Transcript Profiling in Host–Pathogen Interactions

Using genomic technologies, it is now possible to address research hypotheses in the context of entire developmental or biochemical pathways, gene networks, and chromosomal location of relevant genes and their inferred evolutionary history. Through a range of platforms, researchers can survey an entire transcriptome under a variety of experimental and field conditions. Interpretation of such data has led to new insights and revealed previously undescribed phenomena. In the area of plant-pathogen interactions, transcript profiling has provided unparalleled perception into the mechanisms underlying gene-for-gene resistance and basal defense, host vs nonhost resistance, biotrophy vs necrotrophy, and pathogenicity of vascular vs nonvascular pathogens, among many others. In this way, genomic technologies have facilitated a system-wide approach to unifying themes and unique features in the interactions of hosts and pathogens

Development of an Automatic Maize Seedling Phenotyping Platform Using 3D Vision and Industrial Robot Arm

Corp breeding plays an important role in modern agriculture, improving plant adaptability and increase yield. Optimizing genes is the key step to discover the beneficial genetic traits for crop production increasing. Associating genes and their functions needs a mountain of observation and measurement of the phenotypes, which is a dreary and fallible job for human beings. Automatic seedling phenotyping system aims at replacing the manual measurement, reduce the sampling time and increase the allowable work time. In this research, we developed an automated maize seedling phenotyping platform based on a ToF camera and an industrial robot arm. A ToF camera is mounted on the end-effector of the robot arm. The arm brings ToF camera to different viewpoints for acquiring 3D data. Camera-to-arm transformation matrix is calculated from hand-eye calibration, which is applied to transfer different viewpoint into arm base coordinate frame. Filters remove the background and noise in the merged seedling point clouds. 3D-to-2D projection and x-axis pixels density distribution method is used to segment the stem and leaves. Finally, separated leaves are fitted with 3D curves for parameter measurement. In testing experiment, 60 maize plants at early growth stage (V2~V5) were sampled by this platform

A sugarcane mosaic virus vector for gene expression in maize

Zea mays L. ssp. mays (maize) is an important crop plant as well as model system for genetics and plant biology. The ability to select among different virus‐based platforms for transient gene silencing or protein expression experiments is expected to facilitate studies of gene function in maize and complement experiments with stable transgenes. Here, we describe the development of a sugarcane mosaic virus (SCMV) vector for the purpose of protein expression in maize. An infectious SCMV cDNA clone was constructed, and heterologous genetic elements were placed between the protein 1 (P1) and helper component‐proteinase (HC‐Pro) cistrons in the SCMV genome. Recombinant SCMV clones engineered to express green fluorescent protein (GFP), β‐glucuronidase (GUS), or bialaphos resistance (BAR) protein were introduced into sweet corn (Golden × Bantam) plants. Documentation of developmental time courses spanning maize growth from seedling to tasseling showed that the SCMV genome tolerates insertion of foreign sequences of at least 1,809 nucleotides at the P1/HC‐Pro junction. Analysis of insert stability showed that the integrity of GFP and BAR coding sequences was maintained longer than that of the much larger GUS coding sequence. The SCMV isolate from which the expression vector is derived is able to infect several important maize inbred lines, suggesting that this SCMV vector has potential to be a valuable tool for gene functional analysis in a broad range of experimentally important maize genotypes

A viral protease relocalizes in the presence of the vector to promote vector performance

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

A Foxtail mosaic virus Vector for Virus-Induced Gene Silencing in Maize

Plant viruses have been widely used as vectors for foreign gene expression and virus-induced gene silencing (VIGS). A limited number of viruses have been developed into viral vectors for the purposes of gene expression or VIGS in monocotyledonous plants, and among these, the tripartite viruses Brome mosaic virus and Cucumber mosaic virus have been shown to induce VIGS in maize (Zea mays). We describe here a new DNA-based VIGS system derived from Foxtail mosaic virus (FoMV), a monopartite virus that is able to establish systemic infection and silencing of endogenous maize genes homologous to gene fragments inserted into the FoMV genome. To demonstrate VIGS applications of this FoMV vector system, four genes, phytoene desaturase (functions in carotenoid biosynthesis), lesion mimic22 (encodes a key enzyme of the porphyrin pathway), iojap (functions in plastid development), and brown midrib3 (caffeic acid O-methyltransferase), were silenced and characterized in the sweet corn line Golden × Bantam. Furthermore, we demonstrate that the FoMV infectious clone establishes systemic infection in maize inbred lines, sorghum (Sorghum bicolor), and green foxtail (Setaria viridis), indicating the potential wide applications of this viral vector system for functional genomics studies in maize and other monocots

Alternative Hosts for Soybean vein necrosis virus and Feeding Preferences of Its Vector Soybean Thrips

Soybean vein necrosis virus (SVNV), a tospovirus and one of the most widespread soybean viruses in North America, is primarily transmitted by soybean thrips (Neohydatothrips variabilis). Although soybean is not considered the primary plant host for SVNV, there is a dearth of knowledge about alternative host plants for SVNV. We therefore investigated whether commonly present specialty and cover crops in Iowa can serve as alternative hosts for SVNV. Seventeen cover crops and seven specialty crops were tested using mechanical and thrips inoculations. Clear symptoms of SVNV and systemic infection in buckwheat and clear local infection with possible systemic infection on melon were shown. Additionally, we compared soybean thrips feeding on 18 cover crops and determined that they preferred alfalfa, buckwheat, crimson clover, and red clover. Our results suggested that alternative host crops may harbor SVNV and be a possible source of inoculum for soybean

