Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Random mutagenesis of the nucleotide-binding domain of NRC1 (NB-LRR Required for Hypersensitive Response-Associated Cell Death-1), a downstream signalling nucleotide-binding, leucine-rich repeat (NB-LRR) protein, identifies gain-of-function mutations in the nucleotide-binding pocket
    Sueldo, D.J. ; Shimels, M.Z. ; Spiridon, L.N. ; Caldararu, O. ; Petrescu, A.J. ; Joosten, M.H.A.J. ; Tameling, W.I.L. - \ 2015
    New Phytologist 208 (2015)1. - ISSN 0028-646X - p. 210 - 223.
    disease resistance protein - secondary structure prediction - multiple sequence alignment - tobacco-mosaic-virus - immune receptors - arc domain - caenorhabditis-elegans - artificial evolution - gene-products - coiled-coil
    •Plant nucleotide-binding, leucine-rich repeat (NB-LRR) proteins confer immunity to pathogens possessing the corresponding avirulence proteins. Activation of NB-LRR proteins is often associated with induction of the hypersensitive response (HR), a form of programmed cell death. •NRC1 (NB-LRR Required for HR-Associated Cell Death-1) is a tomato (Solanum lycopersicum) NB-LRR protein that participates in the signalling cascade leading to resistance to the pathogens Cladosporium fulvum and Verticillium dahliae. •To identify mutations in NRC1 that cause increased signalling activity, we generated a random library of NRC1 variants mutated in their nucleotide-binding domain and screened them for the ability to induce an elicitor-independent HR in Nicotiana tabacum. Screening of 1920 clones retrieved 11 gain-of-function mutants, with 10 of them caused by a single amino acid substitution. •All substitutions are located in or very close to highly conserved motifs within the nucleotide-binding domain, suggesting modulation of the signalling activity of NRC1. Three-dimensional modelling of the nucleotide-binding domain of NRC1 revealed that the targeted residues are centred around the bound nucleotide. Our mutational approach has generated a wide set of novel gain-of-function mutations in NRC1 and provides insight into how the activity of this NB-LRR is regulated.
    Structural Determinants at the Interface of the ARC2 and LRR Domains Control the Activation of the NB-LRR Plant Immune Receptors Rx1 and Gpa2
    Slootweg, E.J. ; Spiridon, L.N. ; Roosien, J. ; Butterbach, P.B.E. ; Pomp, H. ; Westerhof, L.B. ; Wilbers, R.H.P. ; Bakker, E.H. ; Bakker, J. ; Petrescu, A.J. ; Smant, G. ; Goverse, A. - \ 2013
    Plant Physiology 162 (2013)3. - ISSN 0032-0889 - p. 1510 - 1528.
    secondary structure prediction - disease resistance genes - protein-protein interactions - programmed cell-death - nbs-lrr protein - coiled-coil - pathogen interactions - physical association - multiple alignments - self-association
    Many plant and animal immune receptors have a modular NB-LRR architecture in which a nucleotide-binding switch domain (NB-ARC) is tethered to a leucine-rich repeat sensor domain (LRR). The cooperation between the switch and sensor domains, which regulates the activation of these proteins, is poorly understood. Here we report structural determinants governing the interaction between the NB-ARC and LRR in the highly homologous plant immune receptors Gpa2 and Rx1, which recognize the potato cyst nematode Globodera pallida and Potato Virus X, respectively. Systematic shuffling of polymorphic sites between Gpa2 and Rx1 showed that a minimal region in the ARC2 and the N-terminal repeats of the LRR domain coordinate the activation state of the protein. We identified two closely spaced amino acid residues in this region of the ARC2 (position 401 and 403) that distinguish between autoactivation and effector-triggered activation. Furthermore, a highly acidic loop region in the ARC2 domain and basic patches in the N-terminal end of the LRR domain were demonstrated to be required for the physical interaction between the ARC2 and LRR. The NB-ARC and LRR domains dissociate upon effector-dependent activation and the complementary charged regions are predicted to mediate a fast re-association enabling multiple rounds of activation. Finally, we present a mechanistic model showing how the ARC2, NB and N-terminal half of the LRR form a clamp, which regulates the dissociation and re-association of the switch and sensor domains in NB-LRR proteins.
