Immune signaling in arabidopsis thaliana upon perception of bacterial and viral molecular patterns with a special emphasis on roots

In order to recognize a vast variety of attackers, plants possess a plethora of sophisticated detection systems. Perception of microbe- or pathogen-associated molecular patterns (MAMPs or PAMPs) by the plant pattern recognition receptors (PRRs) leads to subsequent initiation of defense responses, a process collectively referred to as pattern-triggered immunity (PTI). PTI has been extensively studied in plant leaves, especially of the model organism Arabidopsis thaliana, whereas the mechanisms underlying PTI in roots so far attracted less attention. However, since a vast number of plant pests are soil-borne and attack roots in order to propagate and colonize whole plants, understanding the mechanisms underlying basic defense at the root level is of high interest for the development of new tools to combat root pathogens of crop plants. It has been demonstrated that recognition of flg22, the conserved epitope of the bacterial flagellin protein, leads to tissue-specific defense responses in roots. In order to investigate the cause for this tissue-specific induction of downstream responses, several approaches were employed during the course of this work. By studying the cellular localization of the PRR recognizing flg22, FLAGELLIN-SENSING 2 (FLS2), we were able to depict an expression map of FLS2 in wild-type Arabidopsis plants. Our study revealed that FLS2 was expressed in a highly tissue-specific manner in roots and shoots and that the FLS2 promoter activity was inducible upon environmental stimuli as well as during developmental processes, changing not only in intensity in expressing tissues but also in tissue-specificity. These results indicate an important role of the tissue-specific PRR localization in immunity mechanisms. In a parallel study, we expressed FLS2 under the control of several root tissue-specific promoters, which allowed us to analyze the competence of these tissues to detect flg22. Unexpectedly, all investigated root tissues were able to perceive externally applied flg22. In fact, PTI responses could be activated in intact roots as well as in dissected roots, suggesting that the peptide is able to penetrate through the different tissue layers. Remarkably, the expression level of the receptor was not the major parameter determining the magnitude of the immune response output. Thus, we postulated that perception of flg22 by certain tissues leads to stronger PTI responses potentially indicating why plants restrict immune receptor accumulation to tissue-specific locations possibly in order to balance the outcome of the defense activation. Due to the fact that many developmental or immunity processes in plants depend on systemic communication between different plant organs and that beneficial root microbes are known to prime and enhance resistance in aerial plant tissues, we hypothesized that MAMP perception by roots might induce a signaling event from roots to shoots. In order to address the potential existence of such systemic alarm signals, various methods were implemented. However, we encountered several technical limitations mainly concerning elicitor diffusion. Therefore, we focused on the development of an improved application method for studying systemic root-to-shoot signaling in Arabidopsis plants. Our system proved suitable to perform systemic signaling analysis and revealed that at the transcriptional level no systemically activated defense gene modifications were detectable in distal shoots of root-treated plants in our conditions. Like root pathogens, also viruses constitute a major threat in agro-economy and are responsible for immense crop losses. The basal defense response against viruses is thought to be mediated by RNA silencing, a process by which viral replication intermediates are cleaved and degraded by the plant silencing machinery through the recognition of virus-derived small RNAs. Intriguingly, a recent study conducted in our lab demonstrated a role of BRASSINOSTEROID INSENSITIVE1 (BRI1)-ASSOCIATED RECEPTOR KINASE 1 (BAK1), a coreceptor of several PRRs involved in immunity and development, in antiviral defense. These results indicated that PTI may also contribute to antiviral resistance but the exact recognition process remained elusive. Because dsRNA produced during viral replication has been shown to act as a PAMP in animals, we decided to test whether dsRNA is perceived as a viral PAMP in planta as well. We found that natural as well as synthetic dsRNA is indeed perceived as a PAMP by Arabidopsis, leading to the activation of typical PTI responses. Remarkably, dsRNA application also promoted protection of Arabidopsis plants against viral infection. Taken together, this study provides new insights into the recognition mechanisms of bacteria- and virus-associated molecular patterns by different plant organs and contributes to elucidate the molecular defense strategy of plants against agriculturally important diseases

Expression patterns of FLAGELLIN SENSING 2 map to bacterial entry sites in plant shoots and roots

Expression of the flagellin receptor FLS2 is regulated in a cell/tissue-specific and stress-induced manner that correlated with sites of bacterial infection. The vasculature expresses FLS2 and responds to flagelli

Double-stranded RNAs induce a pattern-triggered immune signaling pathway in plants

