The tomato Prf complex is a molecular trap for bacterial effectors based on Pto transphosphorylation

The bacteria Pseudomonas syringae is a pathogen of many crop species and one of the model pathogens for studying plant and bacterial arms race coevolution. In the current model, plants perceive bacteria pathogens via plasma membrane receptors, and recognition leads to the activation of general defenses. In turn, bacteria inject proteins called effectors into the plant cell to prevent the activation of immune responses. AvrPto and AvrPtoB are two such proteins that inhibit multiple plant kinases. The tomato plant has reacted to these effectors by the evolution of a cytoplasmic resistance complex. This complex is compromised of two proteins, Prf and Pto kinase, and is capable of recognizing the effector proteins. How the Pto kinase is able to avoid inhibition by the effector proteins is currently unknown. Our data shows how the tomato plant utilizes dimerization of resistance proteins to gain advantage over the faster evolving bacterial pathogen. Here we illustrate that oligomerisation of Prf brings into proximity two Pto kinases allowing them to avoid inhibition by the effectors by transphosphorylation and to activate immune responses

Communicating Information on Eruptions and Their Impacts from the Earliest Times Until the Late Twentieth Century

Volcanoes hold a fascination for human beings and, before they were recorded by literate observers, eruptions were portrayed in art, were recalled in legends and became incorporated into religious practices: being viewed as agents of punishment, bounty or intimidation depending upon their state of activity and the culture involved. In the Middle East the earliest depiction of an eruption is a wall painting dating from the Neolithic at Çatal Hüyük and the earliest record dates from the third millennium BCE. Knowledge of volcanoes increased over time. In some parts of the world knowledge of eruptions was passed down by oral transmission, but as far as written records were concerned, in the first century CE only 9 volcanoes in the Mediterranean region were recognised, together with Mount Cameroon in West Africa. In the next 1000 years the list grew by 17, some 14 of these volcanoes being in Japan. The first recorded eruptions in Indonesia occurred in 1000 and 1006, and volcanoes in newly settled Iceland increased the number to just 48 in 1380 CE. After this the list continued to increase, with important regions such as New Zealand and Hawaii only being added in the past 200 years. Only from 1900 did the rate of growth decline significantly (Simkin et al. 1981: 23; Simkin, 1993 Siebert et al. 2011; Simkin, 1993), but it is sobering to recall that in the twentieth century major eruptions have occurred from volcanoes that were considered inactive or extinct examples including: Mount Lamington - Papua New Guinea, 1951; Mount Arenal - Costa Rica, 1968 and Nyos - Cameroon, 1986. Although there are instances where the human impact of historical eruptions have been compiled - with examples including the 1883 eruption of Krakatau (Simkin and Fiske (1983) and 1943 -1952 eruption of Parícutin (Luhr and Simkin, 1993) - these are exceptions and there remains a significant gap in knowledge about both the short and long-term effects on societies of major eruptions which occurred before the 1980s. Following a broad review the chapter provides a discussion of the ways in which information has been collected, compiled and disseminated from the earliest times until the 1980s in two case study areas: the Azores Islands (Portugal) and southern Italy. In Italy information on eruptions stretches back to prehistoric times and has become progressively better known over more than 2,000 years of written history, yet even here there remain significant gaps in the record even for events that took place between 1900 and 1990. In contrast, located in the middle of the Atlantic, the Azores have been isolated for much of their history and illustrate the difficulties involved in using indigenous sources to compile, not only assessments of impact, but also at a more basic level a complete list of historical events with accurate dates

Microstructure and precipitation in Al-Li-Cu-Mg-(Mn, Zr) alloys

Hot rolled Al-6Li-1Cu-1Mg-0.2Mn (at.%) (Al-1.6Li-2.2Cu-0.9Mg-0.4Mn, wt.%) and Al-6Li-1Cu-1Mg-0.03Zr (at.%) (Al-1.6Li-2.3Cu-1Mg-0.1Zr, wt.%) alloys developed for age forming were studied by tensile testing, electron backscatter diffraction (EBSD), three-dimensional atom probe (3DAP), transmission electron microscopy (TEM) and differential scanning calorimetry (DSC). For both alloys, DSC analysis shows that ageing at 150°C leads initially to formation of zones/clusters, which are later gradually replaced by S phase. On ageing at 190°C, S phase formation is completed within 12 h. The precipitates identified by 3D atom probe and TEM can be classified into (a) Li-rich clusters containing Cu and Mg, (b) a plate-shaped metastable precipitate (similar to GPB2 zones/S''), (c) S phase and (d) delta' spherical particles rich in Li. The Zr containing alloy also contains beta' (Al3Zr) precipitates and composite beta'/delta' particles. The beta' precipitates reduces recrystallisation and grain growth leading to fine grains and subgrains

Host inhibition of a bacterial virulence effector triggers immunity to infection

Plant pathogenic bacteria secrete effector proteins that attack the host signaling machinery to suppress immunity. Effectors can be recognized by hosts leading to immunity. One such effector is AvrPtoB of Pseudomonas syringae, which degrades host protein kinases, such as tomato Fen, through an E3 ligase domain. Pto kinase, which is highly related to Fen, recognizes AvrPtoB in conjunction with the resistance protein Prf. Here we show that Pto is resistant to AvrPtoB-mediated degradation because it inactivates the E3 ligase domain. AvrPtoB ubiquitinated Fen within the catalytic cleft, leading to its breakdown and loss of the associated Prf protein. Pto avoids this by phosphorylating and inactivating the AvrPtoB E3 domain. Thus, inactivation of a pathogen virulence molecule is one mechanism by which plants resist disease

