Functional Analysis of Type III Effector Proteins in Mesorhizobium loti

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-23-2-0223 ◽

2010 ◽

Vol 23 (2) ◽

pp. 223-234 ◽

Cited By ~ 46

Author(s):

Shin Okazaki ◽

Saori Okabe ◽

Miku Higashi ◽

Yoshikazu Shimoda ◽

Shusei Sato ◽

...

Keyword(s):

Pseudomonas Syringae ◽

Rhizobium Leguminosarum ◽

Plant Pathogens ◽

Effector Proteins ◽

Sequence Motif ◽

Shikimate Kinase ◽

Model Legume ◽

Type Iii Effector ◽

Mesorhizobium Loti ◽

Type Iii

Mesorhizobium loti MAFF303099, a microsymbiont of the model legume Lotus japonicus, possesses a cluster of genes (tts) that encode a type III secretion system (T3SS). In the presence of heterologous nodD from Rhizobium leguminosarum and a flavonoid naringenin, we observed elevated expression of the tts genes and secretion of several proteins into the culture medium. Inoculation experiments with wild-type and T3SS mutant strains revealed that the presence of the T3SS affected nodulation at a species level within the Lotus genus either positively (L. corniculatus subsp. frondosus and L. filicaulis) or negatively (L. halophilus and two other species). By inoculating L. halophilus with mutants of various type III effector candidate genes, we identified open reading frame mlr6361 as a major determinant of the nodulation restriction observed for L. halophilus. The predicted gene product of mlr6361 is a protein of 3,056 amino acids containing 15 repetitions of a sequence motif of 40 to 45 residues and a shikimate kinase-like domain at its carboxyl terminus. Homologues with similar repeat sequences are present in the hypersensitive-response and pathogenicity regions of several plant pathogens, including strains of Pseudomonas syringae, Ralstonia solanacearum, and Xanthomonas species. These results suggest that L. halophilus recognizes Mlr6361 as potentially pathogen derived and subsequently halts the infection process.

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Characterization of Partial Gene Deletions in Type III von Willebrand Disease with Alloantibody Inhibitors

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648835 ◽

1994 ◽

Vol 72 (02) ◽

pp. 180-185 ◽

Cited By ~ 22

Author(s):

David J Mancuso ◽

Elodee A Tuley ◽

Ricardo Castillo ◽

Norma de Bosch ◽

Pler M Mannucci ◽

...

Keyword(s):

Von Willebrand Factor ◽

Von Willebrand Disease ◽

Pcr Analysis ◽

Gene Deletions ◽

Factor Gene ◽

Type Iii ◽

Large Deletions ◽

Von Willebrand ◽

Willebrand Factor

Summaryvon Willebrand factor gene deletions were characterized in four patients with severe type III von Willebrand disease and alloantibodies to von Willebrand factor. A PCR-based strategy was used to characterize the boundaries of the deletions. Identical 30 kb von Willebrand factor gene deletions which include exons 33 through 38 were identified in two siblings of one family by this method. A small 5 base pair insertion (CCTGG) was sequenced at the deletion breakpoint. PCR analysis was used to detect the deletion in three generations of the family, including two family members who are heterozygous for the deletion. In a second family, two type III vWD patients, who are distant cousins, share an -56 kb deletion of exons 22 through 43. The identification and characterization of large vWF gene deletions in these type III vWD patients provides further support for the association between large deletions in both von Willebrand factor alleles and the development of inhibitory alloantibodies.

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