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1.
Mol Cell Proteomics ; 16(4 suppl 1): S230-S243, 2017 04.
Artigo em Inglês | MEDLINE | ID: mdl-27932519

RESUMO

The green peach aphid, Myzus persicae, is a vector of the Potato leafroll virus (PLRV, Luteoviridae), transmitted exclusively by aphids in a circulative manner. PLRV transmission efficiency was significantly reduced when a clonal lineage of M. persicae was reared on turnip as compared with the weed physalis, and this was a transient effect caused by a host-switch response. A trend of higher PLRV titer in physalis-reared aphids as compared with turnip-reared aphids was observed at 24 h and 72 h after virus acquisition. The major difference in the proteomes of these aphids was the up-regulation of predicted lysosomal enzymes, in particular the cysteine protease cathepsin B (cathB), in aphids reared on turnip. The aphid midgut is the site of PLRV acquisition, and cathB and PLRV localization were starkly different in midguts of the aphids reared on the two host plants. In viruliferous aphids that were reared on turnip, there was near complete colocalization of cathB and PLRV at the cell membranes, which was not observed in physalis-reared aphids. Chemical inhibition of cathB restored the ability of aphids reared on turnip to transmit PLRV in a dose-dependent manner, showing that the increased activity of cathB and other cysteine proteases at the cell membrane indirectly decreased virus transmission by aphids. Understanding how the host plant influences virus transmission by aphids is critical for growers to manage the spread of virus among field crops.


Assuntos
Afídeos/virologia , Brassica napus/parasitologia , Catepsina B/metabolismo , Luteoviridae/fisiologia , Physalis/parasitologia , Animais , Afídeos/enzimologia , Afídeos/fisiologia , Trato Gastrointestinal/enzimologia , Trato Gastrointestinal/virologia , Interações Hospedeiro-Parasita , Proteínas de Insetos/metabolismo , Insetos Vetores/enzimologia , Insetos Vetores/fisiologia , Insetos Vetores/virologia , Doenças das Plantas/virologia , Vírus de Plantas/fisiologia , Proteômica/métodos , Regulação para Cima , Carga Viral
2.
J Virol ; 90(4): 1973-87, 2016 02 15.
Artigo em Inglês | MEDLINE | ID: mdl-26656710

RESUMO

UNLABELLED: Demonstrating direct interactions between host and virus proteins during infection is a major goal and challenge for the field of virology. Most protein interactions are not binary or easily amenable to structural determination. Using infectious preparations of a polerovirus (Potato leafroll virus [PLRV]) and protein interaction reporter (PIR), a revolutionary technology that couples a mass spectrometric-cleavable chemical cross-linker with high-resolution mass spectrometry, we provide the first report of a host-pathogen protein interaction network that includes data-derived, topological features for every cross-linked site that was identified. We show that PLRV virions have hot spots of protein interaction and multifunctional surface topologies, revealing how these plant viruses maximize their use of binding interfaces. Modeling data, guided by cross-linking constraints, suggest asymmetric packing of the major capsid protein in the virion, which supports previous epitope mapping studies. Protein interaction topologies are conserved with other species in the Luteoviridae and with unrelated viruses in the Herpesviridae and Adenoviridae. Functional analysis of three PLRV-interacting host proteins in planta using a reverse-genetics approach revealed a complex, molecular tug-of-war between host and virus. Structural mimicry and diversifying selection-hallmarks of host-pathogen interactions-were identified within host and viral binding interfaces predicted by our models. These results illuminate the functional diversity of the PLRV-host protein interaction network and demonstrate the usefulness of PIR technology for precision mapping of functional host-pathogen protein interaction topologies. IMPORTANCE: The exterior shape of a plant virus and its interacting host and insect vector proteins determine whether a virus will be transmitted by an insect or infect a specific host. Gaining this information is difficult and requires years of experimentation. We used protein interaction reporter (PIR) technology to illustrate how viruses exploit host proteins during plant infection. PIR technology enabled our team to precisely describe the sites of functional virus-virus, virus-host, and host-host protein interactions using a mass spectrometry analysis that takes just a few hours. Applications of PIR technology in host-pathogen interactions will enable researchers studying recalcitrant pathogens, such as animal pathogens where host proteins are incorporated directly into the infectious agents, to investigate how proteins interact during infection and transmission as well as develop new tools for interdiction and therapy.


