scholarly journals Re-programming of Pseudomonas syringae pv. actinidiae gene expression during early stages of infection of kiwifruit

2018 ◽  
Author(s):  
Peter A. McAtee ◽  
Lara Brian ◽  
Ben Curran ◽  
Otto van der Linden ◽  
Niels J. Nieuwenhuizen ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Syringae ◽  
Time Course ◽  
Secretion System ◽  
Nutrient Acquisition ◽  
Post Inoculation ◽  
Rna Seq ◽  
Type Iii ◽  
Early Stages ◽  
Expression Of Genes

AbstractBackgroundPseudomonas syringae is a widespread bacterial species complex that includes a number of significant plant pathogens. Amongst these, P. syringae pv. actinidiae (Psa) initiated a worldwide pandemic in 2008 on cultivars of Actinidia chinensis var. chinensis. To gain information about the expression of genes involved in pathogenicity we have carried out transcriptome analysis of Psa during the early stages of kiwifruit infection.ResultsGene expression in Psa was investigated during the first five days after infection of kiwifruit plantlets, using RNA-seq. Principal component and heatmap analyses showed distinct phases of gene expression during the time course of infection. The first phase was an immediate transient peak of induction around three hours post inoculation (HPI) that included genes that code for a Type VI Secretion System and nutrient acquisition (particularly phosphate). This was followed by a significant commitment, between 3 and 24 HPI, to the induction of genes encoding the Type III Secretion System (T3SS) and Type III Secreted Effectors (T3SE). Expression of these genes collectively accounted for 6.3% of the bacterial transcriptome at this stage. There was considerable variation in the expression levels of individual T3SEs but all followed the same temporal expression pattern, with the exception of HopAS1, which peaked later in expression at 48 HPI. As infection progressed over the time course of five days, there was an increase in the expression of genes with roles in sugar, amino acid and sulfur transport and the production of alginate and colanic acid. These are both polymers that are major constituents of extracellular polysaccharide substances (EPS) and are involved in biofilm production. Reverse transcription-quantitative PCR (RT-qPCR) on an independent infection time course experiment showed that the expression profile of selected bacterial genes at each infection phase correlated well with the RNA-seq data.ConclusionsThe results from this study indicate that there is a complex remodeling of the transcriptome during the early stages of infection, with at least three distinct phases of coordinated gene expression. These include genes induced during the immediate contact with the host, those involved in the initiation of infection, and finally those responsible for nutrient acquisition.

scholarly journals Positive Regulation of the Hrp Type III Secretion System in Pseudomonas syringae pv. phaseolicola

2010 ◽  
Vol 23 (5) ◽  
pp. 665-681 ◽  
Author(s):  
Inmaculada Ortiz-Martín ◽  
Richard Thwaites ◽  
Alberto P. Macho ◽  
John W. Mansfield ◽  
Carmen R. Beuzón
Keyword(s):  
Gene Expression ◽  
Pseudomonas Syringae ◽  
Hypersensitive Response ◽  
Type Iii Secretion ◽  
Pathogenic Bacteria ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Virulence Determinants ◽  
Type Iii

Disease in compatible hosts and induction of the hypersensitive response in resistant plants by most plant-pathogenic bacteria require a functional type III secretion system (T3SS). Expression of T3SS genes responds to host and environmental factors and is induced within the plant. In Pseudomonas syringae, expression of the T3SS requires HrpL, which is transcriptionally upregulated by HrpR and HrpS. In some pathovars, expression of the hrpRS genes is upregulated by the GacA/S two-component system. Additionally, HrpA, the major component of the T3SS pilus, has also been linked to the regulation of the hrpRS gene expression. Previous studies concerning regulation of hypersensitive response and pathogenesis/hypersensitive response conserved (hrp/hrc) gene expression have used mostly in vitro inducing conditions, different pathovars, and methodology. Here, we analyze the roles of HrpL, GacA, and HrpA in the bean pathogen, using single, double, and triple mutants as well as strains ectopically expressing the regulators. We use real-time polymerase chain reaction analysis in vitro and in planta to quantify gene expression and competitive indices and other assays to assess bacterial fitness. Our results indicate that i) HrpL acts as a general virulence regulator that upregulates non-T3SS virulence determinants and downregulates flagellar function; ii) GacA modulates the expression of hrpL, and its contribution to virulence is entirely HrpL dependent; iii) there is a basal HrpL-independent expression of the T3SS genes in rich medium that is important for full activation of the system, maybe by keeping the system primed for rapid activation upon contact with the plant; and iv) HrpA upregulates expression of the T3SS genes and is essential to activate expression of the hrpZ operon upon contact with the plant.

scholarly journals Transcriptional profiling of three Pseudomonas syringae pv. actinidiae biovars reveals different responses to apoplast-like conditions related to strain virulence

