Chimeric FLS2 Receptors Reveal the Basis for Differential Flagellin Perception in Arabidopsis and Tomato

Abstract

The flagellin receptor of Arabidopsis thaliana, At-FLAGELLIN SENSING2 (FLS2), has become a model for mechanistic and functional studies on plant immune receptors. Here, we started out with a comparison of At-FLS2 and the orthologous tomato (Solanum lycopersicum) receptor Sl-FLS2. Both receptors specifically responded to picomolar concentrations of the genuine flg22 ligand but proved insensitive to >10 6 -fold higher concentrations of CLV3 peptides that have recently been reported as a second type of ligand for At-FLS2. At-FLS2 and Sl-FLS2 exhibit species-specific differences in the recognition of shortened or sequence-modified flg22 ligands. To map the sites responsible for these species-specific traits on the FLS2 receptors, we performed domain swaps, substituting subsets of the 28 leucine-rich repeats (LRRs) in At-FLS2 with the corresponding LRRs from Sl-FLS2. We found that the LRRs 7 to 10 of Sl-FLS2 determine the high affinity of Sl-FLS2 for the core part RINSAKDD of flg22. In addition, we discovered importance of the LRRs 19 to 24 for the responsiveness to C-terminally modified flagellin peptides. These results indicate that ligand perception in FLS2 is a complex molecular process that involves LRRs from both the outermost and innermost LRRs of the FLS2 ectodomain.

An Overdose of the Arabidopsis Coreceptor BRASSINOSTEROID INSENSITIVE1-ASSOCIATED RECEPTOR KINASE1 or Its Ectodomain Causes Autoimmunity in a SUPPRESSOR OF BIR1-1-Dependent Manner

“…5A) were generated and assayed in mesophyll protoplasts from bak1-4 Arabidopsis. This analysis was done using cotransformation with the BAK1 derivatives and the Luciferase (Luc) reporter gene under the control of the flg22-responsive FRK1 promoter, a well-established and highly sensitive monitoring system for plant immunity (Yoo et al, 2007;Mueller et al, 2012;Albert et al, 2013). Protoplasts cotransformed with GFP (used as a negative control), sECD (for soluble ectodomain of BAK1), and TM-KD or JM-KD (cytoplasmic domains of BAK1) exhibited a low background activity of luciferase.…”

“…Several independent observations indicate that BAK1 and FLS2 are present in close spatial proximity in preformed complexes at the plasma membrane (Chinchilla et al, 2007;Schulze et al, 2010;Roux et al, 2011). Negative regulation of immune signaling prior to ligand perception could happen within the PRR complex and depend on conformational changes following the association of FLS2 with flg22 (Meindl et al, 2000;Schulze et al, 2010;Mueller et al, 2012). Additionally, other partners might prevent the constitutive interaction of BAK1 with FLS2.…”

BR-SIGNALING KINASE1 Physically Associates with FLAGELLIN SENSING2 and Regulates Plant Innate Immunity in Arabidopsis

“…In Arabidopsis thaliana, a conserved 22-amino acid peptide in the N terminus of flagellin (flg22) is specifically recognized by the Leucinerich repeat receptor-like kinase (LRR-RLK) FLAGELLIN SENSING2 (FLS2), and flg22 is sufficient for binding to FLS2 and activation of PTI (Gómez-Gómez and Boller, 2000;Zipfel et al, 2004). FLS2 is present in FLS2-FLS2 complexes in the presence or absence of flg22 (Sun et al, 2012), and ligand perception by FLS2 involves its LRR domains (Mueller et al, 2012). Upon flagellin perception, FLS2 rapidly forms a complex with another LRR-RLK, BRASSI-NOSTEROID INSENSITIVE1 ASSOCIATED KINASE1 (BAK1) (Chinchilla et al, 2007;Schulze et al, 2010), a positive regulator of brassinosteroid (BR) signaling Nam and Li, 2002).…”

Deciphering the Responses of Root Border-Like Cells of Arabidopsis and Flax to Pathogen-Derived Elicitors

“…To this end, we have examined the response of flax and Arabidopsis border-like cells to flagellin22 (flg22) and peptidoglycan (PGN) from Bacillus subtilis, two wellcharacterized MAMPs that are widely used in various plant systems, including Arabidopsis (Felix et al, 1999;Kunze et al, 2004;Mueller et al, 2012). Using microscopical and immunocytochemical techniques in conjunction with specific probes as well as Arabidopsis mutants, we show that border-like cells of both plant species are able to respond specifically to MAMPs by producing ROS, including superoxide (O 2 ·-) and singlet oxygen ( 1 O 2 ), and deposition of callose, a well-known marker of defense.…”