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Abiotic and biotic environments influence a myriad of plant-related processes, including growth, development, and the establishment and maintenance of interaction(s) with microbes. As a driver of this signaling between plants and microbes, the role of plant hormones in both surveillance and signaling has emerged as a point of intersection between plant-abiotic and -biotic responses. In the current study, we elucidate a role for NON-RACE-SPECIFIC DISEASE RESISTANCE1 (NDR1) by exploiting effector-triggered immunity (ETI) to define the regulation of plant host immunity in response to both pathogen infection and elevated temperature. We generated time-series RNA sequencing data of WT Col-0, a NDR1 overexpression line, as well as ndr1 and ics1-2 mutant plants under elevated temperature. Not surprisingly, the NDR1-overexpression line showed genotype-specific gene expression changes related to defense response and immune system function. Interestingly, overexpression of NDR1 revealed a role for NDR1 in immune system function; specifically, we describe a mechanism that intersects with Pseudomonas syringae, type-III effector translocation, R-protein signaling complex stabilization, and sustained levels of SA at elevated temperature during ETI. The results described herein support a role for NDR1 in maintaining cell signaling during simultaneous exposure to elevated temperature and avirulent pathogen stressors.One-sentence summaryNDR1 is required for Pst-AvrRpt2 triggered ETI at elevated temperature.

Pattern-triggered immunity and effector-triggered immunity are two primary forms of innate immunity in land plants. The molecular components and connecting nodes of pattern-triggered immunity and effector-triggered immunity are not fully understood. Here, we report that the Arabidopsis calcium-dependent protein kinase CPK3 is a key regulator of both pattern-triggered immunity and effector-triggered immunity. In vitro and in vivo phosphorylation assays, coupled with genetic and cell biology-based analyses, show that actin-depolymerization factor 4 (ADF4) is a physiological substrate of CPK3, and that phosphorylation of ADF4 by CPK3 governs actin cytoskeletal organization associated with pattern-triggered immunity. CPK3 regulates stomatal closure induced by flg22 and is required for resistance to Pst DC3000. Our data further demonstrates that CPK3 is required for resistance to Pst DC3000 carrying the effector AvrPphB. These results suggest that CPK3 is a missing link between cytoskeleton organization, pattern-triggered immunity and effector-triggered immunity., Remodelling of the actin cytoskeleton occurs during plant immune responses to pathogens. Here Lu et al. show that this process requires the calcium-dependent kinase CPK3 which phosphorylates actin depolymerizing factor 4 and is required for both PAMP and effector-triggered immunity in Arabidopsis.

Throughout their life span, plants confront an endless barrage of pathogens and pests. To successfully defend against biotic threats, plants have evolved a complex immune system responsible for surveillance, perception, and the activation of defense. Plant immunity requires multiple signaling processes, the outcome of which vary according to the lifestyle of the invading pathogen(s). In short, these processes require the activation of host perception, the regulation of numerous signaling cascades, and transcriptome reprograming, all of which are highly dynamic in terms of temporal and spatial scales. At the same time, the development of a single immune event is subjective to the development of plant immune system, which is co-regulated by numerous processes, including plant ontogenesis and the host microbiome. In total, insight into each of these processes provides a fuller understanding of the mechanisms that govern plant-pathogen interactions. In this review, we will discuss the "lifecycle" of plant immunity: the development of individual events of defense, including both local and distal processes, as well as the development and regulation of the overall immune system by ontogenesis regulatory genes and environmental microbiota. In total, we will integrate the output of recent discoveries and theories, together with several hypothetical models, to present a dynamic portrait of plant immunity.