2020 ◽  
Author(s):  
Elodie Vandelle ◽  
Teresa Colombo ◽  
Alice Regaiolo ◽  
Tommaso Libardi ◽  
Vanessa Maurizio ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Molecular Basis ◽  
Expression Profiles ◽  
Type Iii Secretion System ◽  
Secretion System ◽  
Type Iii ◽  
Genes Encoding ◽  
Operon Regulation

AbstractPseudomonas syringae pv. actinidiae (Psa) is a phytopathogen that causes devastating bacterial canker in kiwifruit. Among five biovars defined by genetic, biochemical and virulence traits, Psa3 is the most aggressive and is responsible for the most recent reported outbreaks, but the molecular basis of its heightened virulence is unclear. We therefore designed the first P. syringae multi-strain whole-genome microarray, encompassing biovars Psa1, Psa2 and Psa3 and the well-established model P. syringae pv. tomato, and analyzed early bacterial responses to an apoplast-like minimal medium. Transcriptomic profiling revealed (i) the strong activation in Psa3 of all hrp/hrc cluster genes, encoding components of the type III secretion system required for bacterial pathogenicity and involved in responses to environmental signals; (ii) potential repression of the hrp/hrc cluster in Psa2; and (iii) activation of flagellum-dependent cell motility and chemotaxis genes in Psa1. The detailed investigation of three gene families encoding upstream regulatory proteins (histidine kinases, their cognate response regulators, and proteins with diguanylate cyclase and/or phosphodiesterase domains) indicated that c-di-GMP may be a key regulator of virulence in Psa biovars. The gene expression data were supported by the quantification of biofilm formation. Our findings suggest that diverse early responses to the host apoplast, even among bacteria belonging to the same pathovar, can lead to different virulence strategies and may explain the differing outcomes of infections. Based on our detailed structural analysis of hrp operons, we also propose a revision of hrp cluster organization and operon regulation in P. syringae.Author summaryPseudomonas syringae pv. actinidiae (Psa) is a bacterial pathogen that infects kiwifruit crops. Recent outbreaks have been particularly devastating due to the emergence of a new biovar (Psa3), but the molecular basis of its virulence is unknown so it is difficult to develop mitigation strategies. In this study, we compared the gene expression profiles of Psa3 and various less-virulent biovars in an environment that mimics early infection, to determine the basis of pathogenicity. Genes involved in the assembly and activity of the type III secretion system, which is crucial for the secretion of virulence effectors, were strongly upregulated in Psa3 while lower or not expressed in the other biovars. We also observed the Psa3-specific expression of genes encoding upstream signaling components, confirming that strains of the same bacterial pathovar can respond differently to early contact with their host. Finally, our data suggested a key role in Psa virulence switch ability for the small chemical signaling molecule c-di-GMP, which suppresses the expression of virulence genes. This effect of c-di-GMP levels on Psa3 virulence should be further investigated and confirmed to develop new mitigation methods to target this pathway.

scholarly journals SPLICE-q: a Python tool for genome-wide quantification of splicing efficiency

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Verônica R. de Melo Costa ◽  
Julianus Pfeuffer ◽  
Annita Louloupi ◽  
Ulf A. V. Ørom ◽  
Rosario M. Piro
Keyword(s):  
Gene Expression ◽  
Cancer Progression ◽  
Time Course ◽  
Progressive Increase ◽  
Rna Seq ◽  
Transcript Processing ◽  
Aberrant Transcript ◽  
Genome Wide ◽  
Splicing Efficiency ◽  
Nascent Rna

Abstract Background Introns are generally removed from primary transcripts to form mature RNA molecules in a post-transcriptional process called splicing. An efficient splicing of primary transcripts is an essential step in gene expression and its misregulation is related to numerous human diseases. Thus, to better understand the dynamics of this process and the perturbations that might be caused by aberrant transcript processing it is important to quantify splicing efficiency. Results Here, we introduce SPLICE-q, a fast and user-friendly Python tool for genome-wide SPLICing Efficiency quantification. It supports studies focusing on the implications of splicing efficiency in transcript processing dynamics. SPLICE-q uses aligned reads from strand-specific RNA-seq to quantify splicing efficiency for each intron individually and allows the user to select different levels of restrictiveness concerning the introns’ overlap with other genomic elements such as exons of other genes. We applied SPLICE-q to globally assess the dynamics of intron excision in yeast and human nascent RNA-seq. We also show its application using total RNA-seq from a patient-matched prostate cancer sample. Conclusions Our analyses illustrate that SPLICE-q is suitable to detect a progressive increase of splicing efficiency throughout a time course of nascent RNA-seq and it might be useful when it comes to understanding cancer progression beyond mere gene expression levels. SPLICE-q is available at: https://github.com/vrmelo/SPLICE-q

scholarly journals Pseudomonas syringae Strains Naturally Lacking the Classical P. syringae hrp/hrc Locus Are Common Leaf Colonizers Equipped with an Atypical Type III Secretion System

