RNA-seq on tomato infected with Xanthomonas euvesicatoria
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ABSTRACT: Transcription activator-like effectors (TALEs) from Xanthomonas activate host susceptibility genes to promote disease. We report here the tomato transcriptome changes after inoculation with Xanthomonas strains delivering Brg11*, a TALE-like effector from Ralstonia solanacearum or delivering the Brg11-derivative lacking the DNA binding domain as a control. After diffenential gene expression analysis we identified 117 genes were up-regulated and 8 genes were down-regulated by Brg11. This provides us a candidate gene list facilitating identification of Brg11 direct target genes in tomato.
Project description:Xanthomonas spp. employ transcription activator-like effectors (TALEs) to promote pathogenicity by activating host susceptibility (S) genes. Cotton GhSWEET10 is an S gene targeted by a TALE in an early isolate of Xanthomonas citri pv. malvacearum (Xcm), but not by recent field Xcm isolates. To understand the pathogenicity shift in Xcm and its adaptation to cotton, we assembled the whole genome and the TALE repertoire of three recent Xcm Texas field isolates. A newly evolved TALE, Tal7b, activated different GhSWEET genes, GhSWEET14a and GhSWEET14b. Simultaneous activation of GhSWEET14a and GhSWEET14b resulted in pronounced water-soaked lesions. Transcriptome profiling coupled with TALE-binding element prediction identified a pectin lyase as an additional Tal7b target, quantitatively contributing to Xcm virulence alongside GhSWEET14a/b. CRISPR-Cas9-based gene editing supported the function of GhSWEETs as S genes in cotton bacterial blight and the promise of disrupting the TALE-binding site in these genes to control the disease. Collectively, our findings elucidate the rapid evolution of TALEs in Xanthomonas field isolates and highlight the virulence mechanism wherein TALEs induce multiple S genes simultaneously to promote pathogenicity.
Project description:A particularly promising approach for engineering more precise TALE proteins is to better understand non-canonical RVDs. The RVD repertoire employed by Xanthomonas extends well beyond just the four canonical RVD types, and natural TALE repeat arrays often utilize serial combinations of both canonical and non-canonical RVDs in a manner which is not fully understood. Attempts to characterize these non-canonical RVDs have utilized various approaches, in terms of both TALE design and experimental method. Some studies have considered just TALEs of particular biological relevance, others have looked at large numbers of different RVDs within a fixed repeat-array-context, and still others have designed custom TALEs to address hypotheses about optimal RVD positioning within the repeat array. Methods for inferring the DNA binding activity of these TALEs have included sequencing of cleaved DNA fragments from TALE-nuclease fusions; ELISA assays for TALE-oligonucleotide binding; reporter assays for TALEs’ transcriptional-activating effects; and in silico modeling. In this study we employ a novel “wholesale swap-out” approach to characterize 3 non-canonical RVDs for thymine and 4 non-canonical RVDs for guanine.
Project description:Investigation of whole genome gene expression level changes in the bacterial wilt pathogen Ralstonia solanacearum, strain GMI1000 at 20°C and 28°C in culture and in planta. The tropical strain GMI1000 cannot wilt tomato plants at 20°C although it can cause full-blown disease at 28°C. A 16 array study using total RNA recovered from the following: 8 separate cultures of Ralstonia solanacearum strain GMI1000 grown in rich medium-CPG (4 grown at 20°C and 4 grown at 28°C) 8 separate in planta samples of Ralstonia solanacearum strain GMI1000 harvested from diseased tomato plants cv. BonnyBest (4 recovered from plants grown at 20°C and 4 from plants grown at 28°C) Each array (4-plex format) measures the expression level of 5061 genes from Ralstonia solanacearum strain GMI1000 with 2 to 6, 40-70 mer probes per gene, with two-fold technical redundancy. The arrays also contain probes for intergenic regions with no technical replicates.
