Page 118 - PC2019 Program & Proceedings

PLANT CANADA 2019

S21. Biofilm formation contributes to Pseudomonas syringae pv. tomato success and suppression of
biofilm formation is important for PAMP-triggered immunity in Arabidopsis
*
Nunn, G. ; A. Fufeng; N. Xiao; A. Halim; R. Cameron
McMaster University

Biofilms consist of bacterial cells and associated extracellular poylsaccharides such as alginate,
extracellular (e)DNA, lipids and proteins. Biofilms are thought to protect bacterial pathogens from plant
defense responses. Using the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) and Arabidopsis,
we investigated the role of biofilms in bacterial pathogenicity by asking: Does biofilm formation
contribute to Pst success and does the PAMP-Triggered Immunity (PTI) response limit biofilm formation
in an SA-dependent manner? PTI was induced by flg22 treatment in wild-type Col-0, fls2 (PTI mutant)
and sid2-2 (SA biosynthesis mutant). In vivo bacterial aggregate formation was monitored in leaves using
Pst DC3000 pDSK-GFPuv and epifluorescence microscopy. Pseudomonas aggregate occurrence was
positively correlated with bacterial success in fls2 and sid2-2, while fewer and smaller aggregates were
observed suggesting that Pst aggregate formation was suppressed in an SA-dependent manner during PTI.
In vivo staining with DAPI for eDNA and calcofluor white for polysaccharides, combined with in vitro
bacterial growth and biofilm formation assays with wild-type and algD Pst (alginate biosynthesis
mutant), suggests that Pst aggregates are indeed biofilms (alginate is a common biofilm polysaccharide).
Together these results provide compelling evidence that the ability to form biofilms contributes to Pst
pathogenicity and suppression of biofilm formation is an important component of PTI.

Garrett Nunn (nunngm@mcmaster.ca)

S22. Regulation of plant immune signaling by receptor kinase phosphorylation
4
3
1
2
1
Bender, K. ; D. Couto ; Y. Kadota ; A. Macho ; L. Stransfeld ; C. Zipfel
1
2
1 University of Zurich; University of Geneva
4
3 RIKEN; Chinese Academy of Sciences
Several leucine-rich repeat receptor kinases (LRR-RKs) from the model plant Arabidopsis thaliana
function as cell surface immune receptors which, upon perception of conserved pathogen associated
molecular patterns (PAMPs), activate plant immune responses. The LRR-RK ELONGATION FACTOR
Tu RECEPTOR (EFR) perceives bacterial elongation factor thermo unstable (or its derived peptide elf18)
in concert with the co-receptor LRR-RK BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED
RECEPTOR KINASE 1 (BAK1). Phosphorylation of the EFR cytoplasmic domain is among the earliest
detectable responses to elf18 perception, but the functional significance of receptor phosphorylation is
largely unknown. We reveal that, while EFR is an active protein kinase in vitro, and is phosphorylated in
a PAMP-dependent manner in vivo, kinase-inactive mutants of the receptor are fully functional for elf18-
induced immune responses, indicating a non-catalytic role for the EFR cytoplasmic domain. Analysis of
immuno-purified EFR from elf18-treated plants by LC-MS/MS revealed several Ser and Thr residues
phosphorylated following PAMP perception. To understand the impact of these phosphorylation events
on immune signaling, we generated transgenic plants expressing phospho-null and phospho-mimic site-
directed mutants of EFR and characterized responses to elf18 elicitation. Collectively, our analysis
reveals differential regulation of plant immune signaling by site-specific phosphorylation of EFR. We
propose a model in which site-specific phosphorylation mediates receptor complex assembly/disassembly
during immune signaling and are testing this hypothesis through a combination of quantitative
biochemical and proteomics approaches.

Kyle Bender (kyle.bender@botinst.uzh.ch)

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