2010 ◽  
Vol 23 (2) ◽  
pp. 198-210 ◽  
Author(s):  
Christopher R. Clarke ◽  
Rongman Cai ◽  
David J. Studholme ◽  
David S. Guttman ◽  
Boris A. Vinatzer
Keyword(s):  
Pseudomonas Syringae ◽  
Type Iii Secretion ◽  
Plant Cells ◽  
Type Iii Secretion System ◽  
Ice Nucleation ◽  
Secretion System ◽  
Effector Proteins ◽  
Population Densities ◽  
Type Iii ◽  
Effector Genes

Pseudomonas syringae is best known as a plant pathogen that causes disease by translocating immune-suppressing effector proteins into plant cells through a type III secretion system (T3SS). However, P. syringae strains belonging to a newly described phylogenetic subgroup (group 2c) are missing the canonical P. syringae hrp/hrc cluster coding for a T3SS, flanking effector loci, and any close orthologue of known P. syringae effectors. Nonetheless, P. syringae group 2c strains are common leaf colonizers and grow on some tested plant species to population densities higher than those obtained by other P. syringae strains on nonhost species. Moreover, group 2c strains have genes necessary for the production of phytotoxins, have an ice nucleation gene, and, most interestingly, contain a novel hrp/hrc cluster, which is only distantly related to the canonical P. syringae hrp/hrc cluster. This hrp/hrc cluster appears to encode a functional T3SS although the genes hrpK and hrpS, present in the classical P. syringae hrp/hrc cluster, are missing. The genome sequence of a representative group 2c strain also revealed distant orthologues of the P. syringae effector genes avrE1 and hopM1 and the P. aeruginosa effector genes exoU and exoY. A putative life cycle for group 2c P. syringae is discussed.

scholarly journals BgeeDB, an R package for retrieval of curated expression datasets and for gene list expression localization enrichment tests

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 2748 ◽  
Author(s):  
Andrea Komljenovic ◽  
Julien Roux ◽  
Marc Robinson-Rechavi ◽  
Frederic B. Bastian
Keyword(s):  
Gene Expression ◽  
Gene List ◽  
R Package ◽  
Rna Seq ◽  
Anatomical Structures ◽  
Enrichment Test ◽  
Expression Of Genes ◽  
Affymetrix Microarrays ◽  
New Gene

BgeeDB is a collection of functions to import into R re-annotated, quality-controlled and reprocessed expression data available in the Bgee database. This includes data from thousands of wild-type healthy samples of multiple animal species, generated with different gene expression technologies (RNA-seq, Affymetrix microarrays, expressed sequence tags, and in situ hybridizations). BgeeDB facilitates downstream analyses, such as gene expression analyses with other Bioconductor packages. Moreover, BgeeDB includes a new gene set enrichment test for preferred localization of expression of genes in anatomical structures (“TopAnat”). Along with the classical Gene Ontology enrichment test, this test provides a complementary way to interpret gene lists. Availability: http://www.bioconductor.org/packages/BgeeDB/

scholarly journals Identification and classification of cis-regulatory elements in the amphipod crustacean Parhyale hawaiensis

2021 ◽  
Author(s):  
Dennis A Sun ◽  
Nipam H Patel
Keyword(s):  
Gene Expression ◽  
Genome Annotation ◽  
Time Course ◽  
Regulatory Elements ◽  
Rna Seq ◽  
Genome Wide ◽  
Long Read ◽  
Amphipod Crustacean ◽  
Accessible Chromatin

AbstractEmerging research organisms enable the study of biology that cannot be addressed using classical “model” organisms. The development of novel data resources can accelerate research in such animals. Here, we present new functional genomic resources for the amphipod crustacean Parhyale hawaiensis, facilitating the exploration of gene regulatory evolution using this emerging research organism. We use Omni-ATAC-Seq, an improved form of the Assay for Transposase-Accessible Chromatin coupled with next-generation sequencing (ATAC-Seq), to identify accessible chromatin genome-wide across a broad time course of Parhyale embryonic development. This time course encompasses many major morphological events, including segmentation, body regionalization, gut morphogenesis, and limb development. In addition, we use short- and long-read RNA-Seq to generate an improved Parhyale genome annotation, enabling deeper classification of identified regulatory elements. We leverage a variety of bioinformatic tools to discover differential accessibility, predict nucleosome positioning, infer transcription factor binding, cluster peaks based on accessibility dynamics, classify biological functions, and correlate gene expression with accessibility. Using a Minos transposase reporter system, we demonstrate the potential to identify novel regulatory elements using this approach, including distal regulatory elements. This work provides a platform for the identification of novel developmental regulatory elements in Parhyale, and offers a framework for performing such experiments in other emerging research organisms.Primary Findings-Omni-ATAC-Seq identifies cis-regulatory elements genome-wide during crustacean embryogenesis-Combined short- and long-read RNA-Seq improves the Parhyale genome annotation-ImpulseDE2 analysis identifies dynamically regulated candidate regulatory elements-NucleoATAC and HINT-ATAC enable inference of nucleosome occupancy and transcription factor binding-Fuzzy clustering reveals peaks with distinct accessibility and chromatin dynamics-Integration of accessibility and gene expression reveals possible enhancers and repressors-Omni-ATAC can identify known and novel regulatory elements

