In planta gene expression analysis of Xanthomonas oryzae pathovar oryzae, African strain MAI1

<p>Abstract</p> <p>Background</p> <p>Bacterial leaf blight causes significant yield losses in rice crops throughout Asia and Africa. Although both the Asian and African strains of the pathogen, <it>Xanthomonas oryzae </it>pv. <it>oryzae </it>(<it>Xoo</it>), induce similar symptoms, they are nevertheless genetically different, with the African strains being more closely related to the Asian <it>X. oryzae </it>pv. <it>oryzicola </it>(<it>Xoc</it>).</p> <p>Results</p> <p>Changes in gene expression of the African <it>Xoo </it>strain MAI1 in the susceptible rice cultivar Nipponbare were profiled, using an SSH <it>Xoo </it>DNA microarray. Microarray hybridization was performed comparing bacteria recovered from plant tissues at 1, 3, and 6 days after inoculation (dai) with bacteria grown <it>in vitro</it>. A total of 710 bacterial genes were found to be differentially expressed, with 407 up-regulated and 303 down-regulated. Expression profiling indicated that less than 20% of the 710 bacterial transcripts were induced in the first 24 h after inoculation, whereas 63% were differentially expressed at 6 dai. The 710 differentially expressed genes were one-end sequenced. 535 sequences were obtained from which 147 non-redundant sequences were identified. Differentially expressed genes were related to metabolism, secretion and transport, pathogen adherence to plant tissues, plant cell-wall degradation, IS elements, and virulence. In addition, various other genes encoding proteins with unknown function or showing no similarity to other proteins were also induced. The <it>Xoo </it>MAI1 non-redundant set of sequences was compared against several <it>X. oryzae </it>genomes, revealing a specific group of genes that was present only in MAI1. Numerous IS elements were also found to be differentially expressed. Quantitative real-time PCR confirmed 86% of the identified profile on a set of 14 genes selected according to the microarray analysis.</p> <p>Conclusions</p> <p>This is the first report to compare the expression of <it>Xoo </it>genes <it>in planta </it>across different time points during infection. This work shows that as-yet-unidentified and potentially new virulence factors are appearing in an emerging African pathogen. It also confirms that African <it>Xoo </it>strains do differ from their Asian counterparts, even at the transcriptional level.</p

Identification of Genes Regulating Gene Targeting by a High-Throughput Screening Approach

Homologous gene targeting (HGT) is a precise but inefficient process for genome engineering. Several methods for increasing its efficiency have been developed, including the use of rare cutting endonucleases. However, there is still room for improvement, as even nuclease-induced HGT may vary in efficiency as a function of the nuclease, target site, and cell type considered. We have developed a high-throughput screening assay for the identification of factors stimulating meganuclease-induced HGT. We used this assay to explore a collection of siRNAs targeting 19,121 human genes. At the end of secondary screening, we had identified 64 genes for which knockdown affected nuclease-induced HGT. Two of the strongest candidates were characterized further. We showed that siRNAs directed against the ATF7IP gene, encoding a protein involved in chromatin remodeling, stimulated HGT by a factor of three to eight, at various loci and in different cell types. This method thus led to the identification of a number of genes, the manipulation of which might increase rates of targeted recombination

Ancestral synteny shared between distantly-related plant species from the asterid (Coffea canephora and Solanum Sp.) and rosid (Vitis vinifera) clades

Background: Coffee trees (Rubiaceae) and tomato (Solanaceae) belong to the Asterid clade, while grapevine (Vitaceae) belongs to the Rosid clade. Coffee and tomato separated from grapevine 125 million years ago, while coffee and tomato diverged 83-89 million years ago. These long periods of divergent evolution should have permitted the genomes to reorganize significantly. So far, very few comparative mappings have been performed between very distantly related species belonging to different clades. We report the first multiple comparison between species from Asterid and Rosid clades, to examine both macro-and microsynteny relationships. Results: Thanks to a set of 867 COSII markers, macrosynteny was detected between coffee, tomato and grapevine. While coffee and tomato genomes share 318 orthologous markers and 27 conserved syntenic segments (CSSs), coffee and grapevine also share a similar number of syntenic markers and CSSs: 299 and 29 respectively. Despite large genome macrostructure reorganization, several large chromosome segments showed outstanding macrosynteny shedding new insights into chromosome evolution between Asterids and Rosids. We also analyzed a sequence of 174 kb containing the ovate gene, conserved in a syntenic block between coffee, tomato and grapevine that showed a high-level of microstructure conservation. A higher level of conservation was observed between coffee and grapevine, both woody and long life-cycle plants, than between coffee and tomato. Out of 16 coffee genes of this syntenic segment, 7 and 14 showed complete synteny between coffee and tomato or grapevine, respectively. Conclusions: These results show that significant conservation is found between distantly related species from the Asterid (Coffea canephora and Solanum sp.) and Rosid (Vitis vinifera) clades, at the genome macrostructure and microstructure levels. At the ovate locus, conservation did not decline in relation to increasing phylogenetic distance, suggesting that the time factor alone does not explain divergences. Our results are considerably useful for syntenic studies between supposedly remote species for the isolation of important genes for agronomy

Site-Specific Insertion Polymorphism of the MITE Alex-1 in the Genus Coffea Suggests Interspecific Gene Flow

Miniature Inverted-repeat Transposable Elements (MITEs) are small nonautonomous class-II transposable elements distributed throughout eukaryotic genomes. We identified a novel family of MITEs (named Alex) in the Coffea canephora genome often associated with expressed sequences. The Alex-1 element is inserted in an intron of a gene at the CcEIN4 locus. Its mobility was demonstrated by sequencing the insertion site in C. canephora accessions and Coffea species. Analysis of the insertion polymorphism of Alex-1 at this locus in Coffea species and in C. canephora showed that there was no relationship between the geographical distribution of the species, their phylogenetic relationships, and insertion polymorphism. The intraspecific distribution of C. canephora revealed an original situation within the E diversity group. These results suggest possibly greater gene flow between species than previously thought. This MITE family will enable the study of the C. canephora genome evolution, phylogenetic relationships, and possible gene flows within the Coffea genus

