Publications by authors named "Akiko Sugio"

29 Publications

  • Page 1 of 1

Parasitic modulation of host development by ubiquitin-independent protein degradation.

Cell 2021 Sep 17;184(20):5201-5214.e12. Epub 2021 Sep 17.

Department of Crop Genetics, John Innes Centre, Norwich Research Park, Norwich NR4 7UH, UK. Electronic address:

Certain obligate parasites induce complex and substantial phenotypic changes in their hosts in ways that favor their transmission to other trophic levels. However, the mechanisms underlying these changes remain largely unknown. Here we demonstrate how SAP05 protein effectors from insect-vectored plant pathogenic phytoplasmas take control of several plant developmental processes. These effectors simultaneously prolong the host lifespan and induce witches' broom-like proliferations of leaf and sterile shoots, organs colonized by phytoplasmas and vectors. SAP05 acts by mediating the concurrent degradation of SPL and GATA developmental regulators via a process that relies on hijacking the plant ubiquitin receptor RPN10 independent of substrate ubiquitination. RPN10 is highly conserved among eukaryotes, but SAP05 does not bind insect vector RPN10. A two-amino-acid substitution within plant RPN10 generates a functional variant that is resistant to SAP05 activities. Therefore, one effector protein enables obligate parasitic phytoplasmas to induce a plethora of developmental phenotypes in their hosts.
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http://dx.doi.org/10.1016/j.cell.2021.08.029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8525514PMC
September 2021

Editorial: Plant-Arthropod Interactions: Effectors and Elicitors of Arthropods and Their Associated Microbes.

Front Plant Sci 2020 4;11:610160. Epub 2020 Nov 4.

Department of Nematology, Institute for Integrative Genome Biology, University of California, Riverside, Riverside, CA, United States.

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http://dx.doi.org/10.3389/fpls.2020.610160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7672025PMC
November 2020

Big Genes, Small Effectors: Pea Aphid Cassette Effector Families Composed From Miniature Exons.

Front Plant Sci 2020 2;11:1230. Epub 2020 Sep 2.

Department of Plant Pathology, University of Florida, Gainesville, FL, United States.

Aphids secrete proteins from their stylets that evidence indicates function similar to pathogen effectors for virulence. Here, we describe two small candidate effector gene families of the pea aphid, , that share highly conserved secretory signal peptide coding regions and divergent non-secretory coding sequences derived from miniature exons. The KQY candidate effector family contains eleven members with additional isoforms, generated by alternative splicing. Pairwise comparisons indicate possible four unique KQY families based on coding regions without the secretory signal region. KQY1a, a representative of the family, is encoded by a 968 bp mRNA and a gene that spans 45.7 kbp of the genome. The locus consists of 37 exons, 33 of which are 15 bp or smaller. Additional KQY members, as well as members of the KHI family, share similar features. Differential expression analyses indicate that the genes are expressed preferentially in salivary glands. Proteomic analysis on salivary glands and saliva revealed 11 KQY members in salivary proteins, and KQY1a was detected in an artificial diet solution after aphid feeding. A single KQY locus and two KHI loci were identified in , the peach aphid. Of the genes that can be anchored to chromosomes, loci are mostly scattered throughout the genome, except a two-gene region (KQY4/KQY6). We propose that the KQY family expanded in through combinatorial assemblies of a common secretory signal cassette and novel coding regions, followed by classical gene duplication and divergence.
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http://dx.doi.org/10.3389/fpls.2020.01230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7495047PMC
September 2020

Isolation of Insect Bacteriocytes as a Platform for Transcriptomic Analyses.

Methods Mol Biol 2021 ;2170:185-198

Univ Lyon, INSA-Lyon, INRAE, BF2i, UMR203, F-69621, Villeurbanne, France.

Over the past few decades, various techniques have been developed and optimized for the accurate measurement of RNA abundance in cells or tissues. These methods have been instrumental in gaining insight in complex systems such as host-symbiont associations. The pea aphid model has recently emerged as a powerful and experimentally tractable system for studying symbiotic relationships and it is the subject of a growing number of molecular studies. Nevertheless, the lack of standardized protocols for the collection of bacteriocytes, the specialized host cells harboring the symbionts, has limited its use. This chapter provides a simple, step-by-step dissection protocol for the rapid isolation of aphid bacteriocytes. We then describe an adapted protocol for efficient extraction and purification of bacteriocyte RNA that can be used for most downstream transcriptomic analyses.
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http://dx.doi.org/10.1007/978-1-0716-0743-5_13DOI Listing
March 2021

AcDCXR Is a Cowpea Aphid Effector With Putative Roles in Altering Host Immunity and Physiology.

Front Plant Sci 2020 15;11:605. Epub 2020 May 15.

Graduate Program in Biochemistry and Molecular Biology, University of California, Riverside, Riverside, CA, United States.