NON-RACE-SPECIFIC DISEASE RISISTANCE1 (NDR1) is a key component of plant immune signaling, required for defense against the bacterial pathogen Pseudomonas syringae. Plant stress responses have overlapping molecular, physiological, and cell biology signatures, and given the central role of NDR1 during biotic stress perception and signaling, we hypothesized that NDR1 also functions in abiotic stress responses, including in a role that mediates signaling at the plasma membrane (PM) - cell wall (CW) continuum. Here, we demonstrate that NDR1 is required for the induction of drought stress responses in plants, a role that couples stress signaling in an abscisic acid-dependent manner. We show that NDR1 physically associates with the PM-localized H+-ATPases AHA1, AHA2, and AHA5 and is required for proper regulation of H+-ATPase activity and stomatal guard cell dynamics, providing a mechanistic function of NDR1 during drought responses. In the current study, we demonstrate that NDR1 functions in signaling processes associated with both biotic and abiotic stress response pathways, a function we hypothesize represents NDR1’s role in the maintenance of cellular homeostasis during stress. We propose a role for NDR1 as a core transducer of signaling between cell membrane processes and intercellular stress response activation.

Forecasting spatio-temporal correlated time series of sensor values is crucial in urban applications, such as air pollution alert, biking resource management, and intelligent transportation systems. While recent advances exploit graph neural networks (GNN) to better learn spatial and temporal dependencies between sensors, they cannot model time-evolving spatio-temporal correlation (STC) between sensors, and require pre-defined graphs, which are neither always available nor totally reliable, and target at only a specific type of sensor data at one time. Moreover, since the form of time-series fluctuation is varied across sensors, a model needs to learn fluctuation modulation. To tackle these issues, in this work, we propose a novel GNN-based model, Attention-adjusted Graph Spatio-Temporal Network (AGSTN). In AGSTN, multi-graph convolution with sequential learning is developed to learn time-evolving STC. Fluctuation modulation is realized by a proposed attention adjustment mechanism. Experiments on three sensor data, air quality, bike demand, and traffic flow, exhibit that AGSTN outperforms the state-of-the-art methods., Comment: Published in IEEE ICDM 2020. Code is available at https://github.com/l852888/AGSTN

We evaluated the efficacy of a recombinant adenovirus that expresses a membrane-truncated respiratory syncytial virus (RSV) fusion protein (Ad-F0ΔTM) in newborns via maternal immunization (MI) of pregnant cotton rats. Intranasal Ad-F0ΔTM immunization was given to pregnant female rats, and MI-newborn rats were then challenged intranasally with RSV. Anti-RSV IgGs were observed in the serum of MI-newborn rats after birth. The pulmonary viral loads in Ad-F0ΔTM vs. control vector, Ad-LacZ, and MI-newborns on day 3 post-challenge were reduced by 4 log 10 /g lung. The neutralizing antibody remained for up to 3 weeks in the serum of MI-newborns, which is when weaning began. Ad-F0ΔTM protected MI-newborns from RSV challenge for 1 week. Vertical-transferred protective antibodies were examined in the breast milk and placenta as well. Finally, anti-RSV immunity was not boosted but was only primed during the next RSV exposure in Ad-F0ΔTM-MI-newborns. Maternal Ad-F0ΔTM immunization provides acute protection against RSV infection in neonates.

This paper solves the fake news detection problem under a more realistic scenario on social media. Given the source short-text tweet and the corresponding sequence of retweet users without text comments, we aim at predicting whether the source tweet is fake or not, and generating explanation by highlighting the evidences on suspicious retweeters and the words they concern. We develop a novel neural network-based model, Graph-aware Co-Attention Networks (GCAN), to achieve the goal. Extensive experiments conducted on real tweet datasets exhibit that GCAN can significantly outperform state-of-the-art methods by 16% in accuracy on average. In addition, the case studies also show that GCAN can produce reasonable explanations., To appear in Proceedings of The 58th Annual Meeting of the Association for Computational Linguistics, ACL 2020. Code is available here https://github.com/l852888/GCAN