Differential accumulation of host mRNAs on polyribosomes during obligate pathogen-plant interactions

Plant pathogens elicit dramatic changes in the expression of host genes during both compatible and incompatible interactions. Gene expression profiling studies of plant-pathogen interactions have only considered messenger RNAs (mRNAs) present in total RNA, which contains subpopulations of actively translated mRNAs associated with polyribosomes (polysomes) and non-translated mRNAs that are not associated with polysomes. The goal of this study was to enhance previous gene expression analyses by identifying host mRNAs that become differentially associated with polysomes following pathogen inoculation. Total and polysomal RNA were extracted from barley (Hordeum vulgare) plants at 32 h after inoculation withBlumeria graminis f. sp. hordei, and Arabidopsis thaliana plants at 10 days after inoculation withTurnip mosaic virus. Gene expression profiles were obtained for each pathosystem, which represent diverse plant host-obligate pathogen interactions. Using this approach, host mRNAs were identified that were differentially associated with polysomes in response to pathogen treatment. Approximately 18% and 26% of mRNAs represented by probe sets on the Affymetrix Barley1 and Arabidopsis ATH1 GeneChips, respectively, differentially accumulated in the two populations in one or more combinations of treatment and genotype. Gene ontology analysis of mRNAs sharing the same pattern of accumulation in total and polysomal RNA identified gene sets that contained a significant number of functionally related annotations, suggesting both transcript accumulation and recruitment to polyribosomes are coordinately regulated in these systems

Novel noninvasive in situ probe of protein structure and dynamics

7-Amtryptophan is an ideal noninvasive in situ probe of protein structure and dynamics and provides an alternative to the use of ,tryptophan. 7-Azatryptophan affords a single-exponential fluorescence decay in aqueous solution, unlike tryptophan. Its absorption and fluorescence spectra are distinguishable from those of tryptophan. Its fluorescence spectrum and lifetime are sensitive to the environment. It can be used in peptide synthesis, and it can be incorporated into bacterial protein. These facts render 7-azatryptophan a unique probe that has the potential for widespread use

Differential Requirement of the Ribosomal Protein S6 and Ribosomal Protein S6 Kinase for Plant-Virus Accumulation and Interaction of S6 Kinase with Potyviral VPg

Ribosomal protein S6 (RPS6) is an indispensable plant protein regulated, in part, by ribosomal protein S6 kinase (S6K) which, in turn, is a key regulator of plant responses to stresses and developmental cues. Increased expression of RPS6 was detected in Nicotiana benthamiana during infection by diverse plant viruses. Silencing of the RPS6and S6K genes in N. benthamiana affected accumulation of Cucumber mosaic virus, Turnip mosaic virus (TuMV), and Potato virus A (PVA) in contrast to Turnip crinkle virus and Tobacco mosaic virus. In addition, the viral genome-linked protein (VPg) of TuMV and PVA interacted with S6K in plant cells, as detected by bimolecular fluorescence complementation assay. The VPg–S6K interaction was detected in cytoplasm, nucleus, and nucleolus, whereas the green fluorescent protein-tagged S6K alone showed cytoplasmic localization only. These results demonstrate that the requirement for RPS6 and S6K differs for diverse plant viruses with different translation initiation strategies and suggest that potyviral VPg–S6K interaction may affect S6K functions in both the cytoplasm and the nucleus

Rpp1 encodes a ULP1-NBS-LRR protein that controls immunity to Phakopsora pachyrhizi in soybean

Phakopsora pachyrhizi is the causal agent of Asian soybean rust. Susceptible soybean plants infected by virulent isolates of P. pachyrhizi are characterized by tan-colored lesions and erumpent uredinia on the leaf surface. Germplasm screening and genetic analyses have led to the identification of seven loci, Rpp1 – Rpp7, that provide varying degrees of resistance to P. pachyrhizi (Rpp). Two genes, Rpp1 and Rpp1b, map to the same region on soybean chromosome 18. Rpp1 is unique among the Rpp genes in that it confers an immune response (IR) to avirulent P. pachyrhizi isolates. The IR is characterized by a lack of visible symptoms, whereas resistance provided by Rpp1b – Rpp7 results in red-brown foliar lesions. Rpp1 maps to a region spanning approximately 150 Kb on chromosome 18 between markers Sct_187 and Sat_064 in L85-2378 (Rpp1), an isoline developed from Williams 82 and PI 200492 (Rpp1). To identify Rpp1, we constructed a bacterial artificial chromosome (BAC) library from soybean accession PI 200492. Sequencing of the Rpp1 locus identified three homologous nucleotide binding site-leucine rich repeat (NBS-LRR) candidate resistance genes between Sct_187 and Sat_064. Each candidate gene is also predicted to encode an N-terminal ubiquitin-like protease 1 (ULP1) domain. Co-silencing of the Rpp1 candidates abrogated the immune response in the Rpp1 resistant soybean accession PI 200492, indicating that Rpp1 is a ULP1-NBS-LRR protein and plays a key role in the IR