Cowpea, , is a crop that is essential to semiarid areas of the world like Sub-Sahara Africa. Cowpea is highly susceptible to cowpea aphid, , infestation that can lead to major yield losses. Aphids feed on their host plant by inserting their hypodermal needlelike flexible stylets into the plant to reach the phloem sap. During feeding, aphids secrete saliva, containing effector proteins, into the plant to disrupt plant immune responses and alter the physiology of the plant to their own advantage. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used to identify the salivary proteome of the cowpea aphid. About 150 candidate proteins were identified including diacetyl/L-xylulose reductase (DCXR), a novel enzyme previously unidentified in aphid saliva. DCXR is a member of short-chain dehydrogenases/reductases with dual enzymatic functions in carbohydrate and dicarbonyl metabolism. To assess whether cowpea aphid DCXR (AcDCXR) has similar functions, recombinant AcDCXR was purified and assayed enzymatically. For carbohydrate metabolism, the oxidation of xylitol to xylulose was tested. The dicarbonyl reaction involved the reduction of methylglyoxal, an α-β-dicarbonyl ketoaldehyde, known as an abiotic and biotic stress response molecule causing cytotoxicity at high concentrations. To assess whether cowpea aphids induce methylglyoxal in plants, we measured methylglyoxal levels in both cowpea and pea () plants and found them elevated transiently after aphid infestation. Agrobacterium-mediated transient overexpression of AcDCXR in pea resulted in an increase of cowpea aphid fecundity. Taken together, our results indicate that AcDCXR is an effector with a putative ability to generate additional sources of energy to the aphid and to alter plant defense responses. In addition, this work identified methylglyoxal as a potential novel aphid defense metabolite adding to the known repertoire of plant defenses against aphid pests.
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http://dx.doi.org/10.3389/fpls.2020.00605DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7243947PMC
May 2020

Differential Expression of Candidate Salivary Effector Genes in Pea Aphid Biotypes With Distinct Host Plant Specificity.

Front Plant Sci 2019 22;10:1301. Epub 2019 Oct 22.

INRA, UMR1349, Institute of Genetics, Environment and Plant Protection, Le Rheu, France.

Effector proteins play crucial roles in determining the outcome of various plant-parasite interactions. Aphids inject salivary effector proteins into plants to facilitate phloem feeding, but some proteins might trigger defense responses in certain plants. The pea aphid, , forms multiple biotypes, and each biotype is specialized to feed on a small number of closely related legume species. Interestingly, all the previously identified biotypes can feed on ; hence, it serves as a universal host plant of . We hypothesized that the salivary effector proteins have a key role in determining the compatibility between specific host species and biotypes and that each biotype produces saliva containing a specific mixture of effector proteins due to differential expression of encoding genes. As the first step to address these hypotheses, we conducted two sets of RNA-seq experiments. RNA-seq analysis of dissected salivary glands (SGs) from reference alfalfa- and pea-specialized lines revealed common and line-specific repertoires of candidate salivary effector genes. Based on the results, we created an extended catalogue of salivary effector candidates. Next, we used aphid head samples, which contain SGs, to examine biotype-specific expression patterns of candidate salivary genes. RNA-seq analysis of head samples of alfalfa- and pea-specialized biotypes, each represented by three genetically distinct aphid lines reared on either a universal or specific host plant, showed that a majority of the candidate salivary effector genes was expressed in both biotypes at a similar level. Nonetheless, we identified small sets of genes that were differentially regulated in a biotype-specific manner. Little host plant effect (universal vs. specific) was observed on the expression of candidate salivary genes. Analysis of previously obtained genome re-sequenced data of the two biotypes revealed the copy number variations that might explain the differential expression of some candidate salivary genes. In addition, at least four candidate effector genes that were present in the alfalfa biotype but might not be encoded in the pea biotype were identified. This work sets the stage for future functional characterization of candidate genes potentially involved in the determination of plant specificity of pea aphid biotypes.
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http://dx.doi.org/10.3389/fpls.2019.01301DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6818229PMC
October 2019

Phytoplasma SAP11 effector destabilization of TCP transcription factors differentially impact development and defence of Arabidopsis versus maize.

PLoS Pathog 2019 09 26;15(9):e1008035. Epub 2019 Sep 26.

John Innes Centre, Department of Crop Genetics, Norwich Research Park, Norwich, United Kingdom.

Phytoplasmas are insect-transmitted bacterial pathogens that colonize a wide range of plant species, including vegetable and cereal crops, and herbaceous and woody ornamentals. Phytoplasma-infected plants often show dramatic symptoms, including proliferation of shoots (witch's brooms), changes in leaf shapes and production of green sterile flowers (phyllody). Aster Yellows phytoplasma Witches' Broom (AY-WB) infects dicots and its effector, secreted AYWB protein 11 (SAP11), was shown to be responsible for the induction of shoot proliferation and leaf shape changes of plants. SAP11 acts by destabilizing TEOSINTE BRANCHED 1-CYCLOIDEA-PROLIFERATING CELL FACTOR (TCP) transcription factors, particularly the class II TCPs of the CYCLOIDEA/TEOSINTE BRANCHED 1 (CYC/TB1) and CINCINNATA (CIN)-TCP clades. SAP11 homologs are also present in phytoplasmas that cause economic yield losses in monocot crops, such as maize, wheat and coconut. Here we show that a SAP11 homolog of Maize Bushy Stunt Phytoplasma (MBSP), which has a range primarily restricted to maize, destabilizes specifically TB1/CYC TCPs. SAP11MBSP and SAP11AYWB both induce axillary branching and SAP11AYWB also alters leaf development of Arabidopsis thaliana and maize. However, only in maize, SAP11MBSP prevents female inflorescence development, phenocopying maize tb1 lines, whereas SAP11AYWB prevents male inflorescence development and induces feminization of tassels. SAP11AYWB promotes fecundity of the AY-WB leafhopper vector on A. thaliana and modulates the expression of A. thaliana leaf defence response genes that are induced by this leafhopper, in contrast to SAP11MBSP. Neither of the SAP11 effectors promote fecundity of AY-WB and MBSP leafhopper vectors on maize. These data provide evidence that class II TCPs have overlapping but also distinct roles in regulating development and defence in a dicot and a monocot plant species that is likely to shape SAP11 effector evolution depending on the phytoplasma host range.
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http://dx.doi.org/10.1371/journal.ppat.1008035DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6802841PMC
September 2019

Characterization of two fungal lipoxygenases expressed in Aspergillus oryzae.