The plant cytoskeleton underpins the function of a multitude of cellular mechanisms, including those associated with developmental- and stress-associated signaling processes. In recent years, the actin cytoskeleton has been demonstrated to play a key role in plant immune signaling, including a recent demonstration that pathogens target actin filaments to block plant defense and immunity. Herein, we quantified spatial changes in host actin filament organization after infection with Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), demonstrating that the type-III effector HopG1 is required for pathogen-induced changes to actin filament architecture and host disease symptom development during infection. Using a suite of pathogen effector deletion constructs, coupled with high-resolution microscopy, we found that deletion of hopG1 from Pst DC3000 resulted in a reduction in actin bundling and a concomitant increase in the density of filament arrays in Arabidopsis, both of which correlate with host disease symptom development. As a mechanism underpinning this activity, we further show that the HopG1 effector interacts with an Arabidopsis mitochondrial-localized kinesin motor protein. Kinesin mutant plants show reduced disease symptoms after pathogen infection, which can be complemented by actin-modifying agents. In total, our results support a model in which HopG1 induces changes in the organization of the actin cytoskeleton as part of its virulence function in promoting disease symptom development.

Enterovirus 71 (EV71) is a major cause of hand, foot and mouth disease (HFMD). The current EV71 propagating in Vero (EV-V) or sub-passaged in RD (EV-R) cells was used as a pathogen. Interestingly, EV-R exhibited differential virulence; challenging human scavenger receptor class B2-expressing (hSCARB2-Tg) mice with EV71 revealed that EV-V was more virulent than EV-R: 100% of mice that received lethal amounts of EV-V died, while all the mice that received EV-R survived. Severe pathogenesis correlated with viral burdens and proinflammatory cytokine levels were observed in EV-V-challenged mice, but controversy in EV-R-challenged mice. Consensus sequence analysis revealed EV-R rapidly acquired complete mutations at E145G and S241L and partial mutations at V146I of VP1, and acquired a T to C substitution at nucleotide 494 of the 5'-UTR. EV-R exhibited higher binding affinity for another EV71 receptor, human P-selectin glycoprotein ligand-1 (hPSGL-1), than EV-V. Both EV71s exhibited no significant difference in binding to hSCARB2. The molecular modelling indicate that these mutations might influence EV71 engagement with PSGL-1 and in vivo virulence.

Enterovirus 71 (EV71) is a major cause of hand, foot and mouth disease (HFMD). The current EV71 propagating in Vero (EV-V) or sub-passaged in RD (EV-R) cells was used as a pathogen. Interestingly, EV-R exhibited differential virulence; challenging human scavenger receptor class B2-expressing (hSCARB2-Tg) mice with EV71 revealed that EV-V was more virulent than EV-R: 100% of mice that received lethal amounts of EV-V died, while all the mice that received EV-R survived. Severe pathogenesis correlated with viral burdens and proinflammatory cytokine levels were observed in EV-V-challenged mice, but controversy in EV-R-challenged mice. Consensus sequence analysis revealed EV-R rapidly acquired complete mutations at E145G and S241L and partial mutations at V146I of VP1, and acquired a T to C substitution at nucleotide 494 of the 5′-UTR. EV-R exhibited higher binding affinity for another EV71 receptor, human P-selectin glycoprotein ligand-1 (hPSGL-1), than EV-V. Both EV71s exhibited no significant difference in binding to hSCARB2. The molecular modelling indicate that these mutations might influence EV71 engagement with PSGL-1 and in vivo virulence.

Microbial patterns are recognized by cell-surface receptors to initiate pattern-triggered immunity (PTI) in plants. Receptor-like cytoplasmic kinases (RLCKs), such as BIK1, and calcium-dependent protein kinases (CPKs) are engaged during PTI to activate the NADPH oxidase RBOHD for reactive oxygen species (ROS) production. It is unknown whether protein kinases besides CPKs and RLCKs participate in RBOHD regulation. We screened mutants in all ten Arabidopsis MAP4 kinases (MAP4Ks) and identified the conserved MAP4K SIK1 as a positive regulator of PTI. sik1 mutants were compromised in their ability to elicit the ROS burst in response to microbial features and exhibited compromised PTI to bacterial infection. SIK1 directly interacts with, phosphorylates, and stabilizes BIK1 in a kinase activity-dependent manner. Furthermore, SIK1 directly interacts with and phosphorylates RBOHD upon flagellin perception. Thus, SIK1 positively regulates immunity by stabilizing BIK1 and activating RBOHD to promote the extracellular ROS burst.