Assuntos
Interações Hospedeiro-Patógeno , Luteoviridae/fisiologia , Mapas de Interação de Proteínas , Proteômica/métodos , Proteínas de Plantas/metabolismo , Nicotiana , Proteínas Virais/metabolismo
3.
BMC Biol ; 14(1): 110, 2016 12 14.
Artigo em Inglês | MEDLINE | ID: mdl-27974049

RESUMO

BACKGROUND: The whitefly Bemisia tabaci (Hemiptera: Aleyrodidae) is among the 100 worst invasive species in the world. As one of the most important crop pests and virus vectors, B. tabaci causes substantial crop losses and poses a serious threat to global food security. RESULTS: We report the 615-Mb high-quality genome sequence of B. tabaci Middle East-Asia Minor 1 (MEAM1), the first genome sequence in the Aleyrodidae family, which contains 15,664 protein-coding genes. The B. tabaci genome is highly divergent from other sequenced hemipteran genomes, sharing no detectable synteny. A number of known detoxification gene families, including cytochrome P450s and UDP-glucuronosyltransferases, are significantly expanded in B. tabaci. Other expanded gene families, including cathepsins, large clusters of tandemly duplicated B. tabaci-specific genes, and phosphatidylethanolamine-binding proteins (PEBPs), were found to be associated with virus acquisition and transmission and/or insecticide resistance, likely contributing to the global invasiveness and efficient virus transmission capacity of B. tabaci. The presence of 142 horizontally transferred genes from bacteria or fungi in the B. tabaci genome, including genes encoding hopanoid/sterol synthesis and xenobiotic detoxification enzymes that are not present in other insects, offers novel insights into the unique biological adaptations of this insect such as polyphagy and insecticide resistance. Interestingly, two adjacent bacterial pantothenate biosynthesis genes, panB and panC, have been co-transferred into B. tabaci and fused into a single gene that has acquired introns during its evolution. CONCLUSIONS: The B. tabaci genome contains numerous genetic novelties, including expansions in gene families associated with insecticide resistance, detoxification and virus transmission, as well as numerous horizontally transferred genes from bacteria and fungi. We believe these novelties likely have shaped B. tabaci as a highly invasive polyphagous crop pest and efficient vector of plant viruses. The genome serves as a reference for resolving the B. tabaci cryptic species complex, understanding fundamental biological novelties, and providing valuable genetic information to assist the development of novel strategies for controlling whiteflies and the viruses they transmit.


Assuntos
Genoma de Inseto/genética , Hemípteros/genética , Animais , Hemípteros/efeitos dos fármacos , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Resistência a Inseticidas/genética , Resistência a Inseticidas/fisiologia , Vírus de Plantas/patogenicidade
4.
J Proteome Res ; 15(12): 4601-4611, 2016 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-27762138

RESUMO

Phloem localization of plant viruses is advantageous for acquisition by sap-sucking vectors but hampers host-virus protein interaction studies. In this study, Potato leafroll virus (PLRV)-host protein complexes were isolated from systemically infected potato, a natural host of the virus. Comparing two different co-immunoprecipitation (co-IP) support matrices coupled to mass spectrometry (MS), we identified 44 potato proteins and one viral protein (P1) specifically associated with virus isolated from infected phloem. An additional 142 proteins interact in complex with virus at varying degrees of confidence. Greater than 80% of these proteins were previously found to form high confidence interactions with PLRV isolated from the model host Nicotiana benthamiana. Bioinformatics revealed that these proteins are enriched for functions related to plasmodesmata, organelle membrane transport, translation, and mRNA processing. Our results show that model system proteomics experiments are extremely valuable for understanding protein interactions regulating infection in recalcitrant pathogens such as phloem-limited viruses.


