Plants employ diverse responses mediated by phytohormones to defend themselves against pathogens and herbivores. Adapted pathogens and herbivores often manipulate these responses to their benefit. Previously, we demonstrated that Turnip mosaic virus (TuMV) infection suppresses callose deposition, an important plant defense induced in response to feeding by its aphid vector, the green peach aphid (Myzus persicae), and increases aphid fecundity compared with uninfected control plants. Further, we determined that production of a single TuMV protein, Nuclear Inclusion a-Protease (NIa-Pro) domain, was responsible for changes in host plant physiology and increased green peach aphid reproduction. To characterize the underlying molecular mechanisms of this phenomenon, we examined the role of three phytohormone signaling pathways, jasmonic acid, salicylic acid, and ethylene (ET), in TuMV-infected Arabidopsis (Arabidopsis thaliana), with or without aphid herbivory. Experiments with Arabidopsis mutants ethylene insensitive2 and ethylene response1, and chemical inhibitors of ET synthesis and perception (aminoethoxyvinyl-glycine and 1-methylcyclopropene, respectively), show that the ET signaling pathway is required for TuMV-mediated suppression of Arabidopsis resistance to the green peach aphid. Additionally, transgenic expression of NIa-Pro in Arabidopsis alters ET responses and suppresses aphidinduced callose formation in an ET-dependent manner. Thus, disruption of ET responses in plants is an additional function of NIaPro, a highly conserved potyvirus protein. Virus-induced changes in ET responses may mediate vector-plant interactions more broadly and thus represent a conserved mechanism for increasing transmission by insect vectors across generations.

Many plant and animal viruses are spread by insect vectors. Cauliflower mosaic virus (CaMV) is aphid-transmitted, with the virus being taken up from specialized transmission bodies (TB) formed within infected plant cells. However, the precise events during TB-mediated virus acquisition by aphids are unknown. Here, we show that TBs react instantly to the presence of the vector by ultra-rapid and reversible redistribution of their key components onto microtubules throughout the cell. Enhancing or inhibiting this TB reaction pharmacologically or by using a mutant virus enhanced or inhibited transmission, respectively, confirming its requirement for efficient virus-acquisition. Our results suggest that CaMV can perceive aphid vectors, either directly or indirectly by sharing the host perception. This novel concept in virology, where viruses respond directly or via the host to the outside world, opens new research horizons, that is, investigating the impact of ‘perceptive behaviors’ on other steps of the infection cycle.DOI: http://dx.doi.org/10.7554/eLife.00183.001

SUMMARY Plants employ cell-surface pattern recognition receptors (PRRs) to detect pathogens. Although phytohormones produced during PRR signaling play an essential role in innate immunity, a direct link between PRR activation and hormone regulation is unknown. EFR is a PRR that recognizes bacterial EF-Tu and activates immune signaling. Here we report that EFR regulates the phytohormone jasmonic acid (JA) through direct phosphorylation of a receptor-like cytoplasmic kinase, BIK1. The BIK1 structure revealed that the EFR-phosphorylated sites reside on a uniquely extended loop away from the BIK1 kinase core domain. Phosphomimetic mutations of these sites resulted in increased phytohormones and enhanced resistance to bacterial infections. In addition to its documented plasma membrane localization, BIK1 also localizes to the nucleus and interacts directly with WRKY transcription factors involved in the JA and salicylic acid (SA) regulation. These findings demonstrate the mechanistic basis of signal transduction from PRR to phytohormones, mediated through a PRR-BIK1-WRKY axis.

