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THE APOPLAST AND THE INTEGRATION OF PROCESSES REGULATING DEVELOPMENT AND INNATE IMMUNITY OF PLANTS

Università degli Studi Roma Tre

Abstract

ABSTRACT
The main goal of our research project is to investigate the apoplast-specific events that orchestrate processes related to growth and development and the mechanisms that allow a plant to resist to pathogen infection. Recent findings indicate that the apoplastic perception systems associated to both hormonal or pathogen-derived stimuli are often associated and orchestrate coordinated signal trasduction pathways. In the absence of an adaptive immune system, plants base their defence strategies on an innate immune system, analogous to the one found in animals, that includes kinase receptors with leucine-rich repeat (LRR) domains that are implicated in the recognition of “elicitors”, i.e. pathogen-derived molecules. These include the so-called pathogen-associated molecular patterns (PAMPs) such as fungal chitin and bacterial flagellin. The oligogalacturonides (OGs) are instead host-associated molecular patterns (HAMPs). They are released from the plant cell wall homogalacturonan by the action of pathogen-derived polygalacturonases whose enzymatic activity is modulated by specific apoplastic polygalacturonase inhibiting proteins (PGIPs). The OGs regulate some important developmental processes and share with a number of PAMPs common signal transduction pathways for the activation of defence responses. The LRR-RLK mediated immunity is a multi-component defence strategy that is associated to drastic changes in the apoplast redox state. In particular the production of >>>

Principal Investigator

Riccardo Angelini Università degli Studi ROMA TRE

Research Objectives

The main goal of this integrated research project is to investigate the apoplast-specific events that orchestrate processes related to growth and development and the mechanisms that allow a plant to resist to pathogen infection. The evidence that the brassinosteroid co-receptor BAK1 is also necessary for the function of innate immunity receptors, implicated in the recognition at the apoplastic level of pathogen-associated molecular patterns (PAMP), and that this interaction is at the origin of an integrated regulation of development and defence responses, sheds a new light on these wide research areas. The dualism defence/development applies also to plant-derived elicitors (HAMP: Host-Associated Molecular Patterns) such as the oligogalacturonides (OG). The OG regulate some important developmental processes and share with a number of PAMPs the same signal transduction pathways for the activation of defence responses. They are released from the homogalacturonan of the plant cell wall by pathogen-derived polygalacturonases (PG) whose activity is modulated by specific plant inhibitors, localized at the apoplast [PG Inhibiting Proteins (PGIP)]. The production of reactive oxygen species (ROS) is also a response that is typically associated both to the activation of defence mechanisms (PAMP and HAMP induce an oxidative burst) and to the differentiation processes associated to development, which are also intimately related with secretion processes and endocytosis. The structural and >>>

First Results

EXPECTED RESULTS

OBJECTIVE A. STRUCTURAL CHARACTERIZATION OF THE PG-PGIP INTERACTION AND OF PROTEINS THAT MEDIATE PAMP AND HAMP RECOGNITION. STUDIES ON THE OG-MEDIATED SIGNAL TRASDUCTION PATHWAY

A. 1.IDENTIFICATION AND CHARACTERIZATION OF PGIP “HOT SPOTS” FOR PG RECOGNITION
-We expect to identify the minimal PGIP structural requirements for the recognition of each different PGs. These data will be used to rationally design PGIP2 variants able to recognize fungal PGs that wild-type PGIP2 does not inhibit.

A. 2.CRYSTALLIZATION OF THE PG-PGIP COMPLEX, A CHIMERIC APPROACH
-The minimal expectation is to obtain diffracting crystals of the PG-PGIP chimera and, possibly, to determine its structure within the project time span.

A. 3. STRUCTURAL ANALYSIS OF THE FLS2-FLG22 INTERACTION
-We expect to have obtained by the end of the program a system for the heterologous expression of FLS2-LRR, to have developed a purification protocol to obtain the protein in milligram quantities and to have performed a complete crystallization screen of the protein alone and in complex with flg22. Crystals obtained will be subjected to the crystallographic analysis.
- Availability of constructs for in planta expression of two FLS2/EFR chimeric receptors, also fused with GFP, and of transgenic Arabidopsis ecotype Wassiliewskija (Ws) plants, which normally do not respond to fgl22, expressing these receptors, as well as >>>

Timescale

24 months

National and international background

SCIENTIFIC BACKGROUND
The plant cell apoplast, mainly represented by the cell wall and the wall/plasma membrane interface, is the site where crucial events of perception and signalling occur, involving both endogenous stimuli that regulate differentiation and development, and exogenous signals associated to biotic and abiotic stresses. Recent evidence indicates that the same perception/transduction elements may mediate the response to these two classes of stimuli and orchestrate the activation of signalling pathways for a coordinated regulation of developmental and defence processes.

PLANT INNATE IMMUNITY
In the absence of an adaptive immune system, plants rely for defence against pathogens on a so-called innate immunity system analogous to that found in animals (36). Main elements of this system are germ line-encoded pattern receptors (PRR) for recognition of pathogen-derived molecules indicated as elicitors (generic elicitors or plant-genotype specific elicitors). Generic elicitors are indicated as pathogen-associated molecular patterns (PAMP) and include fungal chitin and bacterial flagellin. Oligogalacturonides (OG) are plant endogenous elicitors (HAMP -Host-Associated Molecular Patterns) that are released from the plant cell wall homogalacturonan (HGA) by pathogen-secreted polygalacturonase (PG), which are also ranked among PAMP (42). OG accumulation is favored by the interaction with specific inhibitors of PGs (PGIP >>>