High spatial and temporal resolution microscopy-based methods are valuable tools for the precise real-time imaging of changes in cellular organization in response to stimulus perception. Here, we describe a quantitative method for the evaluation of the plant actin cytoskeleton during immune stimulus perception and the activation of defense signaling. As a measure of the biotic stress-induced changes in actin filament organization, we present methods for analyzing changes in actin filament organization following elicitation of pattern-triggered immunity and effector-triggered immunity. Using these methods, it is possible to not only quantitatively evaluate changes in actin cytoskeletal organization following biotic stress perception, but to also use these protocols to assess changes in actin filament organization following perception of a wide range of stimuli, including abiotic and developmental cues. As described herein, we present an example application of this method, designed to evaluate changes in actin cytoskeletal organization following pathogen perception and immune signaling.

Adapted filamentous pathogens such as the oomycetes Hyaloperonospora arabidopsidis (Hpa) and Phytophthora infestans (Pi) project specialized hyphae, the haustoria, inside living host cells for the suppression of host defence and acquisition of nutrients. Accommodation of haustoria requires reorganization of the host cell and the biogenesis of a novel host cell membrane, the extrahaustorial membrane (EHM), which envelops the haustorium separating the host cell from the pathogen. Here, we applied live-cell imaging of fluorescent-tagged proteins labelling a variety of membrane compartments and investigated the subcellular changes associated with accommodating oomycete haustoria in Arabidopsis and N. benthamiana. Plasma membrane-resident proteins differentially localized to the EHM. Likewise, secretory vesicles and endosomal compartments surrounded Hpa and Pi haustoria revealing differences between these two oomycetes, and suggesting a role for vesicle trafficking pathways for the pathogen-controlled biogenesis of the EHM. The latter is supported by enhanced susceptibility of mutants in endosome-mediated trafficking regulators. These observations point at host subcellular defences and specialization of the EHM in a pathogen-specific manner. Defence-associated haustorial encasements, a double-layered membrane that grows around mature haustoria, were frequently observed in Hpa interactions. Intriguingly, all tested plant proteins accumulated at Hpa haustorial encasements suggesting the general recruitment of default vesicle trafficking pathways to defend pathogen access. Altogether, our results show common requirements of subcellular changes associated with oomycete biotrophy, and highlight differences between two oomycete pathogens in reprogramming host cell vesicle trafficking for haustoria accommodation. This provides a framework for further dissection of the pathogen-triggered reprogramming of host subcellular changes.

Adapted filamentous pathogens such as the oomycetes Hyaloperonospora arabidopsidis (Hpa) and Phytophthora infestans (Pi) project specialized hyphae, the haustoria, inside living host cells for the suppression of host defence and acquisition of nutrients. Accommodation of haustoria requires reorganization of the host cell and the biogenesis of a novel host cell membrane, the extrahaustorial membrane (EHM), which envelops the haustorium separating the host cell from the pathogen. Here, we applied live-cell imaging of fluorescent-tagged proteins labelling a variety of membrane compartments and investigated the subcellular changes associated with accommodating oomycete haustoria in Arabidopsis and N. benthamiana. Plasma membrane-resident proteins differentially localized to the EHM. Likewise, secretory vesicles and endosomal compartments surrounded Hpa and Pi haustoria revealing differences between these two oomycetes, and suggesting a role for vesicle trafficking pathways for the pathogen-controlled biogenesis of the EHM. The latter is supported by enhanced susceptibility of mutants in endosome-mediated trafficking regulators. These observations point at host subcellular defences and specialization of the EHM in a pathogen-specific manner. Defence-associated haustorial encasements, a double-layered membrane that grows around mature haustoria, were frequently observed in Hpa interactions. Intriguingly, all tested plant proteins accumulated at Hpa haustorial encasements suggesting the general recruitment of default vesicle trafficking pathways to defend pathogen access. Altogether, our results show common requirements of subcellular changes associated with oomycete biotrophy, and highlight differences between two oomycete pathogens in reprogramming host cell vesicle trafficking for haustoria accommodation. This provides a framework for further dissection of the pathogen-triggered reprogramming of host subcellular changes.