Assuntos
Floema/virologia , Mapeamento de Interação de Proteínas/métodos , Biologia Computacional , Interações Hospedeiro-Patógeno , Proteínas de Plantas/metabolismo , Vírus de Plantas/química , Ligação Proteica , Solanum tuberosum/química , Solanum tuberosum/virologia , Proteínas Virais/metabolismo
5.
Proteomics ; 15(12): 2098-112, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25787689

RESUMO

Potato leafroll virus (PLRV) produces a readthrough protein (RTP) via translational readthrough of the coat protein amber stop codon. The RTP functions as a structural component of the virion and as a nonincorporated protein in concert with numerous insect and plant proteins to regulate virus movement/transmission and tissue tropism. Affinity purification coupled to quantitative MS was used to generate protein interaction networks for a PLRV mutant that is unable to produce the read through domain (RTD) and compared to the known wild-type PLRV protein interaction network. By quantifying differences in the protein interaction networks, we identified four distinct classes of PLRV-plant interactions: those plant and nonstructural viral proteins interacting with assembled coat protein (category I); plant proteins in complex with both coat protein and RTD (category II); plant proteins in complex with the RTD (category III); and plant proteins that had higher affinity for virions lacking the RTD (category IV). Proteins identified as interacting with the RTD are potential candidates for regulating viral processes that are mediated by the RTP such as phloem retention and systemic movement and can potentially be useful targets for the development of strategies to prevent infection and/or viral transmission of Luteoviridae species that infect important crop species.


Assuntos
Luteoviridae/fisiologia , Folhas de Planta/metabolismo , Proteínas de Plantas/metabolismo , Mapas de Interação de Proteínas , Proteômica/métodos , Solanum tuberosum/metabolismo , Proteínas Estruturais Virais/metabolismo , Western Blotting , Imunoprecipitação , Espectrometria de Massas , Doenças das Plantas/virologia , Folhas de Planta/virologia , Solanum tuberosum/virologia
6.
Mol Plant Microbe Interact ; 28(4): 467-81, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25496593

RESUMO

Identification of host proteins interacting with the aphidborne Potato leafroll virus (PLRV) from the genus Polerovirus, family Luteoviridae, is a critical step toward understanding how PLRV and related viruses infect plants. However, the tight spatial distribution of PLRV to phloem tissues poses challenges. A polyclonal antibody raised against purified PLRV virions was used to coimmunoprecipitate virus-host protein complexes from Nicotiana benthamiana tissue inoculated with an infectious PLRV cDNA clone using Agrobacterium tumefaciens. A. tumefaciens-mediated delivery of PLRV enabled infection and production of assembled, insect-transmissible virus in most leaf cells, overcoming the dynamic range constraint posed by a systemically infected host. Isolated protein complexes were characterized using high-resolution mass spectrometry and consisted of host proteins interacting directly or indirectly with virions, as well as the nonincorporated readthrough protein (RTP) and three phosphorylated positional isomers of the RTP. A bioinformatics analysis using ClueGO and STRING showed that plant proteins in the PLRV protein interaction network regulate key biochemical processes, including carbon fixation, amino acid biosynthesis, ion transport, protein folding, and trafficking.


