Plant diseases are common and can cause widespread damage to crops, making them a serious threat to stable global food supply. These diseases are spread by pathogens such as bacteria and fungi, who then secrete proteins that cause the damage to the host plants. Understanding the way that these proteins interact with the plant cells is key to developing ways to protect the plants from disease.
In this study, published in Science Advances, an international group of researchers focus on proteins known as NLPs (Necrosis and ethylene-inducing peptide 1 (Nep1)–like proteins), which affect a wide range of widely used crops such as potato, soybean and tobacco. They used a range of biophysical approaches to study how these NLPs interact with the plant plasma membrane, the part of a plant cell that forms a selective barrier between the inside of the cell and the external environment.
In particular, they looked at how NLPPya, from a common water mould, interacted with a model membrane made from tobacco leaf glycosylinositol phosphorylceramides (GIPCs). GIPCs have been identified through other studies as the part of the membrane that the NLPs bind to.
Using computational techniques, the researchers modelled how the NLP and GIPCs interact with one another, and then used neutron reflectometry on the Inter beamline at ISIS to see if their simulations were correct. As well as studying interaction of hydrogenated NLP with GIPC-containing membranes, the researchers also used a deuterated NLP version, so they could see it clearly within the hydrogen-rich membrane.
Their results showed that the NLP molecules form a layer on the outside of the membrane, rather than moving inside it. The GIPC-attached protein molecules may then interact with the surrounding GIPCs in the membrane, causing them to reorganise and cluster. This movement causes pores to form within the membrane, from which small molecules from the plant plasma leak out of, harming the plant.
Membrane damage by formation of pores is a common way for bacteria to breach integrity of animal cells but has been rarely seen in plants. The pathogens must have adapted in a way that means they can produce proteins with a structure that interacts with the specific components present in the membrane, releasing small molecules from the plant cytoplasm that boosts their nutrition.
By understanding the way these pathogens break down the plant membranes, the researchers hope to open up new ways to protect plants and maintain the security of global food production for the future.
"At the ISIS neutron and muon source we have been seeing an increasing number of visiting scientists examine membrane biochemistry using neutron reflectometry," explains Luke Clifton, from the ISIS reflectometry group. "The work of Pirc et al on the interactions of Oomycote toxins with accurate models of plant membranes is a very nice example of how this technique can provide unique structural information which helps us gain a precision understanding how toxins function."
Professor Gregor Anderluh of National Institute of Chemistry, Slovenia, and lead author of the study said: "Nep1-like proteins are widely distributed in microbial pathogens that attack plants and represent an important target for designing strategies aimed at preventing microbe-induced damage to plant cells. Understanding how these proteins damage cell membranes is a critical prerequisite for further development."
The full paper can be found at DOI: 10.1126/sciadv.abj9406