scholarly journals The Ralstonia solanacearum Type III Effector RipAY Is Phosphorylated in Plant Cells to Modulate Its Enzymatic Activity

2017 ◽  
Vol 8 ◽  
Author(s):  
Yali Wei ◽  
Yuying Sang ◽  
Alberto P. Macho
Keyword(s):  
Enzymatic Activity ◽  
Ralstonia Solanacearum ◽  
Plant Cells ◽  
Type Iii Effector ◽  
Type Iii

scholarly journals Identification of novel Ralstonia solanacearum type III effector proteins through translocation analysis of hrpB-regulated gene products

Microbiology ◽  
2009 ◽  
Vol 155 (7) ◽  
pp. 2235-2244 ◽  
Author(s):  
Takafumi Mukaihara ◽  
Naoyuki Tamura
Keyword(s):  
Ralstonia Solanacearum ◽  
Pathogenic Bacteria ◽  
Tandem Repeats ◽  
Plant Cells ◽  
Secretion System ◽  
Effector Proteins ◽  
Gene Products ◽  
Type Iii Effector ◽  
Type Iii ◽  
Effector Genes

The Hrp type III secretion system (TTSS) is essential for the pathogenicity of Ralstonia solanacearum on host plants. Hrp TTSS is a specialized secretion system that injects virulence proteins, the so-called type III effector proteins, into plant cells. In R. solanacearum, the expression of Hrp TTSS-related genes is regulated by an AraC-type transcriptional activator, HrpB. We have identified 30 hrpB-regulated hpx ( hrpB-dependent expression) genes and three well-known hrpB-regulated genes, popA, popB and popC, as candidate effector genes in R. solanacearum strain RS1000. In this study, we newly cloned 11 additional candidate effector genes that share homology with known hpx genes from R. solanacearum RS1000. Using a Cya reporter system, we investigated the translocation of these 44 gene products into plant cells via the Hrp TTSS and identified 34 effector proteins. These include three effector families composed of more than four members, namely the Hpx4, Hpx30 and GALA families. The Hpx30 family effectors are 2200–2500 aa in size and appear to be the largest class of effector proteins among animal- and plant-pathogenic bacteria. Members of this family contain 12–18 tandem repeats of a novel 42 aa motif, designated SKWP repeats.

scholarly journals A Ralstonia solanacearum Type III Effector Directs the Production of the Plant Signal Metabolite Trehalose-6-Phosphate

mBio ◽  
2014 ◽  
Vol 5 (6) ◽  
Author(s):  
M. Poueymiro ◽  
A. C. Cazalé ◽  
J. M. François ◽  
J. L. Parrou ◽  
N. Peeters ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Host Plant ◽  
Ralstonia Solanacearum ◽  
Plant Cells ◽  
Secretion System ◽  
Plant Metabolism ◽  
Signal Molecule ◽  
Content Type ◽  
Type Iii ◽  
Type 3

ABSTRACT  The plant pathogen Ralstonia solanacearum possesses two genes encoding a trehalose-6-phosphate synthase (TPS), an enzyme of the trehalose biosynthetic pathway. One of these genes, named ripTPS, was found to encode a protein with an additional N-terminal domain which directs its translocation into host plant cells through the type 3 secretion system. RipTPS is a conserved effector in the R. solanacearum species complex, and homologues were also detected in other bacterial plant pathogens. Functional analysis of RipTPS demonstrated that this type 3 effector synthesizes trehalose-6-phosphate and identified residues essential for this enzymatic activity. Although trehalose-6-phosphate is a key signal molecule in plants that regulates sugar status and carbon assimilation, the disruption of ripTPS did not alter the virulence of R. solanacearum on plants. However, heterologous expression assays showed that this effector specifically elicits a hypersensitive-like response on tobacco that is independent of its enzymatic activity and is triggered by the C-terminal half of the protein. Recognition of this effector by the plant immune system is suggestive of a role during the infectious process. IMPORTANCE Ralstonia solanacearum, the causal agent of bacterial wilt disease, infects more than two hundred plant species, including economically important crops. The type III secretion system plays a major role in the pathogenicity of this bacterium, and approximately 70 effector proteins have been shown to be translocated into host plant cells. This study provides the first description of a type III effector endowed with a trehalose-6-phosphate synthase enzymatic activity and illustrates a new mechanism by which the bacteria may manipulate the plant metabolism upon infection. In recent years, trehalose-6-phosphate has emerged as an essential signal molecule in plants, connecting plant metabolism and development. The finding that a bacterial pathogen could induce the production of trehalose-6-phosphate in plant cells further highlights the importance of this metabolite in multiple aspects of the molecular physiology of plants.