Prf immune complexes of tomato are oligomeric and contain multiple Pto-like kinases that diversify effector recognition

Cytoplasmic recognition of pathogen virulence effectors by plant NB-LRR proteins leads to strong induction of defence responses termed effector triggered immunity (ETI). In tomato, a protein complex containing the NB-LRR protein Prf and the protein kinase Pto confers recognition of the Pseudomonas syringae effectors AvrPto and AvrPtoB. Although structurally unrelated, AvrPto and AvrPtoB interact with similar residues in the Pto catalytic cleft to activate ETI via an unknown mechanism. Here we show that the Prf complex is oligomeric, containing at least two molecules of Prf. Within the complex, Prf can associate with Pto or one of several Pto family members including Fen, Pth2, Pth3, or Pth5. The dimerization surface for Prf is the novel N-terminal domain, which also coordinates an intramolecular interaction with the remainder of the molecule, and binds Pto kinase or a family member. Thus, association of two Prf N-terminal domains brings the associated kinases into close promixity. Tomato lines containing Prf complexed with Pth proteins but not Pto possessed greater immunity against P. syringae than tomatoes lacking Prf. This demonstrates that incorporation of non-Pto kinases into the Prf complex extends the number of effector proteins that can be recognized

Gene gain and loss during evolution of obligate parasitism in the white rust pathogen of Arabidopsis thaliana

Biotrophic eukaryotic plant pathogens require a living host for their growth and form an intimate haustorial interface with parasitized cells. Evolution to biotrophy occurred independently in fungal rusts and powdery mildews, and in oomycete white rusts and downy mildews. Biotroph evolution and molecular mechanisms of biotrophy are poorly understood. It has been proposed, but not shown, that obligate biotrophy results from (i) reduced selection for maintenance of biosynthetic pathways and (ii) gain of mechanisms to evade host recognition or suppress host defence. Here we use Illumina sequencing to define the genome, transcriptome, and gene models for the obligate biotroph oomycete and Arabidopsis parasite, Albugo laibachii. A. laibachii is a member of the Chromalveolata, which incorporates Heterokonts (containing the oomycetes), Apicomplexa (which includes human parasites like Plasmodium falciparum and Toxoplasma gondii), and four other taxa. From comparisons with other oomycete plant pathogens and other chromalveolates, we reveal independent loss of molybdenum-cofactor-requiring enzymes in downy mildews, white rusts, and the malaria parasite P. falciparum. Biotrophy also requires ‘‘effectors’’ to suppress host defence; we reveal RXLR and Crinkler effectors shared with other oomycetes, and also discover and verify a novel class of effectors, the ‘‘CHXCs’’, by showing effector delivery and effector functionality. Our findings suggest that evolution to progressively more intimate association between host and parasite results in reduced selection for retention of certain biosynthetic pathways, and particularly reduced selection for retention of molybdopterinrequiring biosynthetic pathways. These mechanisms are not only relevant to plant pathogenic oomycetes but also to human pathogens within the Chromalveolata

Transphosphorylation is required for Ser-198 and Thr-199 phosphorylation.

<p>(<b>A</b>) <i>In trans</i> inhibition of Pto S198 and T199 phosphorylation. The slower migrating from of Pto is suppressed <i>in trans</i> by prf^K1128A and pto^D164N, but not pto^S189A/T199A, pto^S189A and pto^T199A. Pto-5Myc, Pto-FLAG, pto mutant-FLAG, AvrPto, Prf-3HA, prf^K1128A-3HA constructs were transiently expressed in wild-type <i>N. benthamiana</i> as indicated, Prf-3HA and prf^K1128A-3HA were immunoprecipitated (IP) using anti-HA antibodies. Immunoblots (IB) were performed with the antibodies indicated on the left. (<b>B</b>) The slower migrating from of Pto was suppressed <i>in trans</i> by prf^K1128A and pto^D164N, but not by pto^S189A pto^T199A and pto^S189A/T199A. Pto-5Myc, Pto-FLAG, pto mutant-FLAG, AvrPto, Prf-3HA, prf^K1128A-3HA constructs were transiently expressed in wild-type <i>N. benthamiana</i> as indicated. Prf-3HA and prf^K1128A-3HA were immunoprecipitated using anti-HA antibodies. The relative abundance of slow- and fast-migrating forms of Pto-5Myc after AvrPto recognition was quantified two days post infiltration, from anti-Myc immunoblots using Quantity One, Bio-Rad (adjusted volume = [CNT*mm2] data counts/mm²).</p

Double phosphorylation of Pto peptide 187–202 and 188–202 upon activation of signalling.

<p>Prf-3HA, prf^K1128A –3HA, prf^D1416V –3HA, N-term -3HA (prf^1–546 –3HA), Pto-FLAG AvrPto and AvrPtoB were expressed transiently in <i>N. benthamiana</i> under the control of 35S promoter; the total amount of Pto-FLAG was immunoprecipitated using anti-FLAG antibodies. The number of peptides identified with 0, 1, and 2 phosphorylation events is indicated.</p>*<p>ligand-independent HR.</p