title: The Tomato Nucleotide-binding Leucine-rich Repeat Immune Receptor I-2 Couples DNA-binding to Nucleotide-binding Domain Nucleotide Exchange.
abstract: Plant nucleotide-binding leucine-rich repeat (NLR) proteins enable plants to recognize and respond to pathogen attack. Previously, we demonstrated that the Rx1 NLR of potato is able to bind and bend DNA in vitro. DNA binding in situ requires its genuine activation following pathogen perception. However, it is unknown whether other NLR proteins are also able to bind DNA. Nor is it known how DNA binding relates to the ATPase activity intrinsic to NLR switch function required to immune activation. Here we investigate these issues using a recombinant protein corresponding to the N-terminal coiled-coil and nucleotide-binding domain regions of the I-2 NLR of tomato. Wild type I-2 protein bound nucleic acids with a preference of ssDNA ≈ dsDNA > ssRNA, which is distinct from Rx1. I-2 induced bending and melting of DNA. Notably, ATP enhanced DNA binding relative to ADP in the wild type protein, the null P-loop mutant K207R, and the autoactive mutant S233F. DNA binding was found to activate the intrinsic ATPase activity of I-2. Because DNA binding by I-2 was decreased in the presence of ADP when compared with ATP, a cyclic mechanism emerges; activated ATP-associated I-2 binds to DNA, which enhances ATP hydrolysis, releasing ADP-bound I-2 from the DNA. Thus DNA binding is a general property of at least a subset of NLR proteins, and NLR activation is directly linked to its activity at DNA.
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: Stepan
authors.initials: S
authors.lastname: Fenyk
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Dixon
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: William H
authors.initials: WH
authors.lastname: Gittens
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: Philip D
authors.initials: PD
authors.lastname: Townsend
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and.
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Sharples
authors.affiliation: the Department of Chemistry, Durham University, South Road, Durham DH1 3LE, United Kingdom and.
authors.forename: Lars-Olof
authors.initials: LO
authors.lastname: Pålsson
authors.affiliation: the Molecular Plant Pathology, Swammerdam Institute for Life Sciences, University of Amsterdam, 1098 XH, Amsterdam, The Netherlands.
authors.forename: Frank L W
authors.initials: FL
authors.lastname: Takken
authors.affiliation: From the School of Biological and Biomedical Sciences, the Biophysical Sciences Institute, and m.j.cann@durham.ac.uk.
authors.forename: Martin J
authors.initials: MJ
authors.lastname: Cann
mesh_terms: D000244:Adenosine Diphosphate / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D019943:Amino Acid Substitution; D001665:Binding Sites; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D006868:Hydrolysis; D000090263:Leucine-Rich Repeat Proteins; D018551:Solanum lycopersicum / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D008958:Models, Molecular; D009154:Mutation; D033704:Nucleotide Transport Proteins / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D057865:Plant Immunity; D010940:Plant Proteins / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D011506:Proteins / Q000819:agonists* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D000090263:Leucine-Rich Repeat Proteins; D033704:Nucleotide Transport Proteins; D010446:Peptide Fragments; D010940:Plant Proteins; D011506:Proteins; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D012313:RNA; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases
keywords: ATPases associated with diverse cellular activities (AAA); DNA binding protein; Nod-like receptor (NLR); cellular immune response; nucleotide; plant biochemistry