Identification of novel genes potentially involved in somatic embryogenesis in chicory (Cichorium intybus L.)

<p>Abstract</p> <p>Background</p> <p>In our laboratory we use cultured chicory (<it>Cichorium intybus</it>) explants as a model to investigate cell reactivation and somatic embryogenesis and have produced 2 chicory genotypes (K59, C15) sharing a similar genetic background. K59 is a responsive genotype (embryogenic) capable of undergoing complete cell reactivation <it>i.e</it>. cell de- and re-differentiation leading to somatic embryogenesis (SE), whereas C15 is a non-responsive genotype (non-embryogenic) and is unable to undergo SE. Previous studies <abbrgrp><abbr bid="B1">1</abbr></abbrgrp> showed that the use of the β-D-glucosyl Yariv reagent (β-GlcY) that specifically binds arabinogalactan-proteins (AGPs) blocked somatic embryo production in chicory root explants. This observation indicates that β-GlcY is a useful tool for investigating somatic embryogenesis (SE) in chicory. In addition, a putative AGP (DT212818) encoding gene was previously found to be significantly up-regulated in the embryogenic K59 chicory genotype as compared to the non-embryogenic C15 genotype suggesting that this AGP could be involved in chicory re-differentiation <abbrgrp><abbr bid="B2">2</abbr></abbrgrp>. In order to improve our understanding of the molecular and cellular regulation underlying SE in chicory, we undertook a detailed cytological study of cell reactivation events in K59 and C15 genotypes, and used microarray profiling to compare gene expression in these 2 genotypes. In addition we also used β-GlcY to block SE in order to identify genes potentially involved in this process.</p> <p>Results</p> <p>Microscopy confirmed that only the K59, but not the C15 genotype underwent complete cell reactivation leading to SE formation. β-GlcY-treatment of explants blocked <it>in vitro </it>SE induction, but not cell reactivation, and induced cell wall modifications. Microarray analyses revealed that 78 genes were differentially expressed between induced K59 and C15 genotypes. The expression profiles of 19 genes were modified by β-GlcY-treatment. Eight genes were both differentially expressed between K59 and C15 genotypes during SE induction and transcriptionally affected by β-GlcY-treatment: <it>AGP </it>(DT212818), <it>26 S proteasome AAA ATPase subunit 6 </it>(<it>RPT6</it>), <it>remorin </it>(<it>REM</it>), <it>metallothionein-1 </it>(<it>MT1</it>), two non-specific lipid transfer proteins genes (<it>SDI-9 and DEA1</it>), <it>3-hydroxy-3-methylglutaryl-CoA reductase </it>(<it>HMG-CoA reductase</it>), and <it>snakin 2 </it>(<it>SN2</it>). These results suggest that the 8 genes, including the previously-identified <it>AGP </it>gene (DT212818), could be involved in cell fate determination events leading to SE commitment in chicory.</p> <p>Conclusion</p> <p>The use of two different chicory genotypes differing in their responsiveness to SE induction, together with β-GlcY-treatment represented an efficient tool to discriminate cell reactivation from the SE morphogenetic pathway. Such an approach, together with microarray analyses, permitted us to identify several putative key genes related to the SE morphogenetic pathway in chicory.</p

The deficiency in EntDD14 bacteriocin synthesis induced metabolic rearrangements to counterbalance its loss in Enterococcus faecalis 14, as revealed by a transcriptomic analysis

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Metabolic Shift of an Isogenic Strain of Enterococcus faecalis 14, Deficient in Its Own Bacteriocin Synthesis, as Revealed by a Transcriptomic Analysis

The production of antimicrobial molecules often involves complex biological pathways. This study aimed at understanding the metabolic and physiological networks of enterocin EntDD14-associated function, in the bacteriocinogenic strain, Enterococcus faecalis 14. A global and comparative transcriptomic study was carried out on E. faecalis 14 and its isogenic mutant Δbac, inactivated in genes coding for EntDD14. The in vitro ability to form biofilm on polystyrene plates was assessed by the crystal violet method, while the cytotoxicity on human colorectal adenocarcinoma Caco-2 cells was determined by the Cell Counting Kit-8. Transcriptomic data revealed that 71 genes were differentially expressed in both strains. As expected, genes coding for EntDD14 were downregulated in the Δbac mutant, whereas the other 69 genes were upregulated. Upregulated genes were associated with phage-related chromosomal islands, biofilm formation capability, resistance to environmental stresses, and metabolic reprogramming. Interestingly, the Δbac mutant showed an improved bacterial growth, a high capacity to form biofilm on inanimate surfaces and a very weak cytotoxicity level. These multiple metabolic rearrangements delineate a new line of defense to counterbalance the loss of EntDD14

Lack of PNPase activity in Enterococcus faecalis 14 increases the stability of EntDD14 bacteriocin transcripts