Pattern-triggered immunity (PTI) is a plant defense response that relies on the perception of conserved microbe- or pathogen-associated molecular patterns (MAMPs or PAMPs, respectively). Recently, it has been recognized that PTI restricts virus infection in plants; however, the nature of the viral or infection-induced PTI elicitors and the underlying signaling pathways are still unknown. As double-stranded RNAs (dsRNAs) are conserved molecular patterns associated with virus replication, we applied dsRNAs or synthetic dsRNA analogs to Arabidopsis thaliana and investigated PTI responses. We show that in vitro-generated dsRNAs, dsRNAs purified from virus-infected plants and the dsRNA analog polyinosinic-polycytidylic acid (poly(I:C)) induce typical PTI responses dependent on the co-receptor SOMATIC EMBRYOGENESIS RECEPTOR-LIKE KINASE 1 (SERK1), but independent of dicer-like (DCL) proteins in Arabidopsis. Moreover, dsRNA treatment of Arabidopsis induces SERK1-dependent antiviral resistance. Screening of Arabidopsis wild accessions demonstrates natural variability in dsRNA sensitivity. Our findings suggest that dsRNAs represent genuine PAMPs in plants, which induce a signaling cascade involving SERK1 and a specific dsRNA receptor. The dependence of dsRNA-mediated PTI on SERK1, but not on DCLs, implies that dsRNA-mediated PTI involves membrane-associated processes and operates independently of RNA silencing. dsRNA sensitivity may represent a useful trait to increase antiviral resistance in cultivated plants

Contemporary Vietnam: Political Opportunities, Conservative Formal Politics, and Patterns of Radical Change

This article examines contemporary Vietnamese politics and argues that many opportunities remain for political rethinking. It examines linguistic arguments that point to a perpetuation of traditional Communist ideas in two crucial areas—village elections and the treatment of “policy.” It then juxtaposes this formal conservatism with two areas of tension. First, problems exist in addressing important policy questions related to rural development, poverty, and participation. These suggest that without major rethinking of fundamental political issues, such problems are increasingly hard to address. Second, it examines the context of formal politics, looking at evidence for successful contestation over leadership positions in villages and the rise of informal farmers’ groups. Both have often led local officials to simple toleration and accommodation rather than exploitation of such trends to assist their own repositioning in ways that could gain them popular political support.Again, such trends highlight the void in formal political rethinking. Both of these areas of tension create considerable difficulties for donors and external partners, whose approaches are often premised upon both a familiar policy-driven role for the state in development and ignorance of local political processes

Responses of PAMP-triggered immunity in roots.

<p>(a) Luminol-based detection of ROS production in isolated roots of Col-0 seedlings treated with 1 μM flg22, chi7 or AtPep1 or without elicitor. Data represent the mean of 12 replicates ± SE. RLU—relative luminescence units (b) Activation of MAP kinases detected by western blot (MPK). Isolated roots from 2-week old seedlings of Col-0 and transgenic lines were treated with 1 μM flg22, chi7 or AtPep1 or without elicitor for 10 min before sampling. Ponceau S staining was used as loading control (PS). The experiment was performed three times with similar results.</p

Root growth inhibition by AtPep1.

<p>(a) Col-0 wild-type and transgenic lines were germinated and grown vertically for 12 days on 0.5x MS containing 10 nM, 100 nM or 1 μM flg22, chi7 or AtPep1 or without elicitor. (b) Analysis of primary root length was performed using Fiji on images recorded under (a) and is shown as box plots representing ≥ 20 roots per sample. The experiment was performed three times for Col-0 and two times for <i>pepr1 pepr2</i>. Statistical analysis was performed using a Student’s t-test comparing with roots grown on control medium: * p < 0.05, ** p < 0.01, *** p < 0.001.</p

Elicitor-triggered responses of <i>pHEL</i>::<i>YFP</i>_<i>N</i>.

<p>(a) Microscopic analysis of root developmental zones of 7-days old <i>pHEL</i>::<i>YFP</i>_<i>N</i> seedlings following 24 h treatment with 100 nM flg22, chi7, AtPep1 or 0.5x MS as control. RC—root cap / meristem, TZ—transition zone, DZ—differentiation zone, MZ—mature zone. Bar 100 μM. (b) Quantification of fluorescent signals from microscopic analysis of different developmental zones of <i>pHEL</i>::<i>YFP</i>_<i>N</i> as described under (a) using Fiji. Bars represent the mean of ≥ 3 images ± SD.</p

Elicitor-dependent activation of defence-related promoters in the TZ and DZ of the root.

<p>(a) Quantification of microscopic analysis of <i>Promoter</i>::<i>YFP</i>_<i>N</i> constructs in the differentiation zone of 7-days old seedlings following treatment with 100 nM flg22, chi7, AtPep1 or 0.5x MS as control. Images were analysed using Fiji. Bars represent the mean of ≥ 3 images ± SD. (b) Overview of inductive and suppressive responses of <i>Promoter</i>::<i>YFP</i>_<i>N</i> constructs in different developmental root zones analysed as described under (a). Numbers in the upper row indicate the sum of significant marker responses in one of the four analysed root zones for a given elicitor, while numbers in the lower row indicate the total sum of significant responses from all markers and all root zones for a given elicitor. Black colour indicates induction, white colour suppression and grey colour non-significant changes.</p