Ionotropic glutamate receptor (iGluR)-like channels mediate MAMP-induced calcium influx in Arabidopsis thaliana

International audienceBinding of specific microbial epitopes (microbe-associated molecular patterns, MAMPs) to pattern recognition receptors (PRRs) and subsequent receptor kinase activation are key steps in plant innate immunity. One of the earliest detectable events after MAMP perception is a rapid and transient rise in cytosolic calcium (Ca2+) levels. In plants, knowledge about the signaling events leading to Ca2+ influx and on the molecular identity of the channels involved is scarce. We used a transgenic Arabidopsis thaliana line stably expressing the luminescent aequorin Ca2+ biosensor to monitor pharmacological interference with Ca2+ signatures following treatment with the bacterial peptide MAMPs flg22 and elf18 and the fungal carbohydrate MAMP chitin. Using a comprehensive set of compounds known to impede Ca2+ transport processes in plants and animals we found strong evidence for a prominent role of amino acid-controlled Ca2+ fluxes, probably through ionotropic glutamate receptor (iGluR)-like channels. Interference with amino acid-mediated Ca2+ fluxes modulates MAMP-triggered mitogen-activated protein kinase (MAPK) activity and affects MAMP-induced accumulation of defense gene transcripts. We conclude that the initiation of innate immune responses upon flg22, elf18 and chitin recognition involves apoplastic Ca2+ influx via iGluR-like channels

Combined Bimolecular Fluorescence Complementation and Förster Resonance Energy Transfer Reveals Ternary SNARE Complex Formation in Living Plant Cells1[W][OA]

Various fluorophore-based microscopic methods, comprising Förster resonance energy transfer (FRET) and bimolecular fluorescence complementation (BiFC), are suitable to study pairwise interactions of proteins in living cells. The analysis of interactions between more than two protein partners using these methods, however, remains difficult. In this study, we report the successful application of combined BiFC-FRET-fluorescence lifetime imaging microscopy and BiFC-FRET-acceptor photobleaching measurements to visualize the formation of ternary soluble N-ethylmaleimide-sensitive factor attachment receptor complexes in leaf epidermal cells. This method expands the repertoire of techniques to study protein-protein interactions in living plant cells by a procedure capable of visualizing simultaneously interactions between three fluorophore-tagged polypeptide partners

Arabidopsis SOMATIC EMBRYOGENESIS RECEPTOR KINASE Proteins Serve Brassinosteroid-Dependent and -Independent Signaling Pathways1[C][W]

The Arabidopsis (Arabidopsis thaliana) SOMATIC EMBRYOGENESIS RECEPTOR KINASE (SERK) genes belong to a small family of five plant receptor kinases that are involved in at least five different signaling pathways. One member of this family, BRASSINOSTEROID INSENSITIVE1 (BRI1)-ASSOCIATED KINASE1 (BAK1), also known as SERK3, is the coreceptor of the brassinolide (BR)-perceiving receptor BRI1, a function that is BR dependent and partially redundant with SERK1. BAK1 (SERK3) alone controls plant innate immunity, is also the coreceptor of the flagellin receptor FLS2, and, together with SERK4, can mediate cell death control, all three in a BR-independent fashion. SERK1 and SERK2 are essential for male microsporogenesis, again independent from BR. SERK5 does not appear to have any function under the conditions tested. Here, we show that the different SERK members are only redundant in pairs, whereas higher order mutant combinations only show additive phenotypes. Surprisingly, SERK members that are redundant within one are not redundant in another pathway. We also show that this evolution of functional pairs occurred by a change in protein function and not by differences in spatial expression. We propose that, in plants, closely related receptor kinases have a minimal homo- or heterodimeric configuration to achieve specificity

A component of the Sec61 ER protein transporting pore is required for plant susceptibility to powdery mildew

Biotrophic pathogens, like the powdery mildew fungi, require living plant cells for their growth and reproduction. During infection, a specialized structure called the haustorium is formed by the fungus. The haustorium is surrounded by a plant cell-derived extrahaustorial membrane (EHM). Over the EHM, the fungus obtains nutrients from and secretes effectors into the plant cell. In the plant cell these effectors interfere with cellular processes such as pathogen defense and membrane trafficking. However, the mechanisms behind effector delivery are largely unknown. This paper provides a model for and new insights into a putative transfer mechanism of effectors into the plant cell. We show that silencing of the barley Sec61βa protein results in decreased susceptibility to the powdery mildew fungus. HvSec61βa is a component of both the endoplasmic reticulum (ER) translocon and retrotranslocon pores, the latter being part of the ER-associated protein degradation machinery (ERAD). We provide support for a model suggesting that the retrotranslocon function of HvSec61βa is required for successful powdery mildew fungal infection. HvSec61βa-GFP and a luminal ER marker were co-localized to the ER, which was found to be in close proximity to the EHM around the haustorial body, but not the haustorial fingers. This differential EHM proximity suggests that the ER, including HvSec61βa, may be actively recruited by the haustorium, potentially to provide efficient effector transfer to the cytosol. Effector transport across this EHM-ER interface may occur by a vesicle-mediated process, while the Sec61 retrotranslocon pore potentially provides an escape route for these proteins to reach the cytosol