scholarly journals Functional diversification of the GALA type III effector family contributes to Ralstonia solanacearum adaptation on different plant hosts

2011 ◽  
Vol 192 (4) ◽  
pp. 976-987 ◽  
Author(s):  
Philippe Remigi ◽  
Maria Anisimova ◽  
Alice Guidot ◽  
Stéphane Genin ◽  
Nemo Peeters
Keyword(s):  
Ralstonia Solanacearum ◽  
Functional Diversification ◽  
Type Iii Effector ◽  
Type Iii ◽  
Plant Hosts

scholarly journals Genome-Wide Identification of a Large Repertoire of Ralstonia solanacearum Type III Effector Proteins by a New Functional Screen

2010 ◽  
Vol 23 (3) ◽  
pp. 251-262 ◽  
Author(s):  
Takafumi Mukaihara ◽  
Naoyuki Tamura ◽  
Masaki Iwabuchi
Keyword(s):  
Ralstonia Solanacearum ◽  
Type Iii Secretion ◽  
Plant Cells ◽  
Amino Acid Sequences ◽  
Effector Proteins ◽  
Type Iii ◽  
Functional Screen ◽  
Genome Wide ◽  
Large Repertoire ◽  
Insertion Mutants

The gram-negative plant-pathogenic bacterium Ralstonia solanacearum utilizes the hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS) to cause disease in plants. To determine the entire repertoire of effector proteins possessed by R. solanacearum RS1000, we constructed a transposon carrying a calmodulin-dependent adenylate cyclase reporter that can be used to specifically detect rip (Ralstonia protein injected into plant cells) genes by monitoring the cAMP level in plant leaves inoculated with insertion mutants. From the new functional screen using this transposon, we identified 38 new Rip proteins translocated into plant cells via the Hrp T3SS. In addition, most of the 34 known effectors of RS1000 could be detected by the screen, except for three effectors that appear to be small in size or only weakly expressed. Finally, we identified 72 Rips in RS1000, which include 68 effector proteins classified into over 50 families and four extracellular components of the Hrp T3SS. Interestingly, one-third of the effectors are specific to R. solanacearum. Many effector proteins contain various repeated amino acid sequences or known enzyme motifs. We also show that most of the R. solanacearum effector proteins, but not Hrp extracellular components, require an Hrp-associated protein, HpaB, for their effective translocation into plant cells.

scholarly journals Ralstonia solanacearum Type III Effector RipAY Is a Glutathione-Degrading Enzyme That Is Activated by Plant Cytosolic Thioredoxins and Suppresses Plant Immunity

mBio ◽  
2016 ◽  
Vol 7 (2) ◽  
Author(s):  
Takafumi Mukaihara ◽  
Tadashi Hatanaka ◽  
Masahito Nakano ◽  
Kenji Oda
Keyword(s):  
Immune System ◽  
Ralstonia Solanacearum ◽  
Plant Immunity ◽  
Plant Cells ◽  
Effector Proteins ◽  
Yeast Cells ◽  
Yeast Growth ◽  
Type Iii Effector ◽  
Plant Immune System ◽  
Degradation Activity

