Clathrin-dependent endocytosis is required for immunity mediated by pattern recognition receptor kinases

Abstract

Sensing of potential pathogenic bacteria is of critical importance for immunity. In plants, this involves plasma membrane-resident pattern recognition receptors, one of which is the FLAGELLIN SENSING 2 (FLS2) receptor kinase. Ligand-activated FLS2 receptors are internalized into endosomes. However, the extent to which these spatiotemporal dynamics are generally present among pattern recognition receptors (PRRs) and their regulation remain elusive. Using live-cell imaging, we show that at least three other receptor kinases associated with plant immunity, PEP RECEPTOR 1/2 (PEPR1/2) and EF-TU RECEPTOR (EFR), internalize in a ligand-specific manner. In all cases, endocytosis requires the coreceptor BRI1-ASSOCIATED KINASE 1 (BAK1), and thus depends on receptor activation status. We also show the internalization of liganded FLS2, suggesting the transport of signaling competent receptors. Trafficking of activated PRRs requires clathrin and converges onto the same endosomal vesicles that are also shared with the hormone receptor BRASSINOSTERIOD INSENSITIVE 1 (BRI1). Importantly, clathrin-dependent endocytosis participates in plant defense against bacterial infection involving FLS2-mediated stomatal closure and callose deposition, but is uncoupled from activation of the flagellin-induced oxidative burst and MAP kinase signaling. In conclusion, immunity mediated by pattern recognition receptors depends on clathrin, a critical component for the endocytosis of signaling competent receptors into a common endosomal pathway.pattern-triggered immunity | clathrin | FLS2 | PEPR1 | EFR

Clathrin in plant defense signaling and execution

“…An interacting coreceptor, BRI1-ASSOCIATED KINASE 1 (BAK1), is necessary for the internalization of all these activated receptors. Requirement for the active RK domain was demonstrated for the FLS2 PRK, but this might be a more general feature also valid for other activated PRK internalization mechanisms (7).…”

“…Until recently, there was only very limited insight into what happens to PRKs in plant cells upon the arrival of the signal: that is, after specific ligand binding resulting in an intracellular kinase domain activation and signaling initiation. Two side-byside reports in PNAS (7,8) show that three different PRKs [PEP receptor 1 (PEPR1), EF-TU RECEPTOR (EFR), and FLAGELLIN SENSING 2 (FLS2)], involved in biotic defense interactions activated by three different ligands, are all removed from the cell surface via clathrin-mediated endocytosis (CME). An interacting coreceptor, BRI1-ASSOCIATED KINASE 1 (BAK1), is necessary for the internalization of all these activated receptors.…”

“…In a pathway seemingly bypassing the TGN/EE, all three internalized ligand-bound receptors converge on the same ARA7/RabF2b marker-positive late-endosomal prevacuolar compartment (PVC), which is identical to the multivesicular body (MVB) and destined finally for the vacuole (7,8) (Fig. 1).…”

“…1). Interestingly, temporal dynamics for endocytotic internalization of the steroid receptor BRI1 were speedier when compared with pathogen-related pattern recognition ligandbound FLS2 or PEPR1 receptors, but the final destination was the same, indicating a possible existence of the late-endosomal compartment as a general depot for internalized activated RKs in plant cells (6)(7)(8).…”

“…Testing the sensitivity of Arabidopsis mutant plants with compromised CME toward infection by different Pseudomonas bacteria strains was straightforward in the case of activated FLS2-dependent defense reactions (7). Plants with disturbed CME are more sensitive to Pseudomonas than WT, regardless of the presence or absence of Pseudomonas functional type III secretion system (important for bacterial effector injection into the host cells) or coronatin production (reopening stomata to support infection).…”