Biology subjects

Zipfel, C.

Publications and source records attributed to Zipfel, C..

53 records · Page 3Linked to original sources

bioRxiv2021

The Arabidopsis immune receptor EFR increases resistance to the bacterial pathogens Xanthomonas and Xylella in transgenic sweet orange

Plants employ cell surface receptors to recognize pathogen (or microbe)-associated molecular patterns (PAMPs/MAMPs), which are crucial for immune system activation. The well-studied Arabidopsis thaliana ELONGATION FACTOR-TU RECEPTOR (EFR) recognizes the conserved bacterial PAMP EF-Tu, and the derived peptides elf18 and elf26. The interfamily transfer of EFR has been shown to increase disease resistance in several crops, such as tomato, rice, wheat, and potato. Here, we generated sweet orange (Citrus sinensis) transgenic lines expressing EFR to test if it would confer broad-spectrum resistance against two important citrus bacterial diseases: citrus canker and citrus variegated chlorosis (CVC). Independent EFR transgenic lines gained responsiveness to elf18 and elf26 peptides from Xanthomonas citri and Xylella fastidiosa, as measured by reactive oxygen species (ROS) production, mitogen-activated protein kinase (MAPK) activation and defense gene expression. Consistently, infection assays showed that Citrus-EFR transgenic plants were more resistant to citrus canker and CVC. Our results show that the EFR immune receptor can improve plant immunity in a perennial crop against bacterial pathogens, opening perspectives to engineer durable broad-spectrum disease resistance under field conditions.

plant biology↗

bioRxiv2021

The Arabidopsis pattern recognition receptor EFR enhances fire blight resistance in apple

Fire blight disease, caused by the bacterium Erwinia amylovora (E. amylovora), is responsible for substantial losses in cultivated apple worldwide. An important mechanism of plant immunity is based on the recognition of conserved microbial molecules, named pathogen- or microbe-associated molecular patterns (PAMPs or MAMPs), through pattern recognition receptors (PRRs), leading to pattern-triggered immunity (PTI). The interspecies transfer of PRRs represents a promising strategy to engineer broad spectrum and durable disease resistance in crops. EFR, the Arabidopsis thaliana PRR for the PAMP elf18 derived from the elongation factor thermal unstable (EF-Tu) proved to be effective in improving bacterial resistance when expressed into Solanaceae and other plant species. In this study, we tested whether EFR can affect the interaction of apple with E. amylovora by its ectopic expression in the susceptible apple rootstock M.26. Stable EFR expression led to the activation of PAMP-triggered immune response in apple leaves upon treatment with supernatant of E. amylovora, as measured by production of reactive oxygen species and the induction of known defense genes. The amount of tissue necrosis associated with E. amylovora infection was significantly reduced in the EFR transgenic rootstock compared to the wild-type. Our results show that the expression of EFR in apple rootstock may be a valuable biotechnology strategy to improve the resistance of apple to fire blight.

plant biology↗

bioRxiv2020

The phagocytosis oxidase/Bem1p (PB1) domain-containing protein PB1CP negatively regulates the NADPH oxidase RBOHD in plant immunity

O_LIPerception of pathogen-associated molecular patterns (PAMPs) by surface-localized pattern-recognition receptors activates RESPIRATORY BURST OXIDASE HOMOLOG D (RBOHD) through direct phosphorylation by BOTRYTIS-INDUCED KINASE 1 (BIK1) and induces the production of reactive oxygen species (ROS). ROS have direct antimicrobial properties but also serve as signaling molecules to activate additional defense responses such as stomatal closure. RBOHD activity must be tightly controlled to avoid the detrimental effects of ROS, but little is known about RBOHD downregulation. C_LIO_LITo better understand the regulation of RBOHD, we used co-immunoprecipitation of RBOHD coupled with mass spectrometry analysis to identify RBOHD-associated proteins. C_LIO_LIAmong RBOHD-associated proteins, we identified PHAGOCYTOSIS OXIDASE/ BEM1P (PB1) DOMAIN-CONTAINING PROTEIN (PB1CP). We found that PB1CP negatively regulates RBOHD and the resistance against the fungal pathogen Colletotrichum higginsianum. PB1CP directly interacts with RBOHD in vitro, and PAMP treatment increases the interaction in vivo. PB1CP is localized at the cell periphery and in cytoplasm, but PAMP treatment induces PB1CP relocalization to small endomembrane compartments. PB1CP overexpression reduces plasma membrane localization of RBOHD, suggesting that PB1CP down-regulates RBOHD function by relocalizing it away from the plasma membrane. C_LIO_LIWe reveal a novel negative regulation mechanism of ROS production through the relocalization of RBOHD by PB1CP. C_LI

plant biology↗

bioRxiv2020

Rhamnolipids and their 3-(3-hydroxyalkanoyloxy)alkanoic acid precursors activate Arabidopsis innate immunity through two independent mechanisms