    Dual regulatory roles of the extended N-terminus for activation of the tomato Mi-1.2 resistance protein
    Lukasik-Shreepaathy, E. ; Slootweg, E.J. ; Richter, H. ; Cornelissen, B.J.C. ; Goverse, A. ; Takken, F.L.W. - \ 2012
    Molecular Plant-Microbe Interactions 25 (2012)8. - ISSN 0894-0282 - p. 1045 - 1057.
    secondary structure prediction - rich repeat domain - cell-death - nucleotide-binding - disease resistance - confers resistance - immune-system - coiled coils - lrr protein - arc domain
    Plant resistance (R) proteins mediate race-specific immunity and initiate host defenses that are often accompanied by a localized cell-death response. Most R proteins belong to the NB-LRR protein family as they carry a central NB-ARC domain fused to an LRR domain. The CC domain at the N-terminus of some Solanaceous NB-LRR proteins is extended with a solanaceae domain (SD). Tomato Mi-1.2, which confers resistance against nematodes, white flies, psyllids and aphids, encodes a typical SD-CNL protein. Here, we analyzed the role of the extended N-terminus for Mi-1.2 activation. Removal of the first part of the N-terminus (Nt1) induced Mi-1.2-mediated cell death that could be suppressed by over-expression of the second half of the N-terminal region (Nt2). Yet, autoactivating NB-ARC-LRR mutants require in trans co-expression of the N-terminal region to induce cell death, indicating that the N-terminus functions both as a negative and a positive regulator. Based on secondary structure predictions we could link both functions to three distinct subdomains; a typical CC domain and two novel, structurally-conserved helical subdomains called SD1 and SD2. A negative regulatory function could be assigned to the SD1 whereas SD2 and the CC together function as positive regulators of Mi-1.2 mediated cell death
    Nucleocytoplasmic distribution is required for activation of resistance by the potato NB-LRR receptor Rx1 and is balanced by its functional domains
    Slootweg, E.J. ; Roosien, J. ; Spiridon, L.N. ; Petrescu, A.J. ; Tameling, W.I.L. ; Joosten, M.H.A.J. ; Pomp, H. ; Schaik, C.C. van; Dees, R.H.L. ; Borst, J.W. ; Smant, G. ; Schots, A. ; Bakker, J. ; Goverse, A. - \ 2010
    The Plant Cell 22 (2010)12. - ISSN 1040-4651 - p. 4195 - 4215.
    secondary structure prediction - plant-disease resistance - nuclear-localization signals - green fluorescent protein - innate immune-response - leucine-rich repeat - to-cell movement - virus-x - arabidopsis-thaliana - coat protein
    The Rx1 protein, as many resistance proteins of the nucleotide binding–leucine-rich repeat (NB-LRR) class, is predicted to be cytoplasmic because it lacks discernable nuclear targeting signals. Here, we demonstrate that Rx1, which confers extreme resistance to Potato virus X, is located both in the nucleus and cytoplasm. Manipulating the nucleocytoplasmic distribution of Rx1 or its elicitor revealed that Rx1 is activated in the cytoplasm and cannot be activated in the nucleus. The coiled coil (CC) domain was found to be required for accumulation of Rx1 in the nucleus, whereas the LRR domain promoted the localization in the cytoplasm. Analyses of structural subdomains of the CC domain revealed no autonomous signals responsible for active nuclear import. Fluorescence recovery after photobleaching and nuclear fractionation indicated that the CC domain binds transiently to large complexes in the nucleus. Disruption of the Rx1 resistance function and protein conformation by mutating the ATP binding phosphate binding loop in the NB domain, or by silencing the cochaperone SGT1, impaired the accumulation of Rx1 protein in the nucleus, while Rx1 versions lacking the LRR domain were not affected in this respect. Our results support a model in which interdomain interactions and folding states determine the nucleocytoplasmic distribution of Rx1
    A Caenorhabditis elegans Wild Type Defies the Temperature-Size Rule Owing to a Single Nucleotide Polymorphism in tra-3
    Kammenga, J.E. ; Doroszuk, A. ; Riksen, J.A.G. ; Hazendonk, E. ; Spiridon, L.N. ; Petrescu, A.J. ; Tijsterman, M. ; Plasterk, R.H.A. ; Bakker, J. - \ 2007
    Plos Genetics 3 (2007)3. - ISSN 1553-7404
    quantitative trait loci - secondary structure prediction - life-history puzzle - drosophila-melanogaster - body-size - cell-size - c-elegans - transcription factor - sex-determination - cold-acclimation
    Ectotherms rely for their body heat on surrounding temperatures. A key question in biology is why most ectotherms mature at a larger size at lower temperatures, a phenomenon known as the temperature¿size rule. Since temperature affects virtually all processes in a living organism, current theories to explain this phenomenon are diverse and complex and assert often from opposing assumptions. Although widely studied, the molecular genetic control of the temperature¿size rule is unknown. We found that the Caenorhabditis elegans wild-type N2 complied with the temperature¿size rule, whereas wild-type CB4856 defied it. Using a candidate gene approach based on an N2 × CB4856 recombinant inbred panel in combination with mutant analysis, complementation, and transgenic studies, we show that a single nucleotide polymorphism in tra-3 leads to mutation F96L in the encoded calpain-like protease. This mutation attenuates the ability of CB4856 to grow larger at low temperature. Homology modelling predicts that F96L reduces TRA-3 activity by destabilizing the DII-A domain. The data show that size adaptation of ectotherms to temperature changes may be less complex than previously thought because a subtle wild-type polymorphism modulates the temperature responsiveness of body size. These findings provide a novel step toward the molecular understanding of the temperature¿size rule, which has puzzled biologists for decades.
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