Assuntos
Luteoviridae/metabolismo , Proteínas de Plantas , Mapas de Interação de Proteínas/fisiologia , Proteínas Virais , Agrobacterium tumefaciens , Imunoprecipitação , Luteoviridae/química , Espectrometria de Massas , Proteínas de Plantas/análise , Proteínas de Plantas/química , Proteínas de Plantas/metabolismo , Nicotiana/química , Nicotiana/virologia , Proteínas Virais/análise , Proteínas Virais/química , Proteínas Virais/metabolismo , Vírion
7.
J Virol ; 88(10): 5652-60, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24600010

RESUMO

UNLABELLED: Numerous animal and plant viruses are transmitted by arthropod vectors in a persistent, circulative manner. Tomato yellow leaf curl virus (TYLCV) is transmitted by the sweet potato whitefly Bemisia tabaci. We report here that infection with Rickettsia spp., a facultative endosymbiont of whiteflies, altered TYLCV-B. tabaci interactions. A B. tabaci strain infected with Rickettsia acquired more TYLCV from infected plants, retained the virus longer, and exhibited nearly double the transmission efficiency compared to an uninfected B. tabaci strain with the same genetic background. Temporal and spatial antagonistic relationships were discovered between Rickettsia and TYLCV within the whitefly. In different time course experiments, the levels of virus and Rickettsia within the insect were inversely correlated. Fluorescence in situ hybridization analysis of Rickettsia-infected midguts provided evidence for niche exclusion between Rickettsia and TYLCV. In particular, high levels of the bacterium in the midgut resulted in higher virus concentrations in the filter chamber, a favored site for virus translocation along the transmission pathway, whereas low levels of Rickettsia in the midgut resulted in an even distribution of the virus. Taken together, these results indicate that Rickettsia, by infecting the midgut, increases TYLCV transmission efficacy, adding further insights into the complex association between persistent plant viruses, their insect vectors, and microorganism tenants that reside within these insects. IMPORTANCE: Interest in bacterial endosymbionts in arthropods and many aspects of their host biology in agricultural and human health systems has been increasing. A recent and relevant studied example is the influence of Wolbachia on dengue virus transmission by mosquitoes. In parallel with our recently studied whitefly-Rickettsia-TYLCV system, other studies have shown that dengue virus levels in the mosquito vector are inversely correlated with bacterial load. Our work here presents evidence of unifying principles between vectors of plant and animal viruses in a role for endosymbionts in manipulating vector biology and pathogen transmission. Our results demonstrate the influence of an interesting and prominent bacterial endosymbiont in Bemisia tabaci in TYLCV transmission, a worldwide disease infecting tomatoes. Besides its agricultural importance, this system provides interesting insights into Bemisia interaction with these newly discovered endosymbionts.


Assuntos
Begomovirus/isolamento & purificação , Portador Sadio/virologia , Hemípteros/microbiologia , Hemípteros/virologia , Rickettsia/isolamento & purificação , Rickettsia/fisiologia , Simbiose , Animais , Trato Gastrointestinal/microbiologia , Trato Gastrointestinal/virologia , Interações Microbianas
8.
J Proteome Res ; 13(4): 2094-108, 2014 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-24588548

RESUMO

Biotypes of aphids and many other insect pests are defined based on the phenotypic response of host plants to the insect pest without considering their intrinsic characteristics and genotypes. Plant breeders have spent considerable effort developing aphid-resistant, small-grain varieties to limit insecticide control of the greenbug, Schizaphis graminum. However, new S. graminum biotypes frequently emerge that break resistance. Mechanisms of virulence on the aphid side of the plant-insect interaction are not well understood. S. graminum biotype H is highly virulent on most small grain varieties. This characteristic makes biotype H ideal for comparative proteomics to investigate the basis of biotype virulence in aphids. In this study, we used comparative proteomics to identify protein expression differences associated with virulence. Aphid proteins involved in the tricarboxylic acid cycle, immune system, cell division, and antiapoptosis pathways were found to be up-regulated in biotype H relative to other biotypes. Proteins from the bacterial endosymbiont of aphids were also differentially expressed in biotype H. Guided by the proteome results, we tested whether biotype H had a fitness advantage compared with other S. graminum biotypes and found that biotype H had a higher reproductive fitness as compared with two other biotypes on a range of different wheat germplasms. Finally, we tested whether aphid genetics can be used to further dissect the genetic mechanisms of biotype virulence in aphids. The genetic data showed that sexual reproduction is a source of biotypic variation observed in S. graminum.