Plant innate immunity is activated upon perception of invasion pattern molecules by plant cell-surface immune receptors. Several bacteria of the genera Pseudomonas and Burkholderia produce rhamnolipids (RLs) from L- rhamnose and (R)-3-hydroxyalkanoate precursors (HAAs). RL and HAA secretion is required to modulate bacterial surface motility, biofilm development, and thus successful colonization of hosts. Here, we show that the lipidic secretome from the opportunistic pathogen Pseudomonas aeruginosa mostly comprising RLs and HAAs stimulates Arabidopsis immunity. We demonstrate that HAAs are sensed by the bulb-type lectin receptor kinase LIPOOLIGOSACCHARIDE-SPECIFIC REDUCED ELICITATION/S-DOMAIN-1-29 (LORE/SD1-29) that also mediates medium-chain 3-hydroxy fatty acid (mc-3-OH-FA) perception in the plant Arabidopsis thaliana. HAA sensing induces canonical immune signaling and local resistance to plant pathogenic Pseudomonas infection. By contrast, RLs trigger an atypical immune response and resistance to Pseudomonas infection independent of LORE. Thus, the glycosyl moieties of RLs, albeit abolishing sensing by LORE, do not impair their ability to trigger plant defense. In addition, our results show that RL-triggered immune response is affected by the sphingolipid composition of the plasma membrane. In conclusion, RLs and their precursors released by bacteria can both be perceived by plants but through distinct mechanisms. SignificanceActivation of plant innate immunity relies on the perception of microorganisms through self and nonself elicitors. Rhamnolipids and their precursor HAAs are exoproducts produced by beneficial and pathogenic bacteria. They are involved in bacterial surface dissemination and biofilm development. As these compounds are released in the extracellular milieu, they have the potential to be perceived by the plant immune system. Our work shows that both compounds independently activate plant immunity. We demonstrate that HAAs are perceived by the receptor protein kinase LORE. By contrast, rhamnolipids are not senses by LORE but activate a non-canonical immune response affected by the sphingolipid composition of the plant plasma membrane. Thus, plants are able to sense bacterial molecules as well as their direct precursors to trigger a distinct immune response.

plant biology↗

bioRxiv2020

The transcriptional landscape of Arabidopsis thaliana pattern-triggered immunity

Plants initiate immunity upon recognition of a wide array of self and non-self molecular patterns. Whether plants tune their immune outputs to patterns of different biological origins or of different biochemical nature remains mostly unclear. Here, we performed a detailed early time-series transcriptomics analysis in Arabidopsis thaliana, revealing that the response to diverse patterns is remarkably congruent. Early transcriptional reprogramming is dominated by a plant general stress response (GSR), which is essential for pattern-induced immunity. The definition of core immunity response genes common and specific to pattern response in addition revealed the function of previously uncharacterized GLUTAMATE RECEPTOR-LIKE calcium-permeable channels in immunity. This study thus illustrates general and unique properties of early immune transcriptional reprogramming and uncovered important components of plant immunity. One Sentence SummaryTime-resolved transcriptomics reveals new properties of pattern-triggered immunity and function of calcium-permeable channels.

plant biology↗

bioRxiv2020

The Arabidopsis E3 ubiquitin ligase PUB4 regulates BIK1 homeostasis and is targeted by a bacterial type-III effector

