Source: CORNELL UNIVERSITY submitted to NRP
DELIVERY OF EFFECTOR PROTEINS TO PLANT CELLS VIA THE HRP TYPE III PROTEIN SECRETION SYSTEM OF PSEUDOMONAS SYRINGAE
Sponsoring Institution
State Agricultural Experiment Station
Project Status
COMPLETE
Funding Source
STATE
Reporting Frequency
Annual
Accession No.
0187451
Grant No.
(N/A)
Cumulative Award Amt.
(N/A)
Proposal No.
(N/A)
Multistate No.
(N/A)
Project Start Date
May 15, 2000
Project End Date
Apr 30, 2004
Grant Year
(N/A)
Program Code
[(N/A)]- (N/A)
Recipient Organization
CORNELL UNIVERSITY
(N/A)
ITHACA,NY 14853
Performing Department
PLANT PATHOLOGY
Non Technical Summary
Bacterial diseases have a significant impact on many crops. A better understanding of bacterial virulence mechanisms will provide us with new targets for disease control.
Animal Health Component
(N/A)
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
21214601040100%
Knowledge Area
212 - Pathogens and Nematodes Affecting Plants;

Subject Of Investigation
1460 - Tomato;

Field Of Science
1040 - Molecular biology;
Goals / Objectives
The specific objectives of this project are to: 1) Identify targeting signals affecting the differential secretion behaviors of proteins traveling the Hrp pathway, and develop improved assays for detecting Hrp-mediated protein translocation into plant cells. 2) Identify additional extracellular components of the P. syringae Hrp secretion system and the conditions maximizing their production. 3) Determine the contribution of each component of the Hrp system to a panel of molecular phenotypes aimed at dissecting Hrp-mediated effector protein secretion and translocation.
Project Methods
The central event in the parasitism of Pseudomonas syringae and other common gram-negative plant pathogens appears to be the Hrp (type III protein secretion system)-mediated translocation of effector proteins into plant cells. An understanding of effector protein delivery mechanisms is key to understanding bacterial plant pathogenicity and is likely to yield novel therapies. The proposed research will use a combination of biochemical and genetic techniques to investigate the targeting signals in proteins traveling the Hrp pathway and the extracellular components of the Hrp secretion machinery.

Progress 05/15/00 to 04/30/04

Outputs
Type III secretion machines are essential to the pathogenicity of Pseudomonas syringae and many other bacterial pathogens of plants and animals. The hrp/hrc genes encoding the type III system in P. syringae are at the center of a pathogenicity island and are flanked by exchangeable and conserved effector loci. We are exploring the role of various components of the system in effector protein delivery, host range determination, and virulence. Candidate effector genes downstream of Hrp promoters were analyzed for patterns associated with type III effector N-terminal targeting signals and then tested for translocation into plant cells. Effectors that possessed the predictive patterns were found to be translocated. HopPtoN is encoded in the P. syringae pv. tomato DC3000 Hrp pathogenicity island. The protein was shown to be a cysteine protease that is translocated into plant cells where it suppresses plant cell death associated with both compatible and incompatible interactions.

Impacts
Protein secretion systems are essential for the virulence of most bacterial pathogens of both plants and animals, and a better understanding of them should lead to new measures for disease prevention and therapy.

Publications

  • Alfano, J. R., and Collmer, A. 2004. Type III secretion system effector proteins: double agents in bacterial disease and plant defense. Annu. Rev. Phytopathol. 42:385-414.
  • Jamir, Y., Guo, M., Oh, H.-S., Petnicki-Ocwieja, T., Chen, S., Tang, X., Dickman, M. B., Collmer, A., and Alfano, J. R. 2004. Identification of Pseudomonas syringae type III secreted effectors that suppress programmed cell death in plants and yeast. Plant J. 37:554-565.
  • Lopez-Solanilla, E., Bronstein, P. A., Schneider, A. R., and Collmer, A. 2004. HopPtoN is a Pseudomonas syringae Hrp (type III secretion system) cyseine protease effector that suppresses pathogen-induced necrosis associated with both compatible and incompatible plant interactions. Mol. Microbiol. 54:353-365.
  • Preston, G. M., and Collmer, A. 2004. The type III secretion systems of plant-associated pseudomonads: genes and proteins on the move. Pages 181-219 in: Pseudomonas, Volume 2. J.-L. Ramos, ed. Plenum Press, New York.
  • Schechter, L. M., Roberts, K. A., Jamir, Y., Alfano, J. R., and Collmer, A. 2004. Pseudomonas syringae type III secretion system targeting signals and novel effectors studied with a Cya translocation reporter. J. Bacteriol. 186:543-555.


Progress 01/01/03 to 12/31/03

Outputs
Type III secretion machines are essential to the pathogenicity of Pseudomonas syringae and many other bacterial pathogens of plants and animals. The hrp/hrc genes encoding the type III system in P. syringae are at the center of a pathogenicity island and are flanked by exchangeable and conserved effector loci. We are exploring the role of various components of the system in effector protein delivery, host range determination, and virulence. Deleting the hrp/hrc cluster of P. syringae pv. tomato DC3000 completely abolishes the ability of DC3000 to grow or elicit any observable responses in plants. Replacing the gene cluster with that from the bean pathogen P. s. syringae 61 restores the characteristic ability of DC3000 to produce disease in tomato and elicit the HR in tomato, which indicates that the secretion machinery has no direct role in host range determination. HopPtoM is an effector protein encoded by the conserved effector locus that has a homolog outside of this region, is secreted in a Hrp-dependent manner, and is important for the ability of P. syringae pv. tomato DC3000 to cause disease lesions in tomato.