scholarly journals Importance of experimental information (metadata) for archived sequence data: case of specific gene bias due to lag time between sample harvest and RNA protection in RNA sequencing

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e11875
Author(s):  
Tomoko Matsuda
Keyword(s):  
Gene Expression ◽  
Rna Sequencing ◽  
Time Course ◽  
Sequence Data ◽  
Specific Gene ◽  
Time Interval ◽  
Short Time Interval ◽  
Rna Seq ◽  
Lysis Buffer ◽  
Rna Protection

Large volumes of high-throughput sequencing data have been submitted to the Sequencing Read Archive (SRA). The lack of experimental metadata associated with the data makes reuse and understanding data quality very difficult. In the case of RNA sequencing (RNA-Seq), which reveals the presence and quantity of RNA in a biological sample at any moment, it is necessary to consider that gene expression responds over a short time interval (several seconds to a few minutes) in many organisms. Therefore, to isolate RNA that accurately reflects the transcriptome at the point of harvest, raw biological samples should be processed by freezing in liquid nitrogen, immersing in RNA stabilization reagent or lysing and homogenizing in RNA lysis buffer containing guanidine thiocyanate as soon as possible. As the number of samples handled simultaneously increases, the time until the RNA is protected can increase. Here, to evaluate the effect of different lag times in RNA protection on RNA-Seq data, we harvested CHO-S cells after 3, 5, 6, and 7 days of cultivation, added RNA lysis buffer in a time course of 15, 30, 45, and 60 min after harvest, and conducted RNA-Seq. These RNA samples showed high RNA integrity number (RIN) values indicating non-degraded RNA, and sequence data from libraries prepared with these RNA samples was of high quality according to FastQC. We observed that, at the same cultivation day, global trends of gene expression were similar across the time course of addition of RNA lysis buffer; however, the expression of some genes was significantly different between the time-course samples of the same cultivation day; most of these differentially expressed genes were related to apoptosis. We conclude that the time lag between sample harvest and RNA protection influences gene expression of specific genes. It is, therefore, necessary to know not only RIN values of RNA and the quality of the sequence data but also how the experiment was performed when acquiring RNA-Seq data from the database.

scholarly journals Concomitant Regulation by a LacI-Type Transcriptional Repressor XylR on Genes Involved in Xylan and Xylose Metabolism and the Type III Secretion System in Rice Pathogen Xanthomonas oryzae pv. oryzae

2018 ◽  
Vol 31 (6) ◽  
pp. 605-613 ◽  
Author(s):  
Yumi Ikawa ◽  
Sayaka Ohnishi ◽  
Akiko Shoji ◽  
Ayako Furutani ◽  
Seiji Tsuge
Keyword(s):  
Gene Expression ◽  
Type Iii Secretion ◽  
Type Iii Secretion System ◽  
Xylose Isomerase ◽  
Secretion System ◽  
Xanthomonas Oryzae ◽  
Upstream Region ◽  
Xylose Metabolism ◽  
Type Iii ◽  
Hrp Genes

The hypersensitive response and pathogenicity (hrp) genes of Xanthomonas oryzae pv. oryzae, the causal agent of bacterial leaf blight of rice, encode components of the type III secretion system and are essential for virulence. Expression of hrp genes is regulated by two key hrp regulators, HrpG and HrpX; HrpG regulates hrpX and hrpA, and HrpX regulates the other hrp genes on hrpB-hrpF operons. We previously reported the sugar-dependent quantitative regulation of HrpX; the regulator highly accumulates in the presence of xylose, followed by high hrp gene expression. Here, we found that, in a mutant lacking the LacI-type transcriptional regulator XylR, HrpX accumulation and hrp gene expression were high even in the medium without xylose, reaching the similar levels present in the wild type incubated in the xylose-containing medium. XylR also negatively regulated one of two xylose isomerase genes (xylA2 but not xylA1) by binding to the motif sequence in the upstream region of the gene. Xylose isomerase is an essential enzyme in xylose metabolism and interconverts between xylose and xylulose. Our results suggest that, in the presence of xylose, inactivation of XylR leads to greater xylan and xylose utilization and, simultaneously, to higher accumulation of HrpX, followed by higher hrp gene expression in the bacterium.

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