Abstract A mutant deficient in polynucleotide phosphorylase (PNPase) activity was previously constructed in Enterococcus faecalis 14; a strain producing a leaderless two-peptide enterocin DD14 (EntDD14). Here, we examined the impact of the absence of PNPase on the expression and synthesis of EntDD14, at the transcriptional and functional levels. As result, EntDD14 synthesis augmented in line with the growth curve, reaching a two- to fourfold increase in the ΔpnpA mutant compared to the E. faecalis 14 wild-type strain (WT). EntDD14 synthesis has reached its highest level after 9 h of growth in both strains. Notably, high expression level of the ddABCDEFGHIJ cluster was registered in ΔpnpA mutant. Transcriptional and in silico analyses support the existence of ddAB and ddCDEFGHIJ independent transcripts, and analysis of the fate of ddAB and ddCDEFGHIJ mRNAs indicated that the differences in mRNA levels and the high EntDD14 activity are likely due to a better stability of the two transcripts in the ΔpnpA mutant, which should result in a higher translation efficiency of the ddAB EntDD14 structural genes and their other protein determinants. Consequently, this study shows a potential link between the mRNA stability and EntDD14 synthesis, secretion and immunity in a genetic background lacking PNPase

The absence of PNPase activity in Enterococcus faecalis results in alterations of the bacterial cell-wall but induces high proteolytic and adhesion activities

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Lack of PNPase activity in Enterococcus faecalis 14 increases the stability of EntDD14 bacteriocin transcripts

10 p.-7 fig.A mutant deficient in polynucleotide phosphorylase (PNPase) activity was previously constructed in Enterococcus faecalis 14; a strain producing a leaderless two-peptide enterocin DD14 (EntDD14). Here, we examined the impact of the absence of PNPase on the expression and synthesis of EntDD14, at the transcriptional and functional levels. As result, EntDD14 synthesis augmented in line with the growth curve, reaching a two- to fourfold increase in the ΔpnpA mutant compared to the E. faecalis 14 wild-type strain (WT). EntDD14 synthesis has reached its highest level after 9 h of growth in both strains. Notably, high expression level of the ddABCDEFGHIJ cluster was registered in ΔpnpA mutant. Transcriptional and in silico analyses support the existence of ddAB and ddCDEFGHIJ independent transcripts, and analysis of the fate of ddAB and ddCDEFGHIJ mRNAs indicated that the differences in mRNA levels and the high EntDD14 activity are likely due to a better stability of the two transcripts in the ΔpnpA mutant, which should result in a higher translation efficiency of the ddAB EntDD14 structural genes and their other protein determinants. Consequently, this study shows a potential link between the mRNA stability and EntDD14 synthesis, secretion and immunity in a genetic background lacking PNPase.This research was funded by Bacterioplus project START’AIR project and the CPER BiHauts Eco de France 2021/2027.Peer reviewe

Efficacy of the bacteriocinogenic Enterococcus faecalis 14 in controlling experimentally induced necrotic enteritis in broilers under floorpen conditions

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In vivo efficacy of Enterococcus faecalis 14 treatment of necrotic enteritis in broiler chickens

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La mycorhization protège le blé contre la septoriose: analyse transcriptomique de la résistance induite

International audienceLa recherche d’alternatives aux produits phytosanitaires, néfastes pour l’environnement et l’être humain, devient prioritaire. En remplacement des fongicides conventionnels, les champignons mycorhiziens à arbuscules (CMA) permettraient de lutter contre les maladies fongiques du blé. Des études ont démontré l’efficacité d’un CMA, Funneliformis mosseae (Fm), à protéger le blé contre l’oïdium et la septoriose, en lien avec l’établissement dans les feuilles d’une résistance induite (MIR pour Mycorrhiza-Induced Resistance). Notre travail vise à caractériser les modifications de l’expression des gènes lors de l’établissement de la MIR dans les feuilles de blé, susceptibles de conduire à la protection contre Zymoseptoria tritici (Zt), agent pathogène responsable de la septoriose. Des feuilles de plants de 3, 4 et 6 semaines, inoculés ou non par Fm, ont été échantillonnées pour étudier l’expression des gènes durant l’établissement de la symbiose racinaire. Puis à 6 semaines, des plantes, mycorhizées ou non par Fm, ont été infectées par Zt ; leur réaction à l’infection a été analysée après 48 heures. A 3 et 4 semaines, 9 gènes, liés au stress abiotique et au métabolisme lipidique, sont sur-exprimés chez les plants mycorhizés. A 6 semaines, 214 gènes, majoritairement liés aux stress biotiques et abiotiques et au métabolisme glucidique, sont sur-exprimés chez les plants mycorhizés. 162 d’entre eux sont également sur-exprimés chez les plants non-mycorhizés et infectés. Par ailleurs,120 gènes sont sous-exprimés chez les plants mycorhizés infectés en comparaison des plants non-mycorhizés et infectés. Enfin, 41 gènes, induits chez des plants mycorhizés non-infectés, sont sous-exprimés chez les plants mycorhizés et infectés. L’analyse fonctionnelle des gènes modulés a été réalisée. Une analyse par RT-qPCR affinera les profils d’expression de gènes modulés par la mycorhization et possiblement impliqués dans la MIR. Leur expression pourra être suivie lors de l’inoculation du blé par d’autres espèces de CMA afin de sélectionner de potentiels marqueurs de la MIR