Functional characterization of cytolytic and non-cytolytic members within the NLP effector superfamily

NEP 1-like proteins (NLPs) represent a superfamiJy of effector proteins which are distributed amongst various oomycetes, fungi and bacteria. Among the best characterized NLPs are those that are expressed and secreted by the pathogens during the late stage of infection and that trigger necrosis in dicotyledooous plants. We could demonstrate that this necrotic activity makes them virulence factors in the host organisms by supporting a necrotrophic life style. The three-dimensional structure of an NLP from Pythium aphanidermatum (NLPPya) is similar to the fold of actinoporins, pore-fonniog toxins from marine invertebrates. Like actinoporins, NLPs affect the integrity of plasma membranes from dicot plants, thereby leading to necrosis. However, the detailed molecular mechanism and the target molecule of NLPs at the plasma membrane arc currently unknown. Several lines of evidence suggest a lipid-like interaction partner at the plasma membrane. A surface-exposed cavity was determined in the NLP(Pya) crystal structure that would be suitable as the binding site for such a molecule. NLP versions mutated in the vicinity of the cavity are used in a photo-affinity Labeling approach for capturing the target molecule. Recent progress in characterizing the mechanism of pore formation and the host docking site ofNLPs will be presented. Besides cytolytic NLPs, another group of NLPs exists which is expressed in the early infection stage. As expected due to the lack of conserved residues required for cytolysis, these NLPs are incapable of-triggering necrosis. They are most abundant in oomycetes and fungi, and are found in biotrophic organisms or pathogens with monocot hosts. Considering the structural relation ofNLPPya not only to actinoporins but also to :fungal lectins, non-cytolytic NLPs could contribute to host cell adhesion. We will provide recent progress in the functional analysis of noncytolYtic l\iLPs and discuss their putative role in the early infection process