StPIP1, a PAMP-induced peptide in potato, elicits plant defenses and is associated with disease symptom severity in a compatible interaction with potato virus Y

The role of small secreted peptides in plant defense responses to viruses has been seldom investigated. Here, we report a role for potato (Solanum tuberosum) PIP1, a gene predicted to encode a member of the PAMP-induced peptide (PIP) family, in the response of potato to potato virus Y infection (PVY). We show that exogenous application of synthetic StPIP1 to potato leaves and nodes increased the production of reactive oxygen species and the expression of plant defense-related genes, showing that StPIP1 triggers early defense responses. In support of this hypothesis, transgenic potato plants that constitutively overexpress StPIP1 had higher levels of leaf callose deposition, and based on measurements of viral RNA titers, were less susceptible to infection by a compatible PVY strain. Interestingly, systemic infection of StPIP1-overexpressing lines with PVY resulted in clear rugose mosaic symptoms that were absent or very mild in infected non-transgenic plants. A transcriptomics analysis revealed that marker genes associated with both pattern-triggered immunity and effector-triggered immunity were induced in infected StPIP1-overexpressors but not in non-transgenic plants. Together, our results reveal a role for StPIP1 in eliciting plant defense responses and in regulating plant anti-viral immunity

Synergistic Activation of Dopamine D1 and TrkB Receptors Mediate Gain Control of Synaptic Plasticity in the Basolateral Amygdala

Fear memory formation is thought to require dopamine, brain-derived neurotrophic factor (BDNF) and zinc release in the basolateral amygdala (BLA), as well as the induction of long term potentiation (LTP) in BLA principal neurons. However, no study to date has shown any relationship between these processes in the BLA. Here, we have used in vitro whole-cell patch clamp recording from BLA principal neurons to investigate how dopamine, BDNF, and zinc release may interact to modulate the LTP induction in the BLA. LTP was induced by either theta burst stimulation (TBS) protocol or spaced 5 times high frequency stimulation (5xHFS). Significantly, both TBS and 5xHFS induced LTP was fully blocked by the dopamine D1 receptor antagonist, SCH23390. LTP induction was also blocked by the BDNF scavenger, TrkB-FC, the zinc chelator, DETC, as well as by an inhibitor of matrix metalloproteinases (MMPs), gallardin. Conversely, prior application of the dopamine reuptake inhibitor, GBR12783, or the D1 receptor agonist, SKF39393, induced robust and stable LTP in response to a sub-threshold HFS protocol (2xHFS), which does not normally induce LTP. Similarly, prior activation of TrkB receptors with either a TrkB receptor agonist, or BDNF, also reduced the threshold for LTP-induction, an effect that was blocked by the MEK inhibitor, but not by zinc chelation. Intriguingly, the TrkB receptor agonist-induced reduction of LTP threshold was fully blocked by prior application of SCH23390, and the reduction of LTP threshold induced by GBR12783 was blocked by prior application of TrkB-FC. Together, our results suggest a cellular mechanism whereby the threshold for LTP induction in BLA principal neurons is critically dependent on the level of dopamine in the extracellular milieu and the synergistic activation of postsynaptic D1 and TrkB receptors. Moreover, activation of TrkB receptors appears to be dependent on concurrent release of zinc and activation of MMPs

Pathogen-induced AdDjSKI of the wild peanut, Arachisdiogoi, potentiates tolerance of multiple stresses in E. coli andtobacco

A gene encoding a serine-rich DnaJIII protein called AdDjSKI that has a 4Fe-4S cluster domain was found to be differentially upregulated in the wild peanut, Arachis diogoi in its resistance responses against the late leaf spot causing fungal pathogen Phaeoisariopsis personata when compared with the cultivated peanut, Arachis hypogaea. AdDjSKI is induced in multiple stress conditions in A. diogoi. Recombinant E. coli cells expressing AdDjSKI showed better growth kinetics when compared with vector control cells under salinity, osmotic, acidic and alkaline stress conditions. Overexpression of this type three J-protein potentiates not only abiotic stress tolerance in Nicotiana tabacum var. Samsun, but also enhances its disease resistance against the phytopathogenic fungi Phytophthora parasitica pv nicotianae and Sclerotinia sclerotiorum. In the present study we show transcriptional upregulation of APX, Mn-SOD and HSP70 under heat stress and increased transcripts of PR genes in response to fungal infection. This transmembrane-domain-containing J protein displays punctate localization in chloroplasts. AdDjSKI appears to ensure proper folding of proteins associated with the photosynthetic machinery under stress

Ancient co-option of an amino acid ABC transporter locus in Pseudomonas syringae for host signal-dependent virulence gene regulation.