Assuntos
Afídeos/fisiologia , Afídeos/patogenicidade , Proteínas de Insetos/análise , Proteoma/análise , Proteoma/fisiologia , Animais , Afídeos/genética , Afídeos/microbiologia , Buchnera , Interações Hospedeiro-Parasita , Proteínas de Insetos/classificação , Proteínas de Insetos/genética , Proteínas de Insetos/metabolismo , Poaceae/parasitologia , Proteoma/genética , Proteoma/metabolismo , Simbiose
9.
J Gen Virol ; 95(Pt 10): 2321-2327, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-24939649

RESUMO

Virions of the RPV strain of Cereal yellow dwarf virus-RPV were purified from infected oat tissue and analysed by MS. Two conserved residues, K147 and K181, in the virus coat protein, were confidently identified to contain epsilon-N-acetyl groups. While no functional data are available for K147, K181 lies within an interfacial region critical for virion assembly and stability. The signature immonium ion at m/z 126.0919 demonstrated the presence of N-acetyllysine, and the sequence fragment ions enabled an unambiguous assignment of the epsilon-N-acetyl modification on K181. We hypothesize that selection favours acetylation of K181 in a fraction of coat protein monomers to stabilize the capsid by promoting intermonomer salt bridge formation.


Assuntos
Proteínas do Capsídeo/metabolismo , Luteoviridae/fisiologia , Lisina/metabolismo , Processamento de Proteína Pós-Traducional , Acetilação , Luteoviridae/química , Espectrometria de Massas
10.
J Proteome Res ; 11(5): 2968-81, 2012 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-22390342

RESUMO

Protein interactions are critical determinants of insect transmission for viruses in the family Luteoviridae. Two luteovirid structural proteins, the capsid protein (CP) and the readthrough protein (RTP), contain multiple functional domains that regulate virus transmission. There is no structural information available for these economically important viruses. We used Protein Interaction Reporter (PIR) technology, a strategy that uses chemical cross-linking and high resolution mass spectrometry, to discover topological features of the Potato leafroll virus (PLRV) CP and RTP that are required for the diverse biological functions of PLRV virions. Four cross-linked sites were repeatedly detected, one linking CP monomers, two within the RTP, and one linking the RTP and CP. Virus mutants with triple amino acid deletions immediately adjacent to or encompassing the cross-linked sites were defective in virion stability, RTP incorporation into the capsid, and aphid transmission. Plants infected with a new, infectious PLRV mutant lacking 26 amino acids encompassing a cross-linked site in the RTP exhibited a delay in the appearance of systemic infection symptoms. PIR technology provided the first structural insights into luteoviruses which are crucially lacking and are involved in vector-virus and plant-virus interactions. These are the first cross-linking measurements on any infectious, insect-transmitted virus.


Assuntos
Afídeos/virologia , Interações Hospedeiro-Patógeno , Insetos Vetores/virologia , Luteoviridae/patogenicidade , Mapeamento de Interação de Proteínas/métodos , Solanum tuberosum/virologia , Agrobacterium tumefaciens/genética , Agrobacterium tumefaciens/metabolismo , Sequência de Aminoácidos , Animais , Afídeos/metabolismo , Sítios de Ligação , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/metabolismo , Comportamento Alimentar , Luteoviridae/isolamento & purificação , Luteoviridae/fisiologia , Espectrometria de Massas , Dados de Sequência Molecular , Doenças das Plantas/virologia , Plasmídeos/genética , Plasmídeos/metabolismo , Nicotiana/virologia , Montagem de Vírus
11.
Proc Natl Acad Sci U S A ; 106(9): 3615-20, 2009 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-19218459