Plant immunity is tightly controlled by a complex and dynamic regulatory network, which ensures optimal activation upon detection of potential pathogens. Accordingly, each component of this network is a potential target for manipulation by pathogens. Here, we report that RipAC, a type III-secreted effector from the bacterial pathogen Ralstonia solanacearum, targets the plant E3 ubiquitin ligase PUB4 to inhibit pattern-triggered immunity (PTI). PUB4 plays a positive role in PTI by regulating the homeostasis of the central immune kinase BIK1. Before PAMP perception, PUB4 promotes the degradation of non-activated BIK1, while, after PAMP perception, PUB4 contributes to the accumulation of activated BIK1. RipAC leads to BIK1 degradation, which correlates with its PTI-inhibitory activity. RipAC causes a reduction in pathogen-associated molecular pattern (PAMP)-induced PUB4 accumulation and phosphorylation. Our results shed light on the role played by PUB4 in immune regulation, and illustrate an indirect targeting of the immune signalling hub BIK1 by a bacterial effector.

plant biology↗

bioRxiv2020

Large-scale identification of ubiquitination sites on membrane-associated proteins in Arabidopsis thaliana seedlings

Protein phosphorylation and ubiquitination are two of the most abundant forms of post-translational modifications in eukaryotes, regulated by thousands of protein kinases, phosphatases, E3 ubiquitin ligases, and ubiquitin proteases. Although previous studies have catalogued several ubiquitinated proteins in plants (Walton et al., 2016), few membrane-localized proteins have been identified. Receptor kinases (RKs) initiate phosphorylation signal relays that regulate plant growth, development, and stress responses. While the regulatory role of phosphorylation on protein kinase function is well-documented (Couto and Zipfel, 2016), considerably less is known about the role of ubiquitination on protein kinase function, even though protein turnover is critical to their signaling competence and cellular homeostasis. Here we describe the large-scale identification of ubiquitination sites on Arabidopsis proteins associated with or integral to the plasma membrane, including over 100 protein kinases.

plant biology↗

bioRxiv2020

Family-wide evaluation of RALF peptides in Arabidopsis thaliana

Plant peptide hormones are important players controlling various aspects of plants’ lives. RAPID ALKALINIZATION FACTOR (RALF) peptides have recently emerged as important players in multiple physiological processes. Numerous studies on RALF peptides focused on broad phylogenetic analysis including multiple species. Thus, progress has been made in understanding the evolutionary processes that shaped this family. Nevertheless, to date, there is no comprehensive, family-wide functional study on RALF peptides. Here, we analysed the phylogeny and function of the proposed multigenic RALF peptide family in the model plant Arabidopsis thaliana, ecotype Col-0. Our phylogenetic analysis reveals that two of the previously proposed RALF peptides are not genuine RALF peptides, which leads us to propose a new consensus AtRALF peptide family annotation. Moreover, we show that the majority of AtRALF peptides are able to induce seedling or root growth inhibition in A. thaliana seedlings when applied exogenously as synthetic peptides. Additionally, we show that most of these responses are dependent on the Catharanthus roseus RLK1-LIKE receptor kinase FERONIA, suggesting a pivotal role in the perception of multiple RALF peptides.One sentence summary The majority of A. thaliana RALF peptides inhibit growth in a FERONIA-dependent mannerView Full Text

plant biology↗

bioRxiv2020

Lumi-Map, a real-time luciferase bioluminescence screen of mutants combined with MutMap, reveals Arabidopsis genes involved in PAMP-triggered immunity

Plants recognize pathogen-associated molecular patterns (PAMPs) to activate PAMP-triggered immunity (PTI). However, our knowledge of PTI signaling remains limited. In this report, we introduce Lumi-Map, a high-throughput platform for identifying causative single nucleotide polymorphisms (SNPs) to studying PTI signaling components. In Lumi-Map, a transgenic reporter plant line is produced that contains a firefly luciferase (LUC) gene driven by a defense gene promoter, which generates luminescence upon PAMP treatment. The line is mutagenized and the mutants with altered luminescence patterns are screened by a high-throughput real-time bioluminescence monitoring system. Selected mutants are subjected to MutMap analysis, a whole genome sequencing (WGS)-based method of rapid mutation identification, to identify the causative SNP responsible for the luminescence pattern change. We generated nine transgenic Arabidopsis reporter lines expressing LUC gene fused to multiple promoter sequences of defense-related genes. These lines generate luminescence upon activation of FLAGELLIN-SENSING 2 (FLS2) by flg22, a PAMP derived from bacterial flagellin. We selected the WRKY29-promoter reporter line to identify mutants in the signaling pathway downstream of FLS2. After screening 24,000 ethylmethanesulfonate (EMS)-induced mutants of the reporter line, we isolated 22 mutants with altered WRKY29 expression upon flg22 treatment (abbreviated as awf mutants). While five flg22-insensitive awf mutants harbored mutations in FLS2 itself, Lumi-Map revealed three genes not previously associated with PTI. Lumi-Map has the potential to identify novel PAMPs and their receptors as well as signaling components downstream of the receptors.