Project description:Investigation of whole genome gene expression level changes in the bacterial wilt pathogen Ralstonia solanacearum, strain UW551 at 20°C and 28°C in culture and in planta. The temperatel strain UW551 can wilt and cause full-blown disease on tomato plants at 28°C as well as at 20°C. A 16 array study using total RNA recovered from the following: 8 separate cultures of Ralstonia solanacearum strain UW551 grown in rich medium-CPG (4 grown at 20°C and 4 grown at 28°C) 8 separate in planta samples of Ralstonia solanacearum strain UW551 harvested from diseased tomato plants cv. BonnyBest (4 recovered from plants grown at 20°C and 4 from plants grown at 28°C) Each array (4-plex format) measures the expression level of 4318 genes from Ralstonia solanacearum strain UW551 with 2 to 6, 40-70 mer probes per gene, with two-fold technical redundancy. The arrays also contain probes for intergenic regions with no technical replicates.
Project description:Biofilm lifestyle is critical for bacterial pathogens to colonize and protect themselves from host immunity and antimicrobial chemicals in plants and animals. The formation and regulation mechanism of phytobacterial biofilm are still obscure. Here, we found that Ralstonia solanacearum Resistance to ultraviolet C (RuvC) is highly abundant in biofilm and positively regulates pathogenicity by governing systemic movement in tomato xylem. RuvC protein accumulates at the later stage of biofilm and specifically targets the Holliday junction (HJ) like structures to disrupt biofilm extracellular DNA (eDNA) lattice, thus facilitating biofilm dispersal. Recombinant RuvC protein can resolve extracellular HJ prevent bacterial biofilm formation. Heterologous expression of R. solanacearum or Xanthomonas oryzae pv. oryzae RuvC with plant secretion signal in tomato or rice confers resistance to bacterial wilt or bacterial blight disease, respectively. Plant chloroplast localized HJ resolvase monokaryotic chloroplast 1 (MOC1) which is structural similar to bacterial RuvC shows a strong inhibit effect on bacterial biofilm formation. Re-localization of SlMOC1 to apoplast in tomato roots leads to increase resistance to bacterial wilt. Our novel finding reveals a critical pathogenesis mechanism of R. solanacearum and provides an efficient biotechnology strategy to improve plant resistance to bacteria vascular disease.
Project description:Purpose: Investigate genes associated to resistance of Xanthomonas perforans race T4 in tomato line with different resistance level Methods: Resistant and susceptible tomato breeding lines were subjected to the inoculation with Xanthomonas perforans race T4 followed by sample collection at 48 hpi and RNA-seq analysis for screening differential expressed genes associated with inoculation of pathogen. Results: Revealed gene expression profiles associated disease resistance and susceptiblilty.
Project description:Investigation of whole genome gene expression level changes in the bacterial wilt pathogen Ralstonia solanacearum, strain GMI1000 at 20°C and 28°C in culture and in planta. The tropical strain GMI1000 cannot wilt tomato plants at 20°C although it can cause full-blown disease at 28°C.
Project description:Investigation of whole genome gene expression level changes in the bacterial wilt pathogen Ralstonia solanacearum, strain UW551 at 20°C and 28°C in culture and in planta. The temperatel strain UW551 can wilt and cause full-blown disease on tomato plants at 28°C as well as at 20°C.
Project description:The goal of the RNA seq was to investigate the transcriptome changes induced by Pseudmonas syringae pv. tomato J4 in wild-type tomato 'Moneymaker' and transgenic 'Moneymaker' overexpressing the Arabidopsis ELP4 (AtELP4) gene. Results showed that P. syringae pv. tomato J4 induced dramatic transcriptional changes in both the wild-type and transgenic tomato plants. Interestingly, a group of defense genes including PR-5x, Pti5, PR1b1, and CHI3/9/14/17, which are associated with resistance to the hemibiotrophic bacterial pathogen Ralstonia solanacearum, were induced to higher levels in the AtELP4 transgenic tomato than in the wild type at 8 and 24 hr after P. syringae pv. tomato J4 infection. These results indicate that overexpression of AtELP4 in tomato leads to faster and/or stronger induction of some defense genes.
Project description:Bacterial wilt caused by Ralstonia solanacearum is a lethal, soil-borne disease of tomato. Control of the disease with chemicals and crop rotation is insufficient, because the pathogen is particularly well adapted for surviving in the soil and rhizosphere. Therefore, cultivar resistance is the most effective means for controlling bacterial wilt, but the molecular mechanisms of resistance responses remain unclear. We used microarrays to obtain the characteristics of the gene expression changes that are induced by R. solanacearum infection in resistant cultivar LS-89 and susceptible cultivar Ponderosa.