J Biosci Bioeng 2018 Oct 24;126(4):436-444. Epub 2018 May 24.

Novozymes Japan Ltd., CB-6 MTG, 1-3 Nakase, Mihama-ku, Chiba 261-8501, Japan. Electronic address:

Two fungal lipoxygenase genes were cloned from a rice pathogen, Magnaporthe salvinii, and the take-all fungus, Gaeumannomyces graminis var. tritici, and successfully expressed in Aspergillus oryzae in secreted form. The lipoxygenases expressed, termed MLOX and GLOX, were purified and characterized to evaluate suitability for industrial applications. Both enzymes were active broadly at pH 4-11 and had optimum temperatures around 60 °C, but they were largely different in substrate specificity. Where MLOX was active broadly on arachidonic acid, EPA and DHA, and even on derivatives of fatty acids, such as methyl linoleate or linoleoyl alcohol, GLOX was more specific to linoleic acid and linolenic acid. The most remarkable difference between the two fungal LOXs was the positional and stereo-specificity of oxygenation reactions on polyunsaturated fatty acids. When using linoleic acid as the substrate, the product of MLOX was 9S-hydroperoxy-(E,Z)-octadecadienoic acid (9S(E,Z)-HPODE), on the other hand, the product of GLOX was 13R(E,Z)-HPODE. The enzymes were evaluated for a couple of potential applications and found to be effective on bleaching colored compounds such as carotenoids.
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http://dx.doi.org/10.1016/j.jbiosc.2018.04.005DOI Listing
October 2018

Fast Evolution and Lineage-Specific Gene Family Expansions of Aphid Salivary Effectors Driven by Interactions with Host-Plants.

Genome Biol Evol 2018 06;10(6):1554-1572

INRA, UMR1349, Institute of Genetics, Environment and Plant Protection, Le Rheu, France.

Effector proteins play crucial roles in plant-parasite interactions by suppressing plant defenses and hijacking plant physiological responses to facilitate parasite invasion and propagation. Although effector proteins have been characterized in many microbial plant pathogens, their nature and role in adaptation to host plants are largely unknown in insect herbivores. Aphids rely on salivary effector proteins injected into the host plants to promote phloem sap uptake. Therefore, gaining insight into the repertoire and evolution of aphid effectors is key to unveiling the mechanisms responsible for aphid virulence and host plant specialization. With this aim in mind, we assembled catalogues of putative effectors in the legume specialist aphid, Acyrthosiphon pisum, using transcriptomics and proteomics approaches. We identified 3,603 candidate effector genes predicted to be expressed in A. pisum salivary glands (SGs), and 740 of which displayed up-regulated expression in SGs in comparison to the alimentary tract. A search for orthologs in 17 arthropod genomes revealed that SG-up-regulated effector candidates of A. pisum are enriched in aphid-specific genes and tend to evolve faster compared with the whole gene set. We also found that a large fraction of proteins detected in the A. pisum saliva belonged to three gene families, of which certain members show evidence consistent with positive selection. Overall, this comprehensive analysis suggests that the large repertoire of effector candidates in A. pisum constitutes a source of novelties promoting plant adaptation to legumes.
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http://dx.doi.org/10.1093/gbe/evy097DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6012102PMC
June 2018

Rapid transcriptional plasticity of duplicated gene clusters enables a clonally reproducing aphid to colonise diverse plant species.

Genome Biol 2017 02 13;18(1):27. Epub 2017 Feb 13.

Earlham Institute, Norwich Research Park, Norwich, NR4 7UZ, UK.

Background: The prevailing paradigm of host-parasite evolution is that arms races lead to increasing specialisation via genetic adaptation. Insect herbivores are no exception and the majority have evolved to colonise a small number of closely related host species. Remarkably, the green peach aphid, Myzus persicae, colonises plant species across 40 families and single M. persicae clonal lineages can colonise distantly related plants. This remarkable ability makes M. persicae a highly destructive pest of many important crop species.

Results: To investigate the exceptional phenotypic plasticity of M. persicae, we sequenced the M. persicae genome and assessed how one clonal lineage responds to host plant species of different families. We show that genetically identical individuals are able to colonise distantly related host species through the differential regulation of genes belonging to aphid-expanded gene families. Multigene clusters collectively upregulate in single aphids within two days upon host switch. Furthermore, we demonstrate the functional significance of this rapid transcriptional change using RNA interference (RNAi)-mediated knock-down of genes belonging to the cathepsin B gene family. Knock-down of cathepsin B genes reduced aphid fitness, but only on the host that induced upregulation of these genes.

Conclusions: Previous research has focused on the role of genetic adaptation of parasites to their hosts. Here we show that the generalist aphid pest M. persicae is able to colonise diverse host plant species in the absence of genetic specialisation. This is achieved through rapid transcriptional plasticity of genes that have duplicated during aphid evolution.
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http://dx.doi.org/10.1186/s13059-016-1145-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5304397PMC
February 2017

Corrigendum: Optimization of Agroinfiltration in Provides a New Tool for Studying the Salivary Protein Functions in the Pea Aphid Complex.

Front Plant Sci 2016 10;7:2046. Epub 2017 Jan 10.

INRA, UMR1349, Institute of Genetics, Environment and Plant Protection Le Rheu, France.