plant biology↗

bioRxiv2020

Conserved Molecular Function and Regulatory Subfunctionalization of the LORELEI Gene Family in Brassicaceae

A signaling complex comprising members of the LORELEI (LRE)-LIKE GPI-anchored protein (LLG) and Catharanthus roseus RECEPTOR-LIKE KINASE 1-LIKE (CrRLK1L) families perceive RAPID ALKALINIZATION FACTOR (RALF) peptides and regulate growth, reproduction, immunity, and stress responses in Arabidopsis. Genes encoding these proteins are members of multi-gene families in most angiosperms and could generate thousands of signaling complex variants. However, the link(s) between expansion of these gene families and the functional diversification of this critical signaling complex as well as the evolutionary factors underlying the maintenance of gene duplicates remain unknown. Here, we investigated LLG gene family evolution, function, and expression in angiosperms. We found that LLGs in monocots and eudicots are descendants of a duplication early in angiosperm evolution and that both ancient and recent LLG duplicates are retained. Complementation and expression analysis showed that expression divergence of LLGs (regulatory subfunctionalization), rather than functional divergence, explains the retention of paralogs in Brassicales. All but one extant monocot and eudicot species examined maintained an LLG copy with preferential expression in male reproductive tissues, with the other duplicate copies showed highest levels of expression in female or vegetative tissues. Interestingly, the single LLG copy in Amborella (sister to all other angiosperms) is expressed vastly higher in male compared to female reproductive or vegetative tissues. Reconstruction of expression evolution showed that the highest inferred expression levels for the single copy ancestral angiosperm LLG was in male reproductive tissues. We propose that expression divergence played an important role in maintenance of LLG duplicates in angiosperms. One Sentence SummaryExpression divergence played an important role in maintenance of two sub-groups of LLG duplicates in angiosperms

plant biology↗

bioRxiv2019

A receptor for herbivore-associated molecular patterns mediates plant immunity

Plant-herbivore interactions are ubiquitous across nature and drive major agricultural losses. Inducible defense responses triggered through immune recognition aid in host plant protection; however, specific ligand-receptor pairs mediating the initial perception of herbivory remain unknown. Plants in the subtribe Phaseolinae detect herbivore-associated peptides in caterpillar oral secretions and the defined ligands are proteolytic fragments of chloroplastic ATP synthase termed inceptins. Using forward genetic mapping of inceptin-induced responses, we identify a cowpea (Vigna unguiculata) leucine-rich repeat receptor-like protein as an inceptin receptor (INR) sufficient for elicitor-induced responses and enhanced defense against armyworms (Spodoptera exigua). INR defines a receptor by which plants perceive herbivore-associated molecular patterns (HAMPs) and expands the paradigm of surface immune recognition to attack with mandibles.\n\nOne Sentence SummaryA plant cell surface receptor directly perceives peptides associated with caterpillar herbivory.

plant biology↗

bioRxiv2019

Chitin perception in plasmodesmata identifies subcellular, context-specific immune signalling in plants

The plasma membrane (PM) that lines plasmodesmata has a distinct protein and lipid composition, underpinning specific regulation of these connections between cells. The plasmodesmal PM can integrate extracellular signals differently from the cellular PM, but it is not known how this specificity is established or how a single stimulus can trigger independent signalling cascades in neighbouring membrane domains. Here we have used the fungal elicitor chitin to investigate signal integration and responses at the plasmodesmal PM. We found that the plasmodesmal PM employs a receptor complex composed of the LysM receptors LYM2 and LYK4 which respectively change their location and interactions in response to chitin. Downstream, signalling is transmitted via a specific phosphorylation signature of an NADPH oxidase and localised callose synthesis that causes plasmodesmata closure. This demonstrates the plasmodesmal PM deploys both plasmodesmata-specific components and differential activation of PM-common components to independently integrate an immune signal.