RESUMO

Cell-to-cell transport in plants occurs through cytoplasmic channels called "plasmodesmata" and is regulated by developmental and environmental factors. Callose deposition modulates plasmodesmal transport in vivo, but little is known about the mechanisms that regulate this process. Here we report a genetic approach to identify mutants affecting plasmodesmal transport. We isolated 5 mutants, named gfp arrested trafficking (gat), affected in GFP unloading from the phloem into the meristem. gat1 mutants were seedling lethal and carried lesions in an m-type thioredoxin that is expressed in non-green plastids of meristems and organ primordia. Callose and hydrogen peroxide accumulated in gat1 mutants, and WT plants subjected to oxidative conditions phenocopied the gat1 trafficking defects. Ectopic expression of GAT1 in mature leaves increased plasmodesmal permeability and led to a delay in senescence and flowering time. We propose a role for the GAT1 thioredoxin in the redox regulation of callose deposition and symplastic permeability that is essential for meristem maintenance in Arabidopsis.


Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Meristema/crescimento & desenvolvimento , Meristema/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Meristema/genética , Fenótipo , Plantas Geneticamente Modificadas , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Tiorredoxinas/genética
12.
Proteomics ; 11(12): 2440-58, 2011 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-21648087

RESUMO

Yellow dwarf viruses cause the most economically important virus diseases of cereal crops worldwide and are vectored by aphids. The identification of vector proteins mediating virus transmission is critical to develop sustainable virus management practices and to understand viral strategies for circulative movement in all insect vectors. Previously, we applied 2-D DIGE to an aphid filial generation 2 population to identify proteins correlated with the transmission phenotype that were stably inherited and expressed in the absence of the virus. In the present study, we examined the expression of the DIGE candidates in previously unstudied, field-collected aphid populations. We hypothesized that the expression of proteins involved in virus transmission could be clinically validated in unrelated, virus transmission-competent, field-collected aphid populations. All putative biomarkers were expressed in the field-collected biotypes, and the expression of nine of these aligned with the virus transmission-competent phenotype. The strong conservation of the expression of the biomarkers in multiple field-collected populations facilitates new and testable hypotheses concerning the genetics and biochemistry of virus transmission. Integration of these biomarkers into current aphid-scouting methodologies will enable rational strategies for vector control aimed at judicious use and development of precision pest control methods that reduce plant virus infection.


Assuntos
Afídeos , Biomarcadores/análise , Eletroforese em Gel Bidimensional/métodos , Proteínas de Insetos/genética , Insetos Vetores/genética , Doenças das Plantas/virologia , Vírus de Plantas/genética , Sequência de Aminoácidos , Animais , Afídeos/classificação , Afídeos/genética , Afídeos/virologia , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Buchnera/química , Estudos de Associação Genética , Variação Genética , Hordeum/virologia , Proteínas de Insetos/metabolismo , Insetos Vetores/metabolismo , Espectrometria de Massas , Dados de Sequência Molecular , Doenças das Plantas/genética , Vírus de Plantas/metabolismo , Proteômica/métodos , Simbiose
13.
Curr Opin Cell Biol ; 16(5): 500-6, 2004 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-15363799

RESUMO

Intercellular transport via plasmodesmata controls cell fate decisions in plants, and is of fundamental importance in viral movement, disease resistance, and the spread of RNAi signals. Although plasmodesmata appear to be unique to plant cells, they may have structural and functional similarities to the newly discovered tunneling nanotubes that connect animal cells. Recently, proteins that localize to plasmodesmata have been identified, and a microtubule-associated protein was found to negatively regulate the trafficking of viral movement proteins. Other advances have delivered new insights into the function and molecular nature of plasmodesmata and have shown that protein trafficking through plasmodesmata is developmentally regulated, opening up the possibility that the genetic control of plasmodesmal function will soon be understood.