Microcollinearity in an ethylene receptor coding gene region of the Coffea canephora genome is extensively conserved with Vitis vinifera and other distant dicotyledonous sequenced genomes

Background: Coffea canephora, also called Robusta, belongs to the Rubiaceae, the fourth largest angiosperm family. This diploid species (2x = 2n = 22) has a fairly small genome size of approximate to 690 Mb and despite its extreme economic importance, particularly for developing countries, knowledge on the genome composition, structure and evolution remain very limited. Here, we report the 160 kb of the first C. canephora Bacterial Artificial Chromosome (BAC) clone ever sequenced and its fine analysis. Results: This clone contains the CcEIN4 gene, encoding an ethylene receptor, and twenty other predicted genes showing a high gene density of one gene per 7.8 kb. Most of them display perfect matches with C. canephora expressed sequence tags or show transcriptional activities through PCR amplifications on cDNA libraries. Twenty-three transposable elements, mainly Class II transposon derivatives, were identified at this locus. Most of these Class II elements are Miniature Inverted-repeat Transposable Elements (MITE) known to be closely associated with plant genes. This BAC composition gives a pattern similar to those found in gene rich regions of Solanum lycopersicum and Medicago truncatula genomes indicating that the CcEIN4 regions may belong to a gene rich region in the C. canephora genome. Comparative sequence analysis indicated an extensive conservation between C. canephora and most of the reference dicotyledonous genomes studied in this work, such as tomato (S. lycopersicum), grapevine (V. vinifera), barrel medic M. truncatula, black cottonwood (Populus trichocarpa) and Arabidopsis thaliana. The higher degree of microcollinearity was found between C. canephora and V. vinifera, which belong respectively to the Asterids and Rosids, two clades that diverged more than 114 million years ago. Conclusion: This study provides a first glimpse of C. canephora genome composition and evolution. Our data revealed a remarkable conservation of the microcollinearity between C. canephora and V. vinifera and a high conservation with other distant dicotyledonous reference genomes. Altogether, these results provide valuable information to identify candidate genes in C. canephora genome and serve as a foundation to establish strategies for whole genome sequencing. Future large-scale sequence comparison between C. canephora and reference sequenced genomes will help in understanding the evolutionary history of dicotyledonous plants

Clinical management of molecular alterations identified by high throughput sequencing in patients with advanced solid tumors in treatment failure: Real-world data from a French hospital

BackgroundIn the context of personalized medicine, screening patients to identify targetable molecular alterations is essential for therapeutic decisions such as inclusion in clinical trials, early access to therapies, or compassionate treatment. The objective of this study was to determine the real-world impact of routine incorporation of FoundationOne analysis in cancers with a poor prognosis and limited treatment options, or in those progressing after at least one course of standard therapy.MethodsA FoundationOneCDx panel for solid tumor or liquid biopsy samples was offered to 204 eligible patients.ResultsSamples from 150 patients were processed for genomic testing, with a data acquisition success rate of 93%. The analysis identified 2419 gene alterations, with a median of 11 alterations per tumor (range, 0–86). The most common or likely pathogenic variants were on TP53, TERT, PI3KCA, CDKN2A/B, KRAS, CCDN1, FGF19, FGF3, and SMAD4. The median tumor mutation burden was three mutations/Mb (range, 0–117) in 143 patients with available data. Of 150 patients with known or likely pathogenic actionable alterations, 13 (8.6%) received matched targeted therapy. Sixty-nine patients underwent Molecular Tumor Board, which resulted in recommendations in 60 cases. Treatment with genotype-directed therapy had no impact on overall survival (13 months vs. 14 months; p = 0.95; hazard ratio = 1.04 (95% confidence interval, 0.48–2.26)].ConclusionsThis study highlights that an organized center with a Multidisciplinary Molecular Tumor Board and an NGS screening system can obtain satisfactory results comparable with those of large centers for including patients in clinical trials

Improving building envelope knowledge from analysis of 219,000 certified on-site air leakage measurements in France

International audienc

French building airtightness database after 10 years of operation: statistical analyses of about 500,000 measurements

International audienc

Description of new shock-induced phases in the meteorites of Shergotty, Zagami, Nakhla and Chassigny

International audienceThe SNC meteorites, Shergotty, Zagami, Nakhla and Chassigny, have beenstudied by analytical transmission electron microscopy. New phases, characteristic ofstrong shock conditions, have been discovered in Shergotty, Zagami, Nakhla andChassigny : calcium-rich majorites in Shergottty, wadsleyite with anomalously elevatediron content in Chassigny and impact melts in Shergotty, Zagami and Nakhla.Cristobalites (α and β polymorphs) observed in Shergotty and Zagami may also berelated to shock and are interpreted as backtransformation products of post-stishovitesilica polymorphs. Strong shocks corresponding to pressure and temperature conditionscharacteristic of the Earth’s transition zone and lower mantle have occurred in thosemeteorites. Moreover, impact melts indicate high temperature conditions in localizedareas. On the other hand, no massive impact melting is observed in those meteorites,consistently with previous descriptions. These observations evidence highlyheterogeneous shock conditions at the scale of few micrometers in these meteorites.Strongly heterogeneous conditions such as those suggested by the present study mayhelp to interpret the preservation in martian meteorites of phases practically unaffectedby shock being very close to strongly shock-metamorphized minerals