Pathogenic bacteria frequently acquire virulence traits via horizontal gene transfer, yet additional evolutionary innovations may be necessary to integrate newly acquired genes into existing regulatory pathways. The plant bacterial pathogen Pseudomonas syringae relies on a horizontally acquired type III secretion system (T3SS) to cause disease. T3SS-encoding genes are induced by plant-derived metabolites, yet how this regulation occurs, and how it evolved, is poorly understood. Here we report that the two-component system AauS-AauR and substrate-binding protein AatJ, proteins encoded by an acidic amino acid-transport (aat) and -utilization (aau) locus in P. syringae, directly regulate T3SS-encoding genes in response to host aspartate and glutamate signals. Mutants of P. syringae strain DC3000 lacking aauS, aauR or aatJ expressed lower levels of T3SS genes in response to aspartate and glutamate, and had decreased T3SS deployment and virulence during infection of Arabidopsis. We identified an AauR-binding motif (Rbm) upstream of genes encoding T3SS regulators HrpR and HrpS, and demonstrated that this Rbm is required for maximal T3SS deployment and virulence of DC3000. The Rbm upstream of hrpRS is conserved in all P. syringae strains with a canonical T3SS, suggesting AauR regulation of hrpRS is ancient. Consistent with a model of conserved function, an aauR deletion mutant of P. syringae strain B728a, a bean pathogen, had decreased T3SS expression and growth in host plants. Together, our data suggest that, upon acquisition of T3SS-encoding genes, a strain ancestral to P. syringae co-opted an existing AatJ-AauS-AauR pathway to regulate T3SS deployment in response to specific host metabolite signals

Opposing functions of the plant TOPLESS gene family during SNC1-mediated autoimmunity.

Regulation of the plant immune system is important for controlling the specificity and amplitude of responses to pathogens and in preventing growth-inhibiting autoimmunity that leads to reductions in plant fitness. In previous work, we reported that SRFR1, a negative regulator of effector-triggered immunity, interacts with SNC1 and EDS1. When SRFR1 is non-functional in the Arabidopsis accession Col-0, SNC1 levels increase, causing a cascade of events that lead to autoimmunity phenotypes. Previous work showed that some members of the transcriptional co-repressor family TOPLESS interact with SNC1 to repress negative regulators of immunity. Therefore, to explore potential connections between SRFR1 and TOPLESS family members, we took a genetic approach that examined the effect of each TOPLESS member in the srfr1 mutant background. The data indicated that an additive genetic interaction exists between SRFR1 and two members of the TOPLESS family, TPR2 and TPR3, as demonstrated by increased stunting and elevated PR2 expression in srfr1 tpr2 and srfr1 tpr2 tpr3 mutants. Furthermore, the tpr2 mutation intensifies autoimmunity in the auto-active snc1-1 mutant, indicating a novel role of these TOPLESS family members in negatively regulating SNC1-dependent phenotypes. This negative regulation can also be reversed by overexpressing TPR2 in the srfr1 tpr2 background. Similar to TPR1 that positively regulates snc1-1 phenotypes by interacting with SNC1, we show here that TPR2 directly binds the N-terminal domain of SNC1. In addition, TPR2 interacts with TPR1 in vivo, suggesting that the opposite functions of TPR2 and TPR1 are based on titration of SNC1-TPR1 complexes by TPR2 or altered functions of a SNC1-TPR1-TPR2 complex. Thus, this work uncovers diverse functions of individual members of the TOPLESS family in Arabidopsis and provides evidence for the additive effect of transcriptional and post-transcriptional regulation of SNC1