[This corrects the article on p. 1171 in vol. 7, PMID: 27555856.].
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http://dx.doi.org/10.3389/fpls.2016.02046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5222837PMC
January 2017

Optimization of Agroinfiltration in Pisum sativum Provides a New Tool for Studying the Salivary Protein Functions in the Pea Aphid Complex.

Front Plant Sci 2016 9;7:1171. Epub 2016 Aug 9.

INRA, UMR1349, Institute of Genetics, Environment and Plant Protection Le Rheu, France.

Aphids are piercing-sucking insect pests and feed on phloem sap. During feeding, aphids inject a battery of salivary proteins into host plant. Some of these proteins function like effectors of microbial pathogens and influence the outcome of plant-aphid interactions. The pea aphid (Acyrthosiphon pisum) is the model aphid and encompasses multiple biotypes each specialized to one or a few legume species, providing an opportunity to investigate the underlying mechanisms of the compatibility between plants and aphid biotypes. We aim to identify the aphid factors that determine the compatibility with host plants, hence involved in the host plant specialization process, and hypothesize that salivary proteins are one of those factors. Agrobacterium-mediated transient gene expression is a powerful tool to perform functional analyses of effector (salivary) proteins in plants. However, the tool was not established for the legume species that A. pisum feeds on. Thus, we decided to optimize the method for legume plants to facilitate the functional analyses of A. pisum salivary proteins. We screened a range of cultivars of pea (Pisum sativum) and alfalfa (Medicago sativa). None of the M. sativa cultivars was suitable for agroinfiltration under the tested conditions; however, we established a protocol for efficient transient gene expression in two cultivars of P. sativum, ZP1109 and ZP1130, using A. tumefaciens AGL-1 strain and the pEAQ-HT-DEST1 vector. We confirmed that the genes are expressed from 3 to 10 days post-infiltration and that aphid lines of the pea adapted biotype fed and reproduced on these two cultivars while lines of alfalfa and clover biotypes did not. Thus, the pea biotype recognizes these two cultivars as typical pea plants. By using a combination of ZP1109 and an A. pisum line, we defined an agroinfiltration procedure to examine the effect of in planta expression of selected salivary proteins on A. pisum fitness and demonstrated that transient expression of one candidate salivary gene increased the fecundity of the aphids. This result confirms that the agroinfiltration can be used to perform functional analyses of salivary proteins in P. sativum and consequently to study the molecular mechanisms underlying host specialization in the pea aphid complex.
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http://dx.doi.org/10.3389/fpls.2016.01171DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4977312PMC
August 2016

Consequences of coinfection with protective symbionts on the host phenotype and symbiont titres in the pea aphid system.

Insect Sci 2017 Oct 29;24(5):798-808. Epub 2016 Sep 29.

IGEPP, Agrocampus Ouest, INRA, Université de Rennes 1, Université Bretagne-Loire, Rennes, France.

Symbiotic associations between microbes and insects are widespread, and it is frequent that several symbionts share the same host individual. Hence, interactions can occur between these symbionts, influencing their respective abundance within the host with consequences on its phenotype. Here, we investigate the effects of multiple infections in the pea aphid, Acyrthosiphon pisum, which is the host of an obligatory and several facultative symbionts. In particular, we study the influence of a coinfection with 2 protective symbionts: Hamiltonella defensa, which confers protection against parasitoids, and Rickettsiella viridis, which provides protection against fungal pathogens and predators. The effects of Hamiltonella-Rickettsiella coinfection on the respective abundance of the symbionts, host fitness and efficacy of enemy protection were studied. Asymmetrical interactions between the 2 protective symbionts have been found: when they coinfect the same aphid individuals, the Rickettsiella infection affected Hamiltonella abundance within hosts but not the Hamiltonella-mediated protective phenotype while the Hamiltonella infection negatively influences the Rickettsiella-mediated protective phenotype but not its abundance. Harboring the 2 protective symbionts also reduced the survival and fecundity of host individuals. Overall, this work highlights the effects of multiple infections on symbiont abundances and host traits that are likely to impact the maintenance of the symbiotic associations in natural habitats.
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http://dx.doi.org/10.1111/1744-7917.12380DOI Listing
October 2017

Differential gene expression according to race and host plant in the pea aphid.

Mol Ecol 2016 Sep 26;25(17):4197-215. Epub 2016 Aug 26.

Department of Biology, University of York, York, UK.

Host-race formation in phytophagous insects is thought to provide the opportunity for local adaptation and subsequent ecological speciation. Studying gene expression differences amongst host races may help to identify phenotypes under (or resulting from) divergent selection and their genetic, molecular and physiological bases. The pea aphid (Acyrthosiphon pisum) comprises host races specializing on numerous plants in the Fabaceae and provides a unique system for examining the early stages of diversification along a gradient of genetic and associated adaptive divergence. In this study, we examine transcriptome-wide gene expression both in response to environment and across pea aphid races selected to cover the range of genetic divergence reported in this species complex. We identify changes in expression in response to host plant, indicating the importance of gene expression in aphid-plant interactions. Races can be distinguished on the basis of gene expression, and higher numbers of differentially expressed genes are apparent between more divergent races; these expression differences between host races may result from genetic drift and reproductive isolation and possibly divergent selection. Expression differences related to plant adaptation include a subset of chemosensory and salivary genes. Genes showing expression changes in response to host plant do not make up a large portion of between-race expression differences, providing confirmation of previous studies' findings that genes involved in expression differences between diverging populations or species are not necessarily those showing initial plasticity in the face of environmental change.
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http://dx.doi.org/10.1111/mec.13771DOI Listing
September 2016

Genomics of adaptation to host-plants in herbivorous insects.