plant biology↗

bioRxiv2018

CrRLK1L receptor-like kinases HERK1 and ANJEA are female determinants of pollen tube reception

Communication between the gametophytes is vital for angiosperm fertilisation. Multiple CrRLK1L-type receptor kinases prevent premature pollen tube burst, while another CrRLK1L protein, FERONIA (FER), is required for pollen tube burst in the female gametophyte. We report here the identification of two additional CrRLK1L homologues, HERCULES RECEPTOR KINASE 1 (HERK1) and ANJEA (ANJ), which act redundantly to promote pollen tube burst at the synergid cells. HERK1 and ANJ localise to the filiform apparatus of the synergid cells in unfertilised ovules, and in herk1 anj mutants a majority of ovules remain unfertilised due to pollen tube overgrowth, together indicating that HERK1 and ANJ act as female determinants for fertilisation. As in fer mutants, the synergid cell-specific, endomembrane protein NORTIA (NTA) is not relocalised after pollen tube reception; however, unlike fer mutants, reactive oxygen species levels are unaffected in herk1 anj double mutants. Both ANJ and HERK1 associate with FER and its proposed co-receptor LORELEI (LRE) in planta. Together, our data indicate that HERK1 and ANJ act with FER to mediate female-male gametophyte interactions during plant fertilisation.

genetics↗

bioRxiv2018

TTL proteins scaffold brassinosteroid signaling components at the plasma membrane to optimize signal transduction in plant cells

Brassinosteroids (BRs) form a group of steroidal hormones essential for plant growth, development and stress responses. Here, we report that plant-specific TETRATRICOPEPTIDE THIOREDOXIN-LIKE (TTL) proteins are positive regulators of BR signaling functioning as scaffold for BR signaling components in Arabidopsis. TTL3 forms a complex with all core components involved in BR signaling, including the receptor kinase BRASSINOSTEROID INSENSITIVE1 (BRI1), the transcription factor BRASSINAZOLE RESISTANT1 (BZR1) and the phosphatase BRI1-SUPPRESSOR1 (BSU1), but excluding the co-receptor BAK1. TTL3 is mainly localized in the cytoplasm, but BR treatment increases its localization at the plasma membrane, where it strengthens the association with BR signaling components. Consistent with a role in BR signaling, mutations in TTL3 and related TTL1 and TTL4 genes cause reduced BR responsiveness. We propose a mechanistic model for BR signaling, in which cytoplasmic/nuclear BR components bound to TTL proteins are recruited to the plasma membrane upon BR perception, which in turn allows the assembly of a BR signaling complex, leading to the de-phosphorylation and nuclear accumulation of the transcription factors BZR1 and BES1.

molecular biology↗

bioRxiv2017

The plant calcium-dependent protein kinase CPK3 phosphorylates REM1.3 to restrict viral infection

Plants respond to pathogens through dynamic regulation of plasma membrane-bound signaling pathways. To date, how the plant plasma membrane is involved in responses to viruses is mostly unknown. Here, we show that plant cells sense the Potato virus X (PVX) COAT PROTEIN and TRIPLE GENE BLOCK 1 proteins and subsequently trigger the activation of a membrane-bound calcium-dependent kinase. We show that the Arabidopsis thaliana CALCIUM-DEPENDENT PROTEIN KINASE 3-interacts with group 1 REMORINs in vivo, phosphorylates the intrinsically disordered N-terminal domain of the Group 1 REMORIN REM1.3, and restricts PVX cell-to-cell movement. REM1.3-s phospho-status defines its plasma membrane nanodomain organization and is crucial for REM1.3-dependent restriction of PVX cell-to-cell movement by regulation of callose deposition at plasmodesmata. This study unveils plasma membrane nanodomain-associated molecular events underlying the plant immune response to viruses.

plant biology↗

bioRxiv2017

Heterologous expression of the immune receptor EFR in Medicago truncatula reduces pathogenic infection, but not rhizobial symbiosis