Vector-borne pathogens influence host characteristics relevant to host–vector contact, increasing pathogen transmission and survival. Previously, we demonstrated that infection with Turnip mosaic virus, a member of one of the largest families of plant-infecting viruses, increases vector attraction and reproduction on infected hosts. These changes were due to a single viral protein, NIa-Pro. Here we show that NIa-Pro responds to the presence of the aphid vector during infection by relocalizing to the vacuole. Remarkably, vacuolar localization is required for NIa-Pro's ability to enhance aphid reproduction on host plants, vacuole localization disappears when aphids are removed, and this phenomenon occurs for another potyvirus, Potato virus Y, suggesting a conserved role for the protein in vector–host interactions. Taken together, these results suggest that potyviruses dynamically respond to the presence of their vectors, promoting insect performance and transmission only when needed.

cCauliflower mosaic virus (CaMV) forms two types of inclusion bodies within infected plant cells: numerous virus factories, which are the sites for viral replication and virion assembly, and a single transmission body (TB), which is specialized for virus transmission by aphid vectors. The TB reacts within seconds to aphid feeding on the host plant by total disruption and redistribution of its principal component, the viral transmission helper protein P2, onto microtubules throughout the cell. At the same time, virions also associate with microtubules. This redistribution of P2 and virions facilitates transmission and is reversible; the TB reforms within minutes after vector departure. Although some virions are present in the TB before disruption, their subsequent massive accumulation on the microtubule network suggests that they also are released from virus factories. Using drug treatments, mutant viruses, and exogenous supply of viral components to infected protoplasts, we show that virions can rapidly exit virus factories and, once in the cytoplasm, accumulate together with the helper protein P2 on the microtubule network. Moreover, we show that during reversion of this phenomenon, virions from the microtubule network can either be incorporated into the reverted TB or return to the virus factories. Our results suggest that CaMV factories are dynamic structures that participate in vector transmission by controlled release and uptake of virions during TB reaction.

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Citrus tristeza virus (CTV), the largest and most complex member of the family Closteroviridae, encodes a unique protein, p33, which shows no homology with other known proteins, however, plays an important role in virus pathogenesis. In this study, we examined some of the characteristics of p33. We show that p33 is a membrane-associated protein that is inserted into the membrane via a transmembrane helix formed by hydrophobic amino acid residues at the C-terminal end of the protein. Removal of this transmembrane domain (TMD) dramatically altered the intracellular localization of p33. Moreover, the TMD alone was sufficient to confer membrane localization of an unrelated protein. Finally, a CTV variant that produced a truncated p33 lacking the TMD was unable to infect sour orange, one of the selected virus hosts, which infection requires p33, suggesting that membrane association of p33 is important for the ability of CTV to extend its host range.

The Arabidopsis resistance protein RPS5 is activated by proteolytic cleavage of the protein kinase PBS1 by the Pseudomonas syringae effector protease AvrPphB. We have previously shown that replacing seven amino acids at the cleavage site of PBS1 with a motif cleaved by the NIa protease of turnip mosaic virus (TuMV) enables RPS5 activation upon TuMV infection. However, this engineered resistance conferred a trailing necrosis phenotype indicative of a cell-death response too slow to contain the virus. We theorized this could result from a positional mismatch within the cell between PBS1TuMV, RPS5, and the NIa protease. To test this, we relocalized PBS1TuMV and RPS5 to cellular sites of NIa accumulation. These experiments revealed that relocation of RPS5 away from the plasma membrane compromised RPS5-dependent cell death in Nicotiana benthamiana, even though PBS1 was efficiently cleaved. As an alternative approach, we tested whether overexpression of plasma membrane–localized PBS1TuMV could enhance RPS5 activation by TuMV. Significantly, overexpressing the PBS1TuMV decoy protein conferred complete resistance to TuMV when delivered by either agrobacterium or by aphid transmission, showing that RPS5-mediated defense responses are effective against bacterial and viral pathogens. Lastly, we have now extended this PBS1 decoy approach to soybean by modifying a soybean PBS1 ortholog to be cleaved by the NIa protease of soybean mosaic virus (SMV). Transgenic overexpression of this soybean PBS1 decoy conferred immunity to SMV, demonstrating that we can use endogenous PBS1 proteins in crop plants to engineer economically relevant disease resistant traits.