Assuntos
Diferenciação Celular/fisiologia , Fenômenos Fisiológicos Vegetais , Plasmodesmos/fisiologia , Transdução de Sinais/fisiologia , Endocitose/fisiologia , Epigênese Genética/fisiologia , Exocitose/fisiologia , Proteínas Associadas aos Microtúbulos/fisiologia , Plantas/virologia , Transporte Proteico/fisiologia
15.
Sci Rep ; 6: 33418, 2016 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-27630042

RESUMO

Candidatus Liberibacter asiaticus (CLas) is a phloem-limited, gram-negative, fastidious bacterium that is associated with the development of citrus greening disease, also known as Huanglongbing (HLB). CLas is transmitted by the Asian citrus psyllid (ACP) Diaphorina citri, in a circulative manner. Two major barriers to transmission within the insect are the midgut and the salivary glands. We performed a thorough microscopic analysis within the insect midgut following exposure to CLas-infected citrus trees. We observed changes in nuclear architecture, including pyknosis and karyorrhexis as well as changes to the actin cytoskeleton in CLas-exposed midgut cells. Further analyses showed that the changes are likely due to the activation of programmed cell death as assessed by Annexin V staining and DNA fragmentation assays. These results suggest that exposure to CLas-infected trees induces apoptotic responses in the psyllid midgut that should be further investigated. Understanding the adaptive significance of the apoptotic response has the potential to create new approaches for controlling HLB.


Assuntos
Citrus/parasitologia , Sistema Digestório/microbiologia , Sistema Digestório/patologia , Hemípteros/citologia , Hemípteros/microbiologia , Rhizobiaceae/fisiologia , Citoesqueleto de Actina/metabolismo , Animais , Anexina A5/metabolismo , Proteínas da Membrana Bacteriana Externa/imunologia , Morte Celular , Núcleo Celular/metabolismo , DNA/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Hibridização in Situ Fluorescente , Rhizobiaceae/imunologia
16.
PLoS One ; 11(12): e0167547, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27997545

RESUMO

Chemical cross-linking mass spectrometry (XL-MS) provides protein structural information by identifying covalently linked proximal amino acid residues on protein surfaces. The information gained by this technique is complementary to other structural biology methods such as x-ray crystallography, NMR and cryo-electron microscopy[1]. The extension of traditional quantitative proteomics methods with chemical cross-linking can provide information on the structural dynamics of protein structures and protein complexes. The identification and quantitation of cross-linked peptides remains challenging for the general community, requiring specialized expertise ultimately limiting more widespread adoption of the technique. We describe a general method for targeted quantitative mass spectrometric analysis of cross-linked peptide pairs. We report the adaptation of the widely used, open source software package Skyline, for the analysis of quantitative XL-MS data as a means for data analysis and sharing of methods. We demonstrate the utility and robustness of the method with a cross-laboratory study and present data that is supported by and validates previously published data on quantified cross-linked peptide pairs. This advance provides an easy to use resource so that any lab with access to a LC-MS system capable of performing targeted quantitative analysis can quickly and accurately measure dynamic changes in protein structure and protein interactions.


Assuntos
Reagentes de Ligações Cruzadas/química , Espectrometria de Massas/métodos , Proteômica/métodos , Soroalbumina Bovina/análise , Software , Animais , Bovinos , Células HeLa , Humanos , Estrutura Quaternária de Proteína , Soroalbumina Bovina/química
17.
Curr Opin Insect Sci ; 8: 69-78, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32846684

RESUMO

During the process of circulative plant virus transmission by insect vectors, viruses interact with different insect vector tissues prior to transmission to a new host plant. An area of intense debate in the field is whether bacterial symbionts of insect vectors are involved in the virus transmission process. We critically review the literature in this area and present a simple model that can be used to quantitatively settle the debate. The simple model determines whether the symbiont is involved in virus transmission and determines what fraction of the pathogen transmission phenotype is contributed by the symbiont. The model is general and can be applied to any vector-pathogen-symbiont interactions.

