Recent advances in understanding gene –for – gene interactions

1. Recent advances in understanding gene –for – gene interactions

2. Avirulence Gene Function – what is currently known Avr Protein     Pathogen     Homology with and/or possible virulence function * Location in the plant cell Common domain for avirulence and virulence function     Location of the matching R gene * AvrPphC     Pseudomonas syringae pv. phaseolicola Interferes with the plant defense response NK ‡ NK     NK     AvrPphF     Pseudomonas syringae pv. phaseolicola Interferes with the plant defense response NK     NK     NK     AvrRpt2     Pseudomonas syringae pv. tomato Interferes with the plant defense response NK     NK     NK     AvrRpm1     Pseudomonas syringae pv. maculicola Enhances growth Plasma membrane [ NK     Plasma membrane AvrPto     Pseudomonas syringae pv. tomato Enhances growth Degradation of resistance gene Plasma membrane No NK     AvrBst     Xanthomonas campestris pv. campestris     Ubiquitin-like protease NK     Yes NK     AvrXa7     Xanthomonas oryzae pv. oryzae Transcriptional activation domain required for virulence Nucleus Yes NK     AvrBs2     Xanthomonas campestris pv. vesicatoria Agrocinopine synthase (from Agrobacterium tumefaciens)     NK     Recognition region localized in the portion with highest homology to synthase NK     AvrBs3     Xanthomonas campestris pv. glycinea Transcriptional activation domain required for virulence   Nucleus    Yes   NK     PthA     Xanthomonas citri Transcriptional activation domain required for virulence/causes cell hyperplasia [53] Nucleus     NK     NK     Avr9     Cladosporium fulvum NK     Exoplasmic     NK     External plasma membrane    

3. Avirulence genes promote virulence Chen et al., (2000) The Pseudomonas syringae avrRpt2 Gene Product Promotes Pathogen Virulence from Inside Plant Cells. Molecular Plant-Microbe Interactions Vol. 13: 1312–1321 Disease symptoms and growth of Pseudomonas syringae pv. tomato strain DC3000 (PstDC3000) on Arabidopsis thaliana leaf tissue. A, Diseasesymptoms caused by PstDC3000 on A. thaliana No-0 rps2 (top) and Col-0 rps2 (bottom) plants. Leaves are shown 4 days after inoculation withPstDC3000 (left) and PstDC3000(avrRpt2) (right). Plants were inoculated by dipping them into bacterial suspensions (3 to 5 × 10 8 CFU/cm 2 ) containingthe surfactant Silwet L-77. B, Growth of PstDC3000 and PstDC3000(avrRpt2) in leaf tissue of Col-0 rps2, No-0 rps2, and No-0 RPS2 plants.. In B, A. thaliana plants were inoculated by vacuum infiltration with the indicatedPstDC3000 strains at an initial density of 1 × 10 5 CFU/cm 2 . The concentration of bacteria in the plant leaves was assayed after 0, 2, and 4 days. Datapoints represent means of three independent determinations ± standard error of the mean. Experiments presented in A, B, were carried out aminimum of three times with similar results

4. Virulence genes suppress avr/R gene interactions Chen et al., (2000) The Pseudomonas syringae avrRpt2 Gene Product Promotes Pathogen Virulence from Inside Plant Cells. Molecular Plant-Microbe Interactions Vol. 13: 1312–1321

5. Fractionation of RPM1::MYC on sucrose gradients. A 12-55% (wt/vol) linear sucrose gradient was used to fractionate total extract from transgenic plants. Aliquots of each fraction were blotted to nitrocellulose and were analyzed with either anti-c-Myc or the subcellular compartment marker antibodies listed at the right of each panel. RPM1::MYC is a peripheral PM protein. C-Myc epitope has been “artificially” added to RPM1 Anti-myc antibody can be used to detect myc- labelled RPM1 Plasma Membrane Cytoplasm and ER Boyeset al., (1998) The Arabidopsis thaliana RPM1 disease resistance gene product is a peripheral plasma membrane protein that is degraded coincident with the hypersensitive response. 95, PNAS USA 15849-15854

6. AvrRpm1 and avrB localise to the plant cell membrane. Protoplasts from rpm1 plants were transformed with plasmid that express either avr-GFP or avrG2A-GFP constructs. Control transformations were with GFP alone. avrRpm1 N-terminal fatty acylation sequence avrRpm1 MGCVSSTSR Soc3a MGCSVSKKK Ca sensor G2A = glycine change to alanine Loss in avirulence

7. RPM1::MYC is degraded after inoculation with avirulent P. syringae isolates, which trigger LZ-NBS-LRR resistance genes Hours post inoculation Boyeset al., (1998) The Arabidopsis thaliana RPM1 disease resistance gene product is a peripheral plasma membrane protein that is degraded coincident with the hypersensitive response. 95, PNAS USA 15849-15854

8. AvrXa7 : a nuclear targeted protein PX086mx53 = NLS- version Resistant Susceptible NORMAL avrXa7 over express avrXa7 express avrXa7 NLS- NORMAL avrXa7 over express avrXa7 express avrXa7 NLS- Fig. 1.   AvrXa7 is a DNA-binding protein. Double-stranded oligonucleotides were end labeled with 32P, mixed with AvrXa7 or GST (glutathione S-transferase), and subjected to polyacrylamide electrophoresis. Arrow indicates position of well. Yanget al., (2000)The virulence factor AvrXa7 of Xanthomonas oryzae pv. oryzae is a type III secretion pathway-dependent nuclear-localized double-stranded DNA-binding protein.Proc Natl Acad Sci U S A 15;:9807

9. “Guard” Hypotheses