Aarhus, Denmark
November 1, 2012
View article on the website of Aarhus University
In collaboration with national and international experts, researchers from Aarhus University have revealed new fundamental features of biomolecular interactions that enable plants to identify and respond appropriately to microorganisms. The new results provide a better understanding of the mechanisms governing the ability of plants to interact with beneficial microorganisms while being resistant to pathogenic bacteria and fungi. This could have implications for future sustainable agriculture, where useful microorganisms are increasingly sought to replace pesticides.
Plant roots are surrounded by thousands of bacteria and fungi living in the soil and on the root surface. To survive in this diverse environment, plants employ sophisticated detection systems to distinguish pathogenic microorganisms from beneficial microorganisms.
Here the so-called chitin molecules from microorganisms, along with modified versions, play an important role as they are detected by the plant surveillance system. Legumes, for example, build a defence against pathogenic microorganisms in response to simple chitin molecules.
However, when the plant detects a specific modified chitin molecule (called a Nod factor) that is secreted from the rhizobium soil bacteria, formation of new organs in the form of "root nodules" occurs. Rhizobium bacteria are allowed to enter and colonise in these symbiotic organs, and they ultimately produce nitrogen for the plant.
The plants’ detection of ligands – such as chitin and modified Nod factors – takes place through protein receptors that are localised on the surface of cells. Research at the Centre for Carbohydrate Recognition and Signalling (CARB) has now shown that ligand recognition through direct Nod factor binding is a key step in the receptor-mediated signal transduction that leads to root nodule development in legumes.
High-affinity binding was observed in the nano-molar range, comparable to the biologically relevant concentrations where Nod factor has in vivo activity. In contrast to this, simple chitin molecules bind to the receptors with low affinity. Structure-dependent ligand specificity and ligands binding affinities at different receptors may therefore determine which response mechanism is activated in plants exposed to different microbes or microbial communities in the environment.
Interdisciplinary approaches combining advanced biochemistry, chemoselective chemistry and microbial genetics made it possible to investigate the molecular mechanisms involved in distinguishing between Nod factor molecules secreted from rhizobia and chitin secreted by pathogenic microorganisms.
The challenging task of purifying plant receptor proteins, which are present in very low amounts in roots of the model legume Lotus japonicus, was successfully accomplished by expressing the receptors in heterologeous plant-based systems and purifying them from membrane fractions.
Another challenge was the establishment of binding assays with the carbohydrate ligands. Nod factor labelling and Nod factor immobilisation facilitated this, following application of chemoselective chemistry.
The researchers behind the results that have just been published in the international journal PNAS are affiliated with the Danish National Research Foundation’s Centre for Carbohydrate Recognition and Signalling at the Department of Molecular Biology and Genetics, Aarhus University (Denmark), Department of Chemistry, University of Copenhagen (Denmark) and Department of Microbiology and Immunology, University of Otago (New Zealand).
Link to the scientific article in PNAS:
Legume receptors perceive the rhizobial lipochitin oligosaccharide signal molecules by direct binding
Angelique Broghammera, Lene Krusella, Mickaël Blaisea, Jørgen Sauera, John T. Sullivana, Nicolai Maolanona,
Maria Vinthera, Andrea Lorentzene, Esben B. Madsena, Knud J. Jensena, Peter Roepstorffe, Søren Thirupa, Clive W. Ronsona, Mikkel B. Thygesena, and Jens Stougaarda.
Photo:
First author: Angelique Broghammer (Aarhus University, Denmark) (photo: Lisbeth Heilesen).
Planter genkender sygdomsfremkaldende og nyttige mikroorganismer
Forskere fra Aarhus Universitet har i samarbejde med nationale og internationale eksperter afsløret nye grundlæggende træk i de biomolekylære interaktioner, der sætter planter i stand til at identificere og reagere hensigtsmæssigt på mikroorganismer. De nye resultater giver en bedre forståelse af de mekanismer, der styrer planters evne til at interagere med nyttige mikroorganismer og samtidig være modstandsdygtige over for sygdomsfremkaldende bakterier og svampe. Dette vil kunne få betydning for fremtidens bæredygtige landbrug, hvor sprøjtemidler i stigende grad søges erstattet med nyttige mikroorganismer.
Planters rødder er omgivet af tusindvis af bakterier og svampe, der lever i jorden og på røddernes overflade. For at overleve i dette mangfoldige miljø anvender planterne avancerede detektionssystemer for at skelne sygdomsfremkaldende mikroorganismer fra nyttige såvel som harmløse mikroorganismer.
Her spiller såkaldte kitin-molekyler og modificerede kitin-molekyler, der stammer fra mikroorganismerne, en vigtig rolle, da de registreres af plantens overvågningssystem. For eksempel opbygger bælgplanter et forsvar mod sygdomsfremkaldende mikroorganismer som en reaktion på simple kitin-molekyler.
Når planten derimod detekterer et modificeret kitin-molekyle udskilt fra rhizobiumjordbakterier, såkaldte Nod-faktorer, fører dette til dannelse af nye organer i form af ”rodknolde”, som tillader rhizobium-bakterierne at invadere og etablere sig i disse symbiotiske organer. Rhizobium-bakterierne producerer derefter kvælstof til planten.
Planters detektion af ligander, såsom kitin og modificerede Nod-faktorer, sker gennem proteinreceptorer, der er lokaliseret i cellernes overflade. Forskningen ved Danmark Grundforskningsfonds Center for Kulhydratgenkendelse og Signallering (CARB) har nu vist, at ligand-genkendelse ved direkte binding af Nod-faktor molekyler er et vigtigt skridt i den process, der fører til udvikling af rodknolde i bælgplanter.
Receptor-bindingen sker ved nano-molære koncentrationer af Nod-faktor, hvilket er sammenligneligt med de biologisk relevante koncentrationer, hvor Nod-faktor har in vivo aktivitet. Simple kitin-ligander binder derimod kun til recepetorerne med lav affinitet ved mikromolære koncentrationer. Receptor-genkendelsen er således bestemt af ligandens opbygning og struktur, og ligand-bindings-affiniteten på forskellige receptorer kan derfor bestemme, hvilken responsmekanisme der aktiveres i planten.
Ved at inddrage flere forskellige tværfaglige discipliner, såsom avanceret biokemi, kemoselektiv kemi og mikrobiel genetik, var det muligt for forskerne at undersøge de molekylære mekanismer, der er involveret i at skelne mellem Nod-faktor-molekyler, der udskilles fra rhizobium, og kitin der udskilles fra sygdomsfremkaldende mikroorganismer.
En af udfordringerne var at oprense receptorproteiner, som er til stede i meget små mængder i rødder af bælgplanten japansk kællingetand (Lotus japonicus), der blev brugt som modelplante. Denne udfordring blev løst ved at udtrykke receptorerne i forskellige plante-baserede systemer og oprense dem fra membranfraktioner.
En anden udfordring var etableringen af bindingsassays med kulhydrat-ligander. Dette blev løst ved kemisk modifikation af Nod-faktor og kitin ved hjælp af kemoselektiv kulhydrat kemi.
Forskerne bag de resultater, som netop er blevet offentliggjort i det internationale tidsskrift PNAS, er tilknyttet Danmark Grundforskningsfonds Center for Kulhydratgenkendelse og -Signalering på Institut for Molekylær Biologi og Genetik ved Aarhus Universitet, Kemisk Institut ved Københavns Universitet og Institut for Mikrobiologi og Immunologi ved Otago Universitet (New Zealand).
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