title: Tomato Pto encodes a functional N-myristoylation motif that is required for signal transduction in Nicotiana benthamiana.
abstract: Pto kinase of tomato (Lycopersicon esculentum) confers resistance to bacterial speck disease caused by Pseudomonas syringae pv. tomato expressing avrPto or avrPtoB. Pto interacts directly with these type-III secreted effectors, leading to induction of defence responses including the hypersensitive response (HR). Signalling by Pto requires the nucleotide-binding site-leucine-rich repeat (NBS-LRR) protein Prf. Little is known of how Pto is controlled prior to or during stimulation, although kinase activity is required for Avr-dependent activation. Here we demonstrate a role for the N-terminus in signalling by Pto. N-terminal residues outside the kinase domain were required for induction of the HR in Nicotiana benthamiana. The N-terminus also contributed to both AvrPto-binding and phosphorylation abilities. Pto residues 1-10 comprise a consensus motif for covalent attachment of myristate, a hydrophobic 14-carbon saturated fatty acid, to the Gly-2 residue. Several lines of evidence indicate that this motif is important for Pto function. A heterologous N-myristoylation motif complemented N-terminal deletion mutants of Pto for Prf-dependent signalling. Signalling by wild-type and mutant forms of Pto was strictly dependent on the Gly-2 residue. The N-myristoylation motif of Pto complemented the cognate motif of AvrPto for avirulence function and membrane association. Furthermore, Pto was myristoylated in vivo dependent on the presence of Gly-2. The subcellular localization of Pto was independent of N-myristoylation, indicating that N-myristoylation is required for some function other than membrane affinity. Consistent with this idea, AvrPtoB was also found to be a soluble protein. The data indicate an important role(s) for the myristoylated N-terminus in Pto signalling.
authors.affiliation: The Sainsbury Laboratory, John Innes Centre, Colney Lane, Norwich, NR4 7UH, UK.
authors.forename: Jeroen S
authors.initials: JS
authors.lastname: de Vries
authors.forename: Vasilios M E
authors.initials: VM
authors.lastname: Andriotis
authors.forename: Ai-Jiuan
authors.initials: AJ
authors.lastname: Wu
authors.forename: John P
authors.initials: JP
authors.lastname: Rathjen
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D000596:Amino Acids / Q000378:metabolism; D018551:Solanum lycopersicum / Q000235:genetics*; D019814:Myristic Acid / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics*; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D015398:Signal Transduction; D014026:Nicotiana / Q000378:metabolism*
chemical_list: D000596:Amino Acids; D010940:Plant Proteins; D019814:Myristic Acid; C084354:Pto protein, Lycopersicon; D017346:Protein Serine-Threonine Kinases