Proteasomal protein turnover during immune priming in Arabidopsis

Principal Investigator: Prof. Dr. Frederik Börnke

Regulated protein turnover mediated by the ubiquitin proteasome system (UPS) is involved in many cellular processes, including immunity. While a role of the UPS in local defence responses in plants appears to be firmly established, its role in systemic immunity and priming for enhanced defence is far less understood.
In preliminary work, we were able to show that Arabidopsis thaliana mutants impaired in proteasome function do not display systemic acquired resistance (SAR) towards a challenge infection of systemic leaves with Pseudomonas syringae after a priming infection of local leaves.
Aim of the project is to understand on the molecular level how UPS activity is mechanistically linked to priming. To this end, the above mentioned Arabidopsis mutants will be subject to a detailed molecular and physiological analysis during a secondary infection in systemic leaves of plants that have been subjected to a priming stimulus by a primary bacterial infection. Read out for systemic defence and priming will be bacterial multiplication in systemic leaves, but also the induction of molecular markers of defence. To identify branches of systemic defence mechanisms that are controlled by the proteasome, primed plants will be extensively profiled before and after triggering infection. This includes the determination of defence hormones (e.g. 326 salicylic acid) and other signalling metabolites known to be involved in SAR. In order to investigate gene regulatory differences between wild type plants and proteasome mutants during priming, whole transcriptome expression analysis will be carried out using RNAseq. All data generated (metabolites and gene expression) will be compared to those obtained from the analysis of locally infected leaves which will aid to identify the steps of priming for enhanced defence affected by reduced proteasome activity. In addition, we will employ a proteomics approach to analyse differential protein ubiquitinylation between wild type plants and proteasome mutants during pathogen priming. Candidate genes and proteins responding differentially in wild type and proteasome mutants will be functionally analysed on the genetic level.

Furthermore, in preliminary work we have identified a WRKY transcription factor and an E3 ubiquitin ligase that together might constitute a regulatory module for SAR. We propose a molecular and biochemical study to uncover how proteasomal turnover of this WKRY transcription factor contributes to SAR.

From an ecological perspective we will investigate how the length of the memory phase during defence priming relates to plant age and whether the bacterial pathogen P. syringae possesses type-III effector proteins that specifically impart a growth advantage during infection of primed host plants.

References

• Bartetzko, V., Sonnewald, S., Vogel, F., Hartner, K., Stadler, R., Hammes, U. Z. and Börnke, F. 2009. The Xanthomonas campestris pv. vesicatoria type III effector protein XopJ inhibits protein secretion: evidence for interference with cell wall – associated defense responses. Mol. Plant-Microbe Int. 22: 655- 664.
• Hoefle, C., Huesmann, C., Schultheiss, H., Börnke, F., Hensel, G., Kumlehn, J. and Hueckelhoven, R. 2011. A barley ROPGTPase ACTIVATING PROTEIN associates with microtubules and regulates entry of the barley powdery mildew fungus into leaf epidermal cells. Plant Cell 23: 2422-2439.
• Lee, A.H., Hurley, B., Felsensteiner, C., Yea, C., Ckurshumova, W., Bartetzko, V., Wang, P.W., Van, Q., Lewis, J.D., Liu, Y.L., Börnke, F., Angers, S., Wilde, A., Guttman, D.S. and Desveaux, D. 2012. A bacterial acetyltransferase destroys plant microtubule networks and blocks secretion. PLoS Pathog. 8: e1002523.
• Üstün, S., Müller, P., Palmisano, P., Hensel, M. and Börnke, F. 2012. SseF, a type III effector protein from the mammalian pathogen Salmonella enterica, requires resistance-gene mediated signalling to activate cell death in the model plant Nicotiana benthamiana. New Phytol. 194: 1046-1060.
• Üstün, S., Bartetzko, V. and Börnke, F. 2013. The Xanthomonas campestris Type III Effector XopJ targets the Host Cell Proteasome to suppress Salicylic-acid mediated Plant Defence. PLoS Pathog. 9: e1003427.
• Üstün, S., König, P., Guttman, D.S. and Börnke, F. 2014. HopZ4 from Pseudomonas syringae, a member of the HopZ type III effector family from the YopJ superfamily, inhibits the proteasome. Mol. Plant Microbe Int. 27: 611-623.
• Üstün, S. and Börnke, F. 2014. Interactions of Xanthomonas type-III effector proteins with the plant ubiquitin system and ubiquitin-like pathways. Front. Plant Sci: doi: 10.3389/fpls.2014.00736.
• Üstün, S. and Börnke, F. 2015. The Xanthomonas campestris type III effector XopJ proteolytically degrades proteasome subunit RPT6. Plant Physiol. 168: 107-119.