Brief Funct Genomics 2015 Nov 6;14(6):413-23. Epub 2015 Apr 6.

Herbivorous insects represent the most species-rich lineages of metazoans. The high rate of diversification in herbivorous insects is thought to result from their specialization to distinct host-plants, which creates conditions favorable for the build-up of reproductive isolation and speciation. These conditions rely on constraints against the optimal use of a wide range of plant species, as each must constitute a viable food resource, oviposition site and mating site for an insect. Utilization of plants involves many essential traits of herbivorous insects, as they locate and select their hosts, overcome their defenses and acquire nutrients while avoiding intoxication. Although advances in understanding insect-plant molecular interactions have been limited by the complexity of insect traits involved in host use and the lack of genomic resources and functional tools, recent studies at the molecular level, combined with large-scale genomics studies at population and species levels, are revealing the genetic underpinning of plant specialization and adaptive divergence in non-model insect herbivores. Here, we review the recent advances in the genomics of plant adaptation in hemipterans and lepidopterans, two major insect orders, each of which includes a large number of crop pests. We focus on how genomics and post-genomics have improved our understanding of the mechanisms involved in insect-plant interactions by reviewing recent molecular discoveries in sensing, feeding, digesting and detoxifying strategies. We also present the outcomes of large-scale genomics approaches aimed at identifying loci potentially involved in plant adaptation in these insects.
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http://dx.doi.org/10.1093/bfgp/elv015DOI Listing
November 2015

Plant-insect interactions under bacterial influence: ecological implications and underlying mechanisms.

J Exp Bot 2015 Feb 10;66(2):467-78. Epub 2014 Nov 10.

INRA, Institut de Génétique, Environnement et Protection des Plantes, UMR 1349 IGEPP, Domaine de la Motte, 35653 Le Rheu Cedex, France

Plants and insects have been co-existing for more than 400 million years, leading to intimate and complex relationships. Throughout their own evolutionary history, plants and insects have also established intricate and very diverse relationships with microbial associates. Studies in recent years have revealed plant- or insect-associated microbes to be instrumental in plant-insect interactions, with important implications for plant defences and plant utilization by insects. Microbial communities associated with plants are rich in diversity, and their structure greatly differs between below- and above-ground levels. Microbial communities associated with insect herbivores generally present a lower diversity and can reside in different body parts of their hosts including bacteriocytes, haemolymph, gut, and salivary glands. Acquisition of microbial communities by vertical or horizontal transmission and possible genetic exchanges through lateral transfer could strongly impact on the host insect or plant fitness by conferring adaptations to new habitats. Recent developments in sequencing technologies and molecular tools have dramatically enhanced opportunities to characterize the microbial diversity associated with plants and insects and have unveiled some of the mechanisms by which symbionts modulate plant-insect interactions. Here, we focus on the diversity and ecological consequences of bacterial communities associated with plants and herbivorous insects. We also highlight the known mechanisms by which these microbes interfere with plant-insect interactions. Revealing such mechanisms in model systems under controlled environments but also in more natural ecological settings will help us to understand the evolution of complex multitrophic interactions in which plants, herbivorous insects, and micro-organisms are inserted.
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http://dx.doi.org/10.1093/jxb/eru435DOI Listing
February 2015

The small phytoplasma virulence effector SAP11 contains distinct domains required for nuclear targeting and CIN-TCP binding and destabilization.

New Phytol 2014 May 19;202(3):838-848. Epub 2014 Feb 19.

Cell and Developmental Biology, The John Innes Centre, Norwich Research Park, Norwich, NR4 7UH, UK.

Phytoplasmas are insect-transmitted bacterial phytopathogens that secrete virulence effectors and induce changes in the architecture and defense response of their plant hosts. We previously demonstrated that the small (± 10 kDa) virulence effector SAP11 of Aster Yellows phytoplasma strain Witches' Broom (AY-WB) binds and destabilizes Arabidopsis CIN (CINCINNATA) TCP (TEOSINTE-BRANCHED, CYCLOIDEA, PROLIFERATION FACTOR 1 AND 2) transcription factors, resulting in dramatic changes in leaf morphogenesis and increased susceptibility to phytoplasma insect vectors. SAP11 contains a bipartite nuclear localization signal (NLS) that targets this effector to plant cell nuclei. To further understand how SAP11 functions, we assessed the involvement of SAP11 regions in TCP binding and destabilization using a series of mutants. SAP11 mutants lacking the entire N-terminal domain, including the NLS, interacted with TCPs but did not destabilize them. SAP11 mutants lacking the C-terminal domain were impaired in both binding and destabilization of TCPs. These SAP11 mutants did not alter leaf morphogenesis. A SAP11 mutant that did not accumulate in plant nuclei (SAP11ΔNLS-NES) was able to bind and destabilize TCP transcription factors, but instigated weaker changes in leaf morphogenesis than wild-type SAP11. Overall the results suggest that phytoplasma effector SAP11 has a modular organization in which at least three domains are required for efficient CIN-TCP destabilization in plants.
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http://dx.doi.org/10.1111/nph.12721DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4235307PMC
May 2014

The genome biology of phytoplasma: modulators of plants and insects.

Curr Opin Microbiol 2012 Jun 28;15(3):247-54. Epub 2012 Apr 28.

Department of Disease and Stress Biology, The John Innes Centre, Norwich Research Park, Norwich, NR4 7UH, United Kingdom.