Impacts
Protein secretion systems are essential for the virulence of most bacterial pathogens of both plants and animals, and a better understanding of them should lead to new measures for disease prevention and therapy.

Publications

  • Deng, W.-L., Rehm, A., Charkowski, A., Rojas, C. M., and Collmer, A. 2003. Pseudomonas syringae exchangeable effector loci: sequence diversity in representative pathovars and virulence function in P. syringae pv. syringae B728a. J. Bacteriol. 185:2592-2602.
  • Badel, J. L., Nomura, K., Bandyopadhyay, S., Shimizu, R., Collmer, A., and He, S. Y. 2003. Pseudomonas syringae pv. tomato DC3000 HopPtoM (CEL ORF3) is important for lesion formation but not growth in tomato and is secreted and translocated by the Hrp type III secretion system in a chaperone-dependent manner. Mol. Microbiol. 49:1239-1251.


Progress 01/01/02 to 12/31/02

Outputs
Type III secretion machines are essential to the pathogenicity of Pseudomonas syringae and many other bacterial pathogens of plants and animals. The hrp/hrc genes encoding the type III system in P. syringae are at the center of a pathogenicity island and are flanked by exchangeable and conserved effector loci. We are exploring the role of various components of the system in effector protein delivery, host range determination, and virulence. Deleting the hrp/hrc cluster of P. syringae pv. tomato DC3000 completely abolishes the ability of DC3000 to grow or elicit any observable responses in plants. Replacing the gene cluster with that from the bean pathogen P. s. syringae 61 restores the characteristic ability of DC3000 to produce disease in tomato and elicit the HR in tomato, which indicates that the secretion machinery has no direct role in host range determination. HopPtoA is an effector protein encoded by the conserved effector locus that has a homolog outside of this region, is secreted in a Hrp-dependent manner, and contributes to efficient formation of P. syringae pv. tomato DC3000 colonies in planta.

Impacts
Protein secretion systems are essential for the virulence of most bacterial pathogens of both plants and animals, and a better understanding of them should lead to new measures for disease prevention and therapy.

Publications

  • Badel, J. L., Charkowski, A. O., Deng, W.-L., and Collmer, A. 2002. A gene in the Pseudomonas syringae pv. tomato Hrp pathogenicity island conserved effector locus, hopPtoA1, contributes to efficient formation of bacterial colonies in planta and is duplicated elsewhere in the genome. Mol. Plant-Microbe Interact. 15:1014-1024.
  • Fouts, D. E., Badel, J. L., Ramos, A. R., Rapp, R. A., and Collmer, A. 2002. A Pseudomonas syringae pv. tomato DC3000 Hrp (type III secretion) deletion mutant expressing the Hrp system of bean pathogen P. syringae pv. syringae 61 retains normal host specificity for tomato. Mol. Plant-Microbe Interact. 16:43-52.


Progress 01/01/01 to 12/31/01

Outputs
Type III secretion machines are essential to the pathogenicity of Pseudomonas syringae and many other bacterial pathogens of plants and animals. The hrp/hrc genes encoding the type III system in P. syringae are at the center of a pathogenicity island and are flanked by exchangeable and conserved effector loci. We are exploring the role of various components of the system in effector protein delivery, host range determination, and virulence. Deleting the hrp/hrc cluster of P. syringae pv. tomato DC3000 completely abolishes the ability of DC3000 to grow or elicit any observable responses in plants. Replacing the gene cluster with that from the bean pathogen P. s. syringae 61 restores the characteristic ability of DC3000 to produce disease in tomato and elicit the HR in tomato, which indicates that the secretion machinery has no direct role in host range determination. Deletion of the 5 genes in the conserved effector locus also abolishes pathogenicity. We have determined that much of that phenotype can be attributed to ORF3. Interestingly, complementation of the ORF3 mutation with a plasmid-borne ORF3 gene substantially increases the size of disease lesions without enhancing growth of the bacteria in planta.

Impacts
Protein secretion systems are essential for the virulence of most bacterial pathogens of both plants and animals, and a better understanding of them should lead to new measures for disease prevention and therapy.

Publications

  • Alfano, J. R., and A. Collmer. 2001. Mechanisms of bacterial pathogenesis in plants: Familiar foes in a foreign kingdom, p. 176-226. In E. A. Groisman (ed.), Principles of Bacterial Pathogenesis. Academic Press, San Diego.
  • Lopez-Solanilla, E., A. Llama-Palacios, A. Collmer, F. Garcia-Olmedo, and P. Rodriguez-Palenzuela. 2001. Relative effects on virulence of mutations in the sap, pel and hrp loci of Erwinia chrysanthemi. Mol. Plant-Microbe Interact. 14:386-393.