Interfamily transfer of plant pattern recognition receptors (PRRs) represents a promising biotechnological approach to engineer broad-spectrum, and potentially durable, disease resistance in crops. It is however unclear whether new recognition specificities to given pathogen-associated molecular patterns (PAMPs) affect the interaction of the recipient plant with beneficial microbes. To test this in a direct reductionist approach, we transferred the Brassicaceae-specific PRR ELONGATION FACTOR-THERMO UNSTABLE RECEPTOR (EFR) from Arabidopsis thaliana to the legume Medicago truncatula, conferring recognition of the bacterial EF-Tu protein. Constitutive EFR expression led to EFR accumulation and activation of immune responses upon treatment with the EF-Tu-derived elf18 peptide in leaves and roots. The interaction of M. truncatula with the bacterial symbiont Sinorhizobium meliloti is characterized by the formation of root nodules that fix atmospheric nitrogen. Although nodule numbers were slightly reduced at an early stage of the infection in EFR-Medicago when compared to control lines, nodulation was similar in all lines at later stages. Furthermore, nodule colonization by rhizobia, and nitrogen fixation were not compromised by EFR expression. Importantly, the M. truncatula lines expressing EFR were substantially more resistant to the root bacterial pathogen Ralstonia solanacearum. Our data suggest that the transfer of EFR to M. truncatula does not impede root nodule symbiosis, but has a positive impact on disease resistance against a bacterial pathogen. In addition, our results indicate that Rhizobium can either avoid PAMP recognition during the infection process, or is able to actively suppress immune signaling.\n\nSignificance StatementCrop engineering helps reducing the economic and environmental costs of plant disease. The genetic transfer of immune receptors across plant species is a promising biotechnological approach to increase disease resistance. Surface-localized pattern-recognition receptors (PRRs), which detect conserved characteristic microbial features, are functional in heterologous taxonomically-diverse plant species, and confer broad-spectrum disease resistance. It was unclear whether PRR transfer negatively impacts the association of the recipient plants with symbiotic microbes. Here, we show that a legume engineered with a novel PRR recognizing a conserved bacterial protein becomes more resistant to an important bacterial pathogen without significant impact on nitrogen-fixing symbiosis with rhizobia. This finding is of particular relevance as attempts to transfer this important symbiosis into non-legume plants are ongoing.

plant biology↗

bioRxiv2017

Pattern-Triggered Immunity And Cell Wall Integrity Maintenance Jointly Modulate Plant Stress Responses

Plant cells are surrounded by walls, which must often meet opposing functional requirements during plant growth and defense. The cells meet them by modifying wall structure and composition in a tightly controlled and adaptive manner. The modifications seem to be mediated by a dedicated cell wall integrity (CWI) maintenance mechanism. Currently the mode of action of the mechanism is not understood and it is unclear how its activity is coordinated with established plant defense signaling. We investigated responses to induced cell wall damage (CWD) impairing CWI and the underlying mechanism in Arabidopsis thaliana. Interestingly inhibitor- and enzyme-derived CWD induced similar, turgor-sensitive stress responses. Genetic analysis showed that the receptor-like kinase (RLK) FEI2 and the mechano-sensitive, plasma membrane-localized Ca2+- channel MCA1 function downstream of the THE1 RLK in CWD perception. Phenotypic clustering with 27 genotypes identified a core group of RLKs and ion channels, required for activation of CWD responses. By contrast, the responses were repressed by pattern-triggered immune (PTI) signaling components including PEPR1 and 2, the receptors for the immune signaling peptide AtPep1. Interestingly AtPep1 application repressed CWD-induced phytohormone accumulation in a PEPR1/2-dependent manner. These results suggest that PTI suppresses CWD-induced defense responses through elicitor peptide-mediated signaling during defense response activation. If PTI is impaired, the suppression of CWD-induced responses is alleviated, thus compensating for defective PTI.\n\nSignificance statementStress resistance and plant growth determine food crop yield and efficiency of bioenergy production from ligno-cellulosic biomass. Plant cell walls are essential elements of the biological processes, therefore functional integrity of the cell walls must be maintained throughout. Here we investigate the plant cell wall integrity maintenance mechanism. We characterize its mode of action, identify essential signaling components and show that the AtPep1 signaling peptide apparently coordinates pattern triggered immunity (PTI) and cell wall integrity maintenance in plants. These results suggest how PTI and cell wall modification coordinately regulate biotic stress responses with plants possibly compensating for PTI impairment through enhanced activation of stress responses regulated by the CWI maintenance mechanism.

plant biology↗