Phytoplasmas are bacterial pathogens of plants that are transmitted by insects. These bacteria uniquely multiply intracellularly in both plants (Plantae) and insects (Animalia). Similarly to bacterial endosymbionts, phytoplasmas have reduced genomes with limited metabolic capabilities. Nonetheless, the chromosomes of many phytoplasmas are rich in repeated DNA consisting of mobile elements. Phytoplasmas produce an arsenal of effectors most of which are encoded on these mobile elements and on plasmids. These effectors target conserved plant transcription factors resulting in witches' broom and leafy flower symptoms and suppression of plant defense to insect vectors that transmit the phytoplasmas. Future studies of these fascinating microbes will generate a wealth of new knowledge about forces that shape genomes and microbial interactions with multicellular hosts.
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http://dx.doi.org/10.1016/j.mib.2012.04.002DOI Listing
June 2012

Phytoplasma protein effector SAP11 enhances insect vector reproduction by manipulating plant development and defense hormone biosynthesis.

Proc Natl Acad Sci U S A 2011 Nov 7;108(48):E1254-63. Epub 2011 Nov 7.

Department of Disease and Stress Biology, The John Innes Centre, Norwich NR4 7UH, United Kingdom.

Phytoplasmas are insect-transmitted phytopathogenic bacteria that can alter plant morphology and the longevity and reproduction rates and behavior of their insect vectors. There are various examples of animal and plant parasites that alter the host phenotype to attract insect vectors, but it is unclear how these parasites accomplish this. We hypothesized that phytoplasmas produce effectors that modulate specific targets in their hosts leading to the changes in plant development and insect performance. Previously, we sequenced and mined the genome of Aster Yellows phytoplasma strain Witches' Broom (AY-WB) and identified 56 candidate effectors. Here, we report that the secreted AY-WB protein 11 (SAP11) effector modulates plant defense responses to the advantage of the AY-WB insect vector Macrosteles quadrilineatus. SAP11 binds and destabilizes Arabidopsis CINCINNATA (CIN)-related TEOSINTE BRANCHED1, CYCLOIDEA, PROLIFERATING CELL FACTORS 1 and 2 (TCP) transcription factors, which control plant development and promote the expression of lipoxygenase (LOX) genes involved in jasmonate (JA) synthesis. Both the Arabidopsis SAP11 lines and AY-WB-infected plants produce less JA on wounding. Furthermore, the AY-WB insect vector produces more offspring on AY-WB-infected plants, SAP11 transgenic lines, and plants impaired in CIN-TCP and JA synthesis. Thus, SAP11-mediated destabilization of CIN-TCPs leads to the down-regulation of LOX2 expression and JA synthesis and an increase in M. quadrilineatus progeny. Phytoplasmas are obligate inhabitants of their plant host and insect vectors, in which the latter transmits AY-WB to a diverse range of plant species. This finding demonstrates that pathogen effectors can reach beyond the pathogen-host interface to modulate a third organism in the biological interaction.
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http://dx.doi.org/10.1073/pnas.1105664108DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3228479PMC
November 2011

Phytoplasma effector SAP54 induces indeterminate leaf-like flower development in Arabidopsis plants.

Plant Physiol 2011 Oct 17;157(2):831-41. Epub 2011 Aug 17.

Department of Disease and Stress Biology , John Innes Centre, Norwich NR47UH, United Kingdom.

Phytoplasmas are insect-transmitted bacterial plant pathogens that cause considerable damage to a diverse range of agricultural crops globally. Symptoms induced in infected plants suggest that these phytopathogens may modulate developmental processes within the plant host. We report herein that Aster Yellows phytoplasma strain Witches' Broom (AY-WB) readily infects the model plant Arabidopsis (Arabidopsis thaliana) ecotype Columbia, inducing symptoms that are characteristic of phytoplasma infection, such as the production of green leaf-like flowers (virescence and phyllody) and increased formation of stems and branches (witches' broom). We found that the majority of genes encoding secreted AY-WB proteins (SAPs), which are candidate effector proteins, are expressed in Arabidopsis and the AY-WB insect vector Macrosteles quadrilineatus (Hemiptera; Cicadellidae). To identify which of these effector proteins induce symptoms of phyllody and virescence, we individually expressed the effector genes in Arabidopsis. From this screen, we have identified a novel AY-WB effector protein, SAP54, that alters floral development, resulting in the production of leaf-like flowers that are similar to those produced by plants infected with this phytoplasma. This study offers novel insight into the effector profile of an insect-transmitted plant pathogen and reports to our knowledge the first example of a microbial pathogen effector protein that targets flower development in a host.
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http://dx.doi.org/10.1104/pp.111.181586DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3192582PMC
October 2011

Diverse targets of phytoplasma effectors: from plant development to defense against insects.

Annu Rev Phytopathol 2011 ;49:175-95

Department of Disease and Stress Biology, The John Innes Centre, Norwich Research Park, Norwich NR1 3LY, United Kingdom.

Phytoplasma research begins to bloom (75). Indeed, this review shows that substantial progress has been made with the identification of phytoplasma effectors that alter flower development, induce witches' broom, affect leaf shape, and modify plant-insect interactions. Phytoplasmas have a unique life cycle among pathogens, as they invade organisms of two distinct kingdoms, namely plants (Plantae) and insects (Animalia), and replicate intracellularly in both. Phytoplasmas release effectors into host cells of plants and insects to target host molecules, and in plants these effectors unload from the phloem to access distal tissues and alter basic developmental processes. The effectors provide phytoplasmas with a fitness advantage by modulating their plant and insect hosts. We expect that further research on the functional characterization of phytoplasma effectors will generate new knowledge that is relevant to fundamental aspects of plant sciences and entomology, and for agriculture by improving yields of crops affected by phytoplasma diseases.
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http://dx.doi.org/10.1146/annurev-phyto-072910-095323DOI Listing
August 2013

The cytosolic protein response as a subcomponent of the wider heat shock response in Arabidopsis.