To successfully infect plants, filamentous pathogens such as the oomycete Hyaloperonospora arabidopsidis (Hpa) are able to penetrate host tissues and form haustoria, a feeding structure, inside the host cell. Reorganization of the host cell is required to accommodate the haustoria. Formation of haustoria is accompanied by the biogenesis of the extrahaustorial membrane (EHM) which surrounds the haustorium and separates the host cell from the pathogen. In this study, available fluorescent marker protein fusions were used to monitor the re-distribution of membrane compartments at the interface between Arabidopsis and Hpa. The aquaporin PIP1;4, the ATPase ACA8, and the plasma membrane (PM) intrinsic protein NPSN12 were excluded from the EHM while the syntaxin PEN1 and the receptor-like kinase FLS2 labelled the EHM. This suggests PM-resident proteins are recruited to the EHM selectively. The nucleus is always observed adjacent to haustoria, suggesting that the presence of haustoria causes migration of the nucleus. Secretory vesicles and endosomal compartments localize around the haustoria, implicating secretory and endocytic pathways in the biogenesis of the EHM. Upon Hpa infection, haustorial encasements develop around mature haustoria. All examined plant proteins accumulate at haustorial encasements, indicating that formation of encasements is derived by default redirection of vesicle trafficking pathways. With the aim to genetically dissect endosomal trafficking regulators, I took advantage of quantitative high throughput confocal imaging system and transgenic plants containing the fluorescent biosensor GFP-2xFYVE to perform a forward genetic screen. Different numbers of GFP-2xFYVE positive endosomes were found in two reference lines, Ler/GFP-2xFYVE and Col-0/YFP-2xFYVE suggesting the endosomal levels may vary in different ecotypes of Arabidopsis. Mutants with altered numbers of FYVE Endosomal Levels (fel) have been previous identified and were re-confirmed in this study. fel1, fel2, fel3, fel6, fel9, and fel12 revealed genetically recessive mutations while fel10 could not reveal its genetic inheritance. Two mutants, fel2 and fel9 exhibited more GFP-2xFYVE compartments than wild-type reference plants. These two mutants are affected in endosome trafficking and fel2 is likely tissue specific. We identified gene loci by classical mapping and whole genome sequencing. Fel2 co-segregated with the lower arm of chromosome 4. Fel9 was mapped to two chromosome loci. Investigation of genes in the rough mapping region will unravel regulators of endocytosis or multivesicular bodies (MVBs) biogenesis. Because only few mutant phenotypes recovered in the F2 of backcrossed fel2 and fel9, identification of FEL2 and FEL9 was hampered. Additionally, basal differences of endosomal numbers in the reference lines lead to the limitation for genetic screen based on quantitative changes in endosomal numbers. Altogether, these results show that there are common elements in the subcellular changes associated with biotrophic oomycete between different pathogens. For Hpa and other fungal/oomycete pathogens, reprogramming host cell vesicle trafficking occurs to accommodate haustorial structures. A genetic screen for novel endocytosis mutants, based on quantitative measurements of endosomal numbers, was performed with advanced microscopy technology. Fel mutant plants may be further used to study molecular mechanisms for membrane trafficking, as well as subcellular rearrangement in plant-pathogen interactions.