18.
Curr Opin Virol ; 15: 1-8, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26196230

RESUMO

Begomoviruses comprise an emerging and economically important group of plant viruses exclusively transmitted by the sweetpotato whitefly Bemisia tabaci in many regions of the world. The past twenty years have witnessed significant progress in studying the molecular interactions between members of this virus group and B. tabaci. Mechanisms and proteins encoded by the insect vector and its bacterial symbionts, which have been shown to be important for virus transmission, have been identified and thoroughly studied. Despite the economic importance of this group of viruses and their impact on the global agriculture, progress in investigating the virus-vector interactions is moving slowly when compared with similar virus-vector systems in plants and animals. Major advances in this field and future perspectives will be discussed in this review.


Assuntos
Begomovirus/fisiologia , Hemípteros/virologia , Insetos Vetores/virologia , Agricultura , Animais , Bactérias/virologia , Interações Hospedeiro-Patógeno/fisiologia , Doenças das Plantas/virologia , Simbiose
19.
PLoS One ; 10(11): e0140826, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26580079

RESUMO

'Candidatus Liberibacter asiaticus' (CLas), the bacterial pathogen associated with citrus greening disease, is transmitted by Diaphorina citri, the Asian citrus psyllid. Interactions among D. citri and its microbial endosymbionts, including 'Candidatus Profftella armatura', are likely to impact transmission of CLas. We used quantitative mass spectrometry to compare the proteomes of CLas(+) and CLas(-) populations of D. citri, and found that proteins involved in polyketide biosynthesis by the endosymbiont Profftella were up-regulated in CLas(+) insects. Mass spectrometry analysis of the Profftella polyketide diaphorin in D. citri metabolite extracts revealed the presence of a novel diaphorin-related polyketide and the ratio of these two polyketides was changed in CLas(+) insects. Insect proteins differentially expressed between CLas(+) and CLas(-) D. citri included defense and immunity proteins, proteins involved in energy storage and utilization, and proteins involved in endocytosis, cellular adhesion, and cytoskeletal remodeling which are associated with microbial invasion of host cells. Insight into the metabolic interdependence between the insect vector, its endosymbionts, and the citrus greening pathogen reveals novel opportunities for control of this disease, which is currently having a devastating impact on citrus production worldwide.


Assuntos
Proteínas de Bactérias/genética , Citrus/microbiologia , Hemípteros/microbiologia , Proteínas de Insetos/genética , Policetídeos/metabolismo , Proteoma/genética , Animais , Proteínas de Bactérias/metabolismo , Regulação da Expressão Gênica , Hemípteros/genética , Hemípteros/imunologia , Proteínas de Insetos/metabolismo , Redes e Vias Metabólicas , Anotação de Sequência Molecular , Doenças das Plantas/microbiologia , Proteoma/metabolismo , Rhizobiaceae/fisiologia , Simbiose
20.
Adv Virus Res ; 89: 141-99, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24751196

RESUMO

Species of plant viruses within the Luteoviridae, Geminiviridae, and Nanoviridae are transmitted by phloem-feeding insects in a circulative, nonpropagative manner. The precise route of virus movement through the vector can differ across and within virus families, but these viruses all share many biological, biochemical, and ecological features. All share temporal and spatial constraints with respect to transmission efficiency. The viruses also induce physiological changes in their plant hosts resulting in behavioral changes in the insects that optimize the transmission of virus to new hosts. Virus proteins interact with insect, endosymbiont, and plant proteins to orchestrate, directly and indirectly, virus movement in insects and plants to facilitate transmission. Knowledge of these complex interactions allows for the development of new tools to reduce or prevent transmission, to quickly identify important vector populations, and to improve the management of these economically important viruses affecting agricultural and natural plant populations.


Assuntos
Interações Hospedeiro-Parasita , Insetos Vetores/virologia , Doenças das Plantas/virologia , Plantas/virologia , Animais , Controle de Insetos , Doenças das Plantas/prevenção & controle
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