Plant Cell 2009 Feb 24;21(2):642-54. Epub 2009 Feb 24.

John Ines Centre, Colney, Norwich NR4 7UH, United Kingdom.

In common with a range of environmental and biological stresses, heat shock results in the accumulation of misfolded proteins and a collection of downstream consequences for cellular homeostasis and growth. Within this complex array of responses, the sensing of and responses to misfolded proteins in specific subcellular compartments involves specific chaperones, transcriptional regulators, and expression profiles. Using biological (ectopic protein expression and virus infection) and chemical triggers for misfolded protein accumulation, we have profiled the transcriptional features of the response to misfolded protein accumulation in the cytosol (i.e., the cytoplasmic protein response [CPR]) and identified the effects as a subcomponent of the wider effects induced by heat shock. The CPR in Arabidopsis thaliana is associated with the heat shock promoter element and the involvement of specific heat shock factors (HSFs), notably HSFA2, which appears to be regulated by alternative splicing and non-sense-mediated decay. Characterization of Arabidopsis HSFA2 knockout and overexpression lines showed that HSFA2 is one of the regulatory components of the CPR.
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http://dx.doi.org/10.1105/tpc.108.062596DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2660624PMC
February 2009

Two type III effector genes of Xanthomonas oryzae pv. oryzae control the induction of the host genes OsTFIIAgamma1 and OsTFX1 during bacterial blight of rice.

Proc Natl Acad Sci U S A 2007 Jun 11;104(25):10720-5. Epub 2007 Jun 11.

Department of Plant Pathology, Kansas State University, Manhattan, KS 66506, USA.

Xanthomonas oryzae pv. oryzae strain PXO99(A) induces the expression of the host gene Os8N3, which results in increased host susceptibility to bacterial blight of rice. Here, we show that PXO99(A) affects the expression of two additional genes in a type III secretion system-dependent manner, one encoding a bZIP transcription factor (OsTFX1) and the other the small subunit of the transcription factor IIA located on chromosome 1 (OsTFIIAgamma1). Induction of OsTFX1 and OsTFIIAgamma1 depended on the type III effector genes pthXo6 and pthXo7, respectively, both encoding two previously undescribed members of the transcription activator-like (TAL) effector family. pthXo7 is strain-specific and may reflect adaptation to the resistance mediated by xa5, an allele of OsTFIIAgamma5 encoding a second form of the TFIIA small subunit on chromosome 5 of rice. The loss of pthXo6 resulted in reduced pathogen virulence, and ectopic expression of OsTFX1 abrogated the requirement for pthXo6 for full virulence. X. oryzae pv. oryzae therefore modulates the expression of multiple host genes using multiple TAL effectors from a single strain, and evidence supports the hypothesis that expression of the associated host genes contributes to host susceptibility to disease.
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http://dx.doi.org/10.1073/pnas.0701742104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1965579PMC
June 2007

Os8N3 is a host disease-susceptibility gene for bacterial blight of rice.

Proc Natl Acad Sci U S A 2006 Jul 23;103(27):10503-10508. Epub 2006 Jun 23.

Department of Plant Pathology, Kansas State University, Manhattan, KS 66506

Many bacterial diseases of plants depend on the interaction of type III effector genes of the pathogen and disease-susceptibility genes of the host. The host susceptibility genes are largely unknown. Here, we show that expression of the rice gene Os8N3, a member of the MtN3 gene family from plants and animals, is elevated upon infection by Xanthomonas oryzae pv. oryzae strain PXO99(A) and depends on the type III effector gene pthXo1. Os8N3 resides near xa13, and PXO99(A) failed to induce Os8N3 in rice lines with xa13. Silencing of Os8N3 by inhibitory RNA produced plants that were resistant to infection by strain PXO99(A) yet remained susceptible to other strains of the pathogen. The effector gene avrXa7 from strain PXO86 enabled PXO99(A) compatibility on either xa13- or Os8N3-silenced plants. The findings indicate that Os8N3 is a host susceptibility gene for bacterial blight targeted by the type III effector PthXo1. The results support the hypothesis that X. oryzae pv. oryzae commandeers the regulation of otherwise developmentally regulated host genes to induce a state of disease susceptibility. Furthermore, the results support a model in which the pathogen induces disease susceptibility in a gene-for-gene manner.
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http://dx.doi.org/10.1073/pnas.0604088103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1502487PMC
July 2006

Inhibition of resistance gene-mediated defense in rice by Xanthomonas oryzae pv. oryzicola.

Mol Plant Microbe Interact 2006 Mar;19(3):240-9

Department of Plant Pathology, Iowa State University, Ames, IA 50014, USA.