The bacterial type III-secreted protein AvrRps4 is a bipartite effector

<div><p>Bacterial effector proteins secreted into host plant cells manipulate those cells to the benefit of the pathogen, but effector-triggered immunity (ETI) occurs when effectors are recognized by host resistance proteins. The RPS4/RRS1 pair recognizes the <i>Pseudomonas syringae</i> pv. pisi effector AvrRps4. AvrRps4 is processed <i>in planta</i> into AvrRps4^N (133 amino acids), homologous to the N-termini of other effectors including the native <i>P</i>. <i>syringae</i> pv. tomato strain DC3000 effector HopK1, and AvrRps4^C (88 amino acids). Previous data suggested that AvrRps4^C alone is necessary and sufficient for resistance when overexpressed in heterologous systems. We show that delivering AvrRps4^C from DC3000, but not from a DC3000 <i>hopK1</i>^<i>-</i> strain, triggers resistance in the Arabidopsis accession Col-0. Delivering AvrRps4^C in tandem with AvrRps4^N, or as a chimera with HopK1^N, fully complements AvrRps4-triggered immunity. AvrRps4^N in the absence of AvrRps4^C enhances virulence in Col-0. In addition, AvrRps4^N triggers a hypersensitive response in lettuce that is attenuated by coexpression of AvrRps4^C, further supporting the role of AvrRps4^N as a bona fide effector domain. Based on these results we propose that evolutionarily, fusion of AvrRps4^C to AvrRps4^N may have counteracted recognition of AvrRps4^N, and that the plant <i>RPS4/RRS1</i> resistance gene pair was selected as a countermeasure. We conclude that AvrRps4 represents an unusual chimeric effector, with recognition in Arabidopsis by RPS4/RRS1 requiring the presence of both processed effector moieties.</p></div

Bacteria secreting AvrRps4^N and AvrRps4^C in tandem trigger resistance comparable to wild-type AvrRps4.

<p>Col-0 plants were inoculated with 5x10⁴ cfu/mL suspensions of the indicated DC3000 <i>hopK1</i>^<i>-</i> strains. AvrRps4 variants were delivered from separate broad host range plasmids (top: pML123 constructs; bottom: pVSP61 constructs). Error bars denote standard deviation. Values are averages from four independent experiments with triplicate samples, and error bars denote standard deviation, with letters indicating statistically significant differences (<i>P</i><0.05).</p

AvrRps4^N overexpression enhances bacterial virulence.

<p><i>In planta</i> bacterial growth was measured in two independent Col-0 AvrRps4^N lines (N2 and N8) and untransformed Col-0 after inoculation with DC3000 (top) and DC3000 <i>hopK1</i>^<i>-</i> (bottom) at 5x10⁴ cfu/mL in the presence of dexamethasone (Dex) or ethanol (mock). Values are averages from two independent experiments with quadruplicate samples, and error bars denote standard deviation. Asterisks denote statistically significant differences compared to bacterial growth in mock- and Dex-treated Col-0 based on two-tailed Student's t-tests (ns: non-significant, *<i>P</i><0.05, **<i>P</i><0.01, ****<i>P</i><0.0001).</p

AvrRps4^N and HopK1^N are sufficient to trigger a hypersensitive response in lettuce.

<p>(A) Agrobacterium C58C1 containing the constructs for transient expression of N-terminally HA-tagged AvrRps4 full-length (4^FL), AvrRps4^N (4^N), AvrRps4^C (4^C), and HA-pBA empty vector (EV) was infiltrated into <i>Lactuca sativa</i> cv. Kordaat at an O.D. of 0.3. (B) Equivalent experiment with HopK1 full-length (K1^FL), HopK1^N (K1^N), HopK1^C (K1^C), and HA-pBA empty vector (EV). Cell death phenotypes in (A) and (B) were visualized three days post-infiltration. These experiments were repeated four times with identical results.</p

ETI is attenuated when AvrRpm1SP-AvrRps4 is delivered from DC3000 <i>hopK1</i>^- compared to DC3000.

<p>(A) Col-0 plants were inoculated with 5x10⁴ cfu/mL suspensions of either wild-type DC3000 or the <i>hopK1</i> deletion strain DC3000 <i>hopK1</i>^<i>-</i> containing empty vector (EV), or expressing AvrRpm1SP-AvrRps4 (A1SP-AvrRps4) or AvrRps4. Values are averages from three (DC3000) or four (DC3000 <i>hopK1</i>^<i>-</i>) independent experiments with triplicate samples, and error bars denote standard deviation, with letters indicating statistically significant differences (<i>P</i><0.0001). (B) Model of HopK1^N contribution to AvrRpm1SP-AvrRps4-triggered immunity.</p