Biology subjects

Bortlik, J.

Publications and source records attributed to Bortlik, J..

3 recordsLinked to original sources

bioRxiv2024

XopM, a FFAT motif containing type-III effector protein from Xanthomonas, suppresses PTI responses at the plant plasma membrane

Many Gram-negative pathogenic bacteria use type-III effector proteins (T3Es) as essential virulence factors to suppress host immunity and to cause disease. However, in many cases the molecular function of T3Es remains unknown. The plant pathogen Xanthomonas campestris pv. vesicatoria (Xcv) is the causal agent of bacterial spot disease on tomato and pepper plants and is known to translocate around 30 T3Es into its host cell, which collectively suppress plant defense and promote infection. XopM is an Xcv core T3E with unknown function that has no similarity to any other known protein. We found that XopM interacts with vesicle-associated membrane protein (VAMP)-associated proteins (VAPs) in an isoform specific manner. The endoplasmic reticulum (ER) integral membrane protein VAP is a common component of membrane contact sites involved in both tethering and lipid transfer by binding directly to proteins containing a FFAT [two phenylalanines (FF) in an acidic tract (AT)] motif. Sequence analyses revealed that XopM displays two FFAT motifs that cooperatively mediated the interaction of XopM with VAP. When expressed in plants, XopM supports growth of a non-pathogenic bacterial strain and dampens the production of reactive oxygen species, indicating its ability to suppress plant immunity. Further analyses revealed that the interaction with VAP and the ability to suppress PTI are structurally and functionally separable. Although XopM requires localization to the host membrane system for full PTI suppression activity. We discuss a working model in which XopM uses FFAT-motifs to target the membrane in order to interfere with early PTI responses.

plant biology↗

bioRxiv2022

DUF581-9 (At2g44670; FLZ3) negatively regulates SnRK1 activity by interference with T-loop phosphorylation

In plants, SUCROSE NON-FERMENTING RELATED KINASE 1 (SnRK1) is a key energy-sensor that orchestrates large-scale transcriptional reprogramming to maintain cellular homeostasis under energy deficit. SnRK1 activity is under tight negative control, although the exact mechanisms leading to its activation are far from being understood. We show that the Arabidopsis DOMAIN OF UNKNOWN FUNCTION (DUF581) protein DUF581-9/FLZ3 binds to the catalytic SnRK1 subunit KIN10 to inhibit its activation by GRIK-dependent T-loop phosphorylation. Overexpression of DUF581-9 in Arabidopsis dampens SnRK1 signalling and interferes with adaptation to dark-induced starvation. The presence of DUF581-9 significantly reduced SnRK1 activity in protoplasts and in vitro. This was accompanied by a reduction in T175 T-loop phosphorylation and also diminished KIN10 auto-phosphorylation. Furthermore, DUF581-9 reduced binding of the up-stream activating kinase GRIK2 to KIN10, explaining the reduced KIN10 T-loop phosphorylation. Ectopically expressed DUF581-9 protein was rapidly turned over by the proteasome when Arabidopsis plants were subjected to starvation treatment, likely releasing its inhibitory activity on the SnRK1 complex. Taken together, our results support a model in which DUF581-9 negatively regulates SnRK1 activity under energy sufficient conditions. Turnover over of the protein provides a rapid way for SnRK1 activation under energy deficit without the need of de-novo protein synthesis.

plant biology↗

bioRxiv2019

Prioritization of abiotic and biotic stress responses by direct linkage of ABI1 phosphatase and CPK5 calcium-dependent protein kinase

In nature plants are constantly challenged by simultaneous abiotic and biotic stresses, and under conflicting stress scenarios prioritization of stress responses is required for plant survival. Calcium-dependent protein kinase CPK5 is a central hub in local and distal immune signaling, required for hormone salicylic acid (SA)-dependent immunity and pathogen resistance. Here we show that CPK5-dependent immune responses and pathogen resistance are inhibited upon abscisic acid (ABA) treatment or in genetic mutant backgrounds lacking PP2C phosphatase activities including abi1-2, whereas immune responses are enhanced by co-expression of active ABI1 phosphatase variants. Biochemical studies and mass spectrometry-based phospho-site analysis reveal a direct ABI1 phosphatase-catalyzed de-phosphorylation of CPK5 auto-phosphorylation site T98. Mimicking continuous de-phosphorylation in CPK5T98A leads to enhanced ROS production and more resistant plants, mimicking the auto-phosphorylated status in CPK5T98D, reduces CPK5-mediated immune responses. Mechanistic insight identifies differential phosphorylation at T98 in the N-terminal domain of CPK5 to control the level of interaction between the kinase and its substrate protein rather than CPK5 catalytic activity. Thus, CPK5-catalyzed immune signaling may become discontinued even at an elevated cytoplasmic calcium concentration. Our work reveals an elegant mechanism for stress response prioritization in plants: The ABA-dependent phosphatase ABI1, negative regulator of abiotic responses, functions as positive regulator of biotic stress responses, stabilizing CPK5-dependent immune signaling in the absence of ABA. Continuous pathogen survey activates plant immunity in environmentally friendly conditions, whereas under severe abiotic stress the phosphatase/kinase pair prohibits immune signaling through a direct biochemical switch involving two key regulatory enzymes of these antagonistic pathways. Significance StatementPlants challenged by simultaneous abiotic and biotic stresses must prioritize in conflicting scenarios to guarantee survival. Pathogen resistance and immune memory depends on the phytohormone salicylic acid (SA). Adaptation to abiotic stress signaling involves the phytohormone abscisic acid (ABA). We identify a direct biochemical switch by which ABA-mediated abiotic signaling prioritizes over SA-dependent immune responses via reversible phosphorylation at a single protein mark involving two key regulatory enzymes of these antagonistic pathways. Phosphatase ABI1 de-phosphorylates calcium-dependent protein kinase CPK5 at an auto-phosphorylation site T98, which effects the interaction efficiency between the kinase and its substrate. Under abiotic stress ABA mediates phosphatase inhibition, which facilitates prolonged auto-phosphorylation of CPK5, preventing CPK5 substrate interaction and ultimately stop CPK5-mediated immune signaling.

plant biology↗