Xanthomonas oryzae pv. oryzae and the closely related X. oryzae pv. oryzicola cause bacterial blight and bacterial leaf streak of rice, respectively. Although many rice resistance (R) genes and some corresponding avirulence (avr) genes have been characterized for bacterial blight, no endogenous avr/R gene interactions have been identified for leaf streak. Genes avrXa7 and avrXa10 from X. oryzae pv. oryzae failed to elicit the plant defense-associated hypersensitive reaction (HR) and failed to prevent development of leaf streak in rice cultivars with the corresponding R genes after introduction into X. oryzae pv. oryzicola despite the ability of this pathovar to deliver an AvrXa10:Cya fusion protein into rice cells. Furthermore, coinoculation of X. oryzae pv. oryzicola inhibited the HR of rice cultivar IRBB10 to X. oryzae pv. oryzae carrying avrXa10. Inhibition was quantitative and dependent on the type III secretion system of X. oryzae pv. oryzicola. The results suggest that one or more X. oryzae pv. oryzicola type III effectors interfere with avr/R gene-mediated recognition or signaling and subsequent defense response in the host. Inhibition of R gene-mediated defense by X. oryzae pv. oryzicola may explain, in part, the apparent lack of major gene resistance to leaf streak.
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http://dx.doi.org/10.1094/MPMI-19-0240DOI Listing
March 2006

Characterization of the hrpF pathogenicity peninsula of Xanthomonas oryzae pv. oryzae.

Mol Plant Microbe Interact 2005 Jun;18(6):546-54

Department of Plant Pathology, Throckmorton Hall, Kansas State University, Manhattan 66506-5502, USA.

The hrp gene cluster of Xanthomonas spp. contains genes for the assembly and function of a type III secretion system (TTSS). The hrpF genes reside in a region between hpaB and the right end of the hrp cluster. The region of the hrpF gene of Xanthomonas oryzae pv. oryzae is bounded by two IS elements and also contains a homolog of hpaF of X. campestris pv. vesicatoria and two newly identified genes, hpa3 and hpa4. A comparison of the hrp gene clusters of different species of Xanthomonas revealed that the hrpF region is a constant yet more variable peninsula of the hrp pathogenicity island. Mutations in hpaF, hpa3, and hpa4 had no effect on virulence, whereas hrpF mutants were severely reduced in virulence on susceptible rice cultivars. The hrpF genes from X. campestris pv. vesicatoria, X. campestris pv. campestris, and X. axonopodis pv. citri each were capable of restoring virulence to the hrpF mutant of X. oryzae pv. oryzae. Correspondingly, none of the Xanthomonas pathovars with hrpF from X. oryzae pv. oryzae elicited a hypersensitive reaction in their respective hosts. Therefore, no evidence was found for hrpF as a host-specialization factor. In contrast to the loss of Bs3-dependent reactions by hrpF mutants of X. campestris pv. vesicatoria, hrpF mutants of X. oryzae pv. oryzae with either avrXa10 or avrXa7 elicited hypersensitive reactions in rice cultivars with the corresponding R genes. A double hrpFxoo-hpa1 mutant also elicited an Xa10-dependent resistance reaction. Thus, loss of hrpF, hpal, or both may reduce delivery or effectiveness of type III effectors. However, the mutations did not completely prevent the delivery of effectors from X. oryzae pv. oryzae into the host cells.
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http://dx.doi.org/10.1094/MPMI-18-0546DOI Listing
June 2005

Avoidance of host recognition by alterations in the repetitive and C-terminal regions of AvrXa7, a type III effector of Xanthomonas oryzae pv. oryzae.

Mol Plant Microbe Interact 2005 Feb;18(2):142-9

Department of Plant Pathology, Kansas State University, Manhattan, KS 66502, USA.

avrXa7 is a member of the avrBs3/pthA gene family. The gene is a critical type III effector in several strains of Xanthomonas oryzae pv. oryzae (virulence activity), and in the presence of the Xa7 host gene for resistance, controls the elicitation of resistance in rice (avirulence activity). The ability of strains containing avrXa7 to adapt to the presence of Xa7 in the host population is dependent, in part, on the genetic plasticity of avrXa7. The potential for the conversion of avrXa7 to a virulence effector without Xa7-dependent elicitor activity was examined. Internal reorganization of avrXa7 by artificially deleting a portion of the central repetitive region resulted in gene pthXo4, which retained virulence activity and lost Xa7-dependent avirulence activity. Similarly, spontaneous rearrangements between repetitive regions of avrXa7 during bacterial culture gave rise to gene pthXo5, which also had virulence activity without Xa7-dependent avirulence activity. pthXo5 appeared to be the result of recombination between avrXa7 and a related gene in the genome. Loss of avirulence activity and retention of virulence activity also resulted from replacement of a portion of the C-terminal coding region of avrXa7 with the corresponding sequence from avrBs3. The results demonstrated the potential for a critical virulence effector to lose avirulence activity while retaining effector function. The results also demonstrated that features of both repetitive and nonrepetitive C-terminal regions of AvrXa7 are involved in avirulence specificity.
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http://dx.doi.org/10.1094/MPMI-18-0142DOI Listing
February 2005

Directed evolution of xylanase J from alkaliphilic Bacillus sp. strain 41M-1: restore of alkaliphily of a mutant with an acidic pH optimum.

Nucleic Acids Res Suppl 2003 (3):315-6

Department of Bioengineering, Tokyo Institute of Technology, Midori-ku, Yokohama 226-8501, Japan.

Alkaliphilic Bacillus sp. strain 41M-1 produces an alkaliphilic xylanase (xylanase J). The newly constructed mutant E177Q deltaJC had an acidic pH optimum and showed almost no activity at pH 8.0. The alkaliphily of the enzyme was restored by directed evolution. The evolved mutants, Y176S/E177Q deltaJC and G32V/Y176D/E177Q deltaJC, retained about 30% and 43% activity of their maximal activities at pH 6.0, respectively.
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http://dx.doi.org/10.1093/nass/3.1.315DOI Listing
October 2003
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