ABSTRACT The plant pathogen Ralstonia solanacearum uses a large repertoire of type III effector proteins to succeed in infection. To clarify the function of effector proteins in host eukaryote cells, we expressed effectors in yeast cells and identified seven effector proteins that interfere with yeast growth. One of the effector proteins, RipAY, was found to share homology with the ChaC family proteins that function as γ-glutamyl cyclotransferases, which degrade glutathione (GSH), a tripeptide that plays important roles in the plant immune system. RipAY significantly inhibited yeast growth and simultaneously induced rapid GSH depletion when expressed in yeast cells. The in vitro GSH degradation activity of RipAY is specifically activated by eukaryotic factors in the yeast and plant extracts. Biochemical purification of the yeast protein identified that RipAY is activated by thioredoxin TRX2. On the other hand, RipAY was not activated by bacterial thioredoxins. Interestingly, RipAY was activated by plant h -type thioredoxins that exist in large amounts in the plant cytosol, but not by chloroplastic m -, f -, x -, y - and z -type thioredoxins, in a thiol-independent manner. The transient expression of RipAY decreased the GSH level in plant cells and affected the flg22-triggered production of reactive oxygen species (ROS) and expression of pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) marker genes in Nicotiana benthamiana leaves. These results indicate that RipAY is activated by host cytosolic thioredoxins and degrades GSH specifically in plant cells to suppress plant immunity. IMPORTANCE Ralstonia solanacearum is the causal agent of bacterial wilt disease of plants. This pathogen injects virulence effector proteins into host cells to suppress disease resistance responses of plants. In this article, we report a biochemical activity of R. solanacearum effector protein RipAY. RipAY can degrade GSH, a tripeptide that plays important roles in the plant immune system, with its γ-glutamyl cyclotransferase activity. The high GSH degradation activity of RipAY is considered to be a good weapon for this bacterium to suppress plant immunity. However, GSH also plays important roles in bacterial tolerance to various stresses and growth. Interestingly, RipAY has an excellent safety mechanism to prevent unwanted firing of its enzyme activity in bacterial cells because RipAY is specifically activated by host eukaryotic thioredoxins. This study also reveals a novel host plant protein acting as a molecular switch for effector activation.

Secretion and translocation signals and DspB/F-binding domains in the type III effector DspA/E of Erwinia amylovora

Microbiology ◽  
2010 ◽  
Vol 156 (4) ◽  
pp. 1211-1220 ◽  
Author(s):  
Chang-Sik Oh ◽  
Sara C. D. Carpenter ◽  
Marshall L. Hayes ◽  
Steven V. Beer
Keyword(s):  
Amino Acids ◽  
Erwinia Amylovora ◽  
Active Form ◽  
Bacterial Pathogen ◽  
Plant Cells ◽  
Binding Motif ◽  
Type Iii Effector ◽  
Type Iii ◽  
Binding Domains ◽  
Chaperone Binding

DspA/E is a type III effector of Erwinia amylovora, the bacterial pathogen that causes fire blight disease in roseaceous plants. This effector is indispensable for disease development, and it is translocated into plant cells. A DspA/E-specific chaperone, DspB/F, is necessary for DspA/E secretion and possibly for its translocation. In this work, DspB/F-binding sites and secretion and translocation signals in the DspA/E protein were determined. Based on yeast two-hybrid assays, DspB/F was found to bind DspA/E within the first 210 amino acids of the protein. Surprisingly, both DspB/F and OrfA, the putative chaperone of Eop1, also interacted with the C-terminal 1059 amino acids of DspA/E; this suggests another chaperone-binding site. Secretion and translocation assays using serial N-terminal lengths of DspA/E fused with the active form of AvrRpt2 revealed that at least the first 109 amino acids, including the first N-terminal chaperone-binding motif and DspB/F, were required for efficient translocation of DspA/E, although the first 35 amino acids were sufficient for its secretion and the presence of DspB/F was not required. These results indicate that secretion and translocation signals are present in the N terminus of DspA/E, and that at least one DspB/F-binding motif is required for efficient translocation into plant cells.

Sign in / Sign up

Export Citation Format

Share Document