Meccanismi di percezione di oligosaccaridi alla base della difesa delle piante da agenti patogeni

Responsabili di progetto

Rita Berisio, Hanae Kaku

Accordo

GIAPPONE - JSPS - Japan Society for the Promotion of Science

Bando

CNR-JSPS 2016-2017

Dipartimento

Scienze chimiche e tecnologie dei materiali

Area tematica

Scienze chimiche e tecnologie dei materiali

Stato del progetto

Nuovo

Proposta di ricerca

Plants are under constant attack from microbes. They defend themselves against attacks by microbes using a variety of sophisticated defense mechanisms. The first line of defense in plants is the perception of conserved molecules characteristic of many microbes denoted as microbe-associated molecular patterns (MAMPs) by pattern recognition receptors (PRRs) localized on the surface of plant cells. The MAMP-triggered immunity plays a primary role in the establishments of resistance to microbes (1). It has been shown that MAMP-induced defense responses include alterations in the plant cell wall, induction of antimicrobial compounds and the production of reactive oxygen species and reactive nitrogen species (2). Plants can detect infecting fungi through the perception of chitin oligosaccharides by lysin motif of proteins such as CEBiP and CERK1 (3,4). CEBiP (chitin elicitor-binding protein) is a plasma membrane glycoprotein, identified by the Meiji Unit, which plays a key role in the perception of chitin elicitor in rice, since its knock-down transformants exhibit suppression of chitin-induced defense responses (4). CEBiP lacks any obvious intracellular domains for signal transduction, indicating it may require additional factors for signaling (4). Consistently, it has been identified a novel key component for chitin elicitor signaling, denoted as OsCERK1 (Chitin Elicitor Receptor Kinase). Different than CEBiP, this other protein contains an intracellular Ser/Thr kinase domain. As a common feature, these two proteins contain extra-cellular LysM domains, which are likely to bind chitin oligosaccharides (3-4). However, how CEBiP and OsCERK1 molecules communicate to signal oligosaccharide perception through the plasma membrane into the cytoplasm is hitherto unknown.

This project aims at elucidating plant-pathogen interactions at a molecular level in order to shed light on mechanisms of plant defense reactions. With this aim, we will investigate mechanisms of chitin oligosaccharide perception by the two key receptor molecules CEBiP and OsCERK1 by coupling a variety of structural biology approaches with plant biology.
Structural aspects of host-pathogen interactions will be investigated by analyzing interactions between chitin oligosaccharides and its receptors. Also, results will elucidate the mechanism of OsCERK1 activation upon chitin binding by CEBiP. This project will necessarily require a tight interaction among molecular biologists, structural biologists and plant biologists. All these competences are covered by the Units of this project.Recent results of an ongoing collaboration between the Units provide the basis for the planned activities. These activities will be developed using different and complementary approaches, both theoretical and experimental.

Altogether, adopted methodologies will lead to an improved understanding of the chemical interactions between MAMPs and their plant receptors and therefore provide molecular basis of MAMP elicitor perception. A better understanding of the mechanisms by which pathogen determinants elicit defense responses will have a considerable impact on the improvement of plant health and disease resistance with consequent benefit to society.

These studies will suggest the identification of candidate agrochemicals that will enhance the resistance of plants to pathogens or enhance plant growth under abiotic stress conditions (nutrient stress, water deficit). This will be exploited as a possible way to protect crops from fungal diseases in agriculture. On the other hand, they have the potential to increase the crop production while reducing the use of current agrochemicals with the aim to diminish environmental pollution (diminish antibiotic resistance). This project will aim for maximally productive outcomes by a widespread engagement through publicity of the project in the scientific community at meetings, in publications and through existing communication networks (www.costbm1003.info). This project plans extensive exchange of researchers to expand the scientific expertise of both partners. The project will also translate in joint Patents and joint high impact publications and positively impact S&T bilateral relationships.

The fruitful collaboration between the two partners (CNR-IBB and Meiji) has already translated in a joint project, coordinated by Dr Berisio and funded by the Mizutani Foundation for Glycoscience (http://mizutanifdn.or.jp/grant/grantees/grantinfo.html) and one paper in PNAS (5).
Preliminary results obtained by the collaborative work of the two groups have proven the feasibility of the work planned in this project. The structural characterization of plant proteins directly involved in the innate immunity reaction will elucidate specific chitin oligosaccharide binding modes and will be accomplished using an array of molecular biology and structural biology approaches, routinely adopted at CNR-IBB. By combining this molecular view of the elicitation process with plant biology, we will provide molecular basis of MAMP elicitor perception.

References:
1. Jones J.D., Dangl J.L., "The plant immune system", Nature. 2006 Nov 16;444(7117):323-9.
2. Newman M.A., Sundelin T., Nielsen J.T., Erbs G. "MAMP (microbe-associated molecular pattern) triggered immunity in plants" Front Plant Sci. 2013 May 16;4:139.
3. Shimizu T. et al., "Two LysM receptor molecules, CEBiP and OsCERK1, cooperatively regulate chitin elicitor signaling in rice" Plant J. 2010 Oct; 64(2): 204-214.
4. Shinya T., Nakagawa T., Kaku H., Shibuya N. "Chitin-mediated plant-fungal interactions: catching, hiding and handshaking" Curr Opin Plant Biol. 2015 Aug;26:64-71.
5. Hayafune M., Berisio R., Marchetti R., Silipo A., Kayama M., Desaki Y., Arima S., Squeglia F., Ruggiero A., Tokuyasu K., Molinaro A., Kaku H. and Shibuya N. (2014) Chitin-induced activation of immune signaling by the rice receptor CEBiP relies on a unique sandwich-type dimerization, PNAS 111, E404-413.

Obiettivi della ricerca

Despite the importance of chitin-mediated immunity in plants, molecular machineries involved in the perception of chitin and their signal transduction have been poorly understood. The overall aim of this proposal is to investigate mechanisms of plant defense reactions at a molecular level by coupling a variety of structural biology approaches with plant biology.
The interaction between experts in plant biology experts (Meiji University), who have identified the two key receptor molecules CEBiP and OsCERK1, and structural biology experts (CNR-IBB) is a strong need for the understanding of the mechanisms of chitin oligosaccharide perception and signaling.

Activities will be developed using different structural biology approaches, both theoretical and experimental, complemented with phenotype analysis, in terms of OsCERK1 expression and elicitor response. As an ongoing collaboration (funded by the Mizutani Foundation of Glycoscience), researchers of the Italian and Japanese groups have already proven the feasibility of the planned activities.

The objectives of this project are:
- To obtain highly pure CEBiP and OsCERK1 receptors
- To crystallize receptors and complexes with MAMPs
- To solve three-dimensional structures of receptors and complexes
- To investigate interaction of chitin oligosaccharides with receptor mutants with reduced activity
- To investigate interactions between CEBIP and OsCERK1

Ultimo aggiornamento: 08/06/2025