Abstract
SummaryCell wall appositions (CWAs) are produced reactively by the plant immune system to arrest microbial invasion through the local inversion of plant cell growth. This process requires the controlled invagination of the plasma membrane (PM) in coordination with the export of barrier material to the volume between the plant PM and cell wall. Plant actin dynamics are essential to this response, but it remains unclear how exocytosis and the cytoskeleton are linked in space and time to form functional CWAs. Here, we show that actin-dependent trafficking to immune response sites of Arabidopsis thaliana delivers membrane-integrated FORMIN4, which in turn contributes to local cytoskeletal dynamics. Total internal reflection fluorescence (TIRF) microscopy combined with controlled induction of FORMIN4-GFP expression reveals a dynamic population of vesicular bodies that accumulate to form clusters at the PM through an actin-dependent process. Deactivation of FORMIN4 and its close homologs partially compromises subsequent defense and alters filamentous actin (F-actin) distribution at mature CWAs. The localization of FORMIN4 is stable and segregated from the dynamic traffic of the endosomal network. Moreover, the tessellation of FORMIN4 at the PM with meso-domains of PEN3 reveals a fine spatial segregation of destinations for actin-dependent immunity cargo. Together, our data suggest a model where FORMIN4 is a spatial feedback element in a multi-layered, temporally defined sequence of cytoskeletal response. This positional feedback makes a significant contribution to the distribution of actin filaments at the dynamic CWA boundary and to the outcomes of pre-invasion defense.
Highlights
We confirmed phytopathogen-responsive transcriptional behavior of the FORMIN4 gene by linking its promoter to the uidA reporter gene and infecting stable transformant A. thaliana with the powdery mildew Blumeria graminis f. sp. hordei (Bgh) (Figure 1A)
The impact of actin-mediated trafficking is apparent during hyphal invasion, where the rapid successful assembly of a focused cell wall apposition (CWA) beneath the microbial appressorium and between the plant plasma membrane (PM) and cell wall prevents hyphal invasion and biotrophic haustorium formation [6]
To gain mechanistic insight into sustained actin-mediated trafficking in response to microbial interactions, we searched for A. thaliana genes encoding known and predicted actin-binding proteins (ABPs)
Summary
We confirmed phytopathogen-responsive transcriptional behavior of the FORMIN4 gene by linking its promoter to the uidA reporter gene and infecting stable transformant A. thaliana with the powdery mildew Blumeria graminis f. sp. hordei (Bgh) (Figure 1A). We confirmed phytopathogen-responsive transcriptional behavior of the FORMIN4 gene by linking its promoter to the uidA reporter gene and infecting stable transformant A. thaliana with the powdery mildew Blumeria graminis f. Bgh is adapted to barley and provokes a non-host response in A. thaliana that is commonly used to identify genes contributing to penetration resistance and preinvasion defense [16, 17]. We made a translational fusion of the complete FORMIN4 gene, under the control of its o
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