Synthesis of trehalose-based chemical tools for the study of the mycobacterial membrane

Corynebacteriales including the causative agent of many diseases such as tuberculosis are known to be extremely resistant against external stress as well as to antibiotic treatments which is believed to be related to the singular architecture of their mycomembrane. Over the last decades, both bioorthogonal chemical reporters and fluorescent probes for the metabolic labeling of bacterial cell glycans were developed including several trehalose-based probes to study the dynamics of mycomembrane components. This review presents an exhaustive view on the reported syntheses of trehalose-based probes enabling the study of the mycomembrane biogenesis

Trehalose and Mycolic Acid-Based Probes for the Study of Mycomembrane Biogenesis

Les Corynebactériales possèdent une membrane externe atypique, nommée mycomembrane, contenant des acides gras uniques appelés acides mycoliques. Ils sont notamment associés à un disaccharide, le tréhalose, formant le mono- ou dimycolate de tréhalose (TMM, TDM). La mycomembrane est résistante et imperméable aux antibiotiques et l’étude de ses constituants et de leurs voies de biosynthèse pourrait être utile au développement de nouveaux antibiotiques. Depuis quelques années, des analogues de composants de la mycomembrane, comportant une fonction bioorthogonale, ont été développés permettant son marquage métabolique selon la stratégie du rapporteur chimique. Cependant, aucun analogue bioorthogonal comportant la structure complexe des acides mycoliques naturels n’a été décrit. Ainsi, l’un des objectifs de ces travaux a été de développer des voies de synthèse permettant l’accès soit à des sondes bioorthogonales qui miment fidèlement la structure du TMM naturel, soit à des analogues de tréhalose clickables ou fluorescents. Ces nouvelles sondes ont ensuite été utilisées pour réaliser le marquage métabolique de la mycomembrane de C. glutamicum. Un autre objectif a été l’étude d’une mycoloyltransférase, MytC, utilisant le TMM comme donneur de mycoloyl sur des protéines membranaires. Un test in vitro a été mis au point grâce à du TMM synthétique et différents analogues de TMM ont été synthétisés et utilisés in vitro afin de réaliser une étude structure-activité. Ces outils chimiques ont apporté de nouvelles informations sur le mode d’action de cette enzyme de la biogénèse de la mycomembrane.Corynebacteriales are characterized by an atypical envelope, called mycomembrane (MM), containing unique fatty acids: the mycolic acids. They are esterified to trehalose to form trehalose mono- or dimycolate (TMM, TDM). MM forms an impermeable barrier that contribute to its exceptional resistance. MM components and their biosynthesis are extensively studied because they could provide opportunities for antibiotic development. Over the last decade, bioorthogonal chemical reporters have emerged as valuable tools for detecting bacterial cell surface components. However, these reporters do not mimic the native structure of mycolic acids. One of the objectives of this work was to develop synthetic ways allowing access to either bioorthogonal anlogues that closely mimic the native structure of mycolic acids, or clickable or fluorescent trehalose analogues. These new probes were then used in vivo to label C. glutamicum mycomembrane Another objective was the study of the mycoloyltransferase MytC, which is the only enzyme able to transfer the mycoloyl chain of a TMM onto membrane proteins. An in vitro assay was developed using synthetic TMM and different analogues of TMM were synthesized and used in vitro in a structure-relationship study

Sydnone‐Cyanines as Clickable Probes for Fluorescent Labelling

International audienceAbstract The synthesis of four clickable sydnone‐heptamethine cyanine derivatives is described in this article. The synthetic route is based on a palladium‐cross coupling reactions of sydnone boronates affording the desired sydnone‐cyanine conjugates in only five steps. These compounds were shown to react smoothly with cyclooctynes to form the corresponding pyrazoles clicked products quantitatively at room temperature and with rate constants up to 18 m −1 ⋅ s −1 , affording interesting new tools for biorthogonal fluorescent labelling of (bio)molecules. Fluorescence properties of both sydnone‐ and pyrazole‐cyanines are described, as well

Development of highly-multiplexed SNP arrays in Maritime pine for multi-objective genetic applications

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Synthesis of Sydnonimines from Sydnones and their use for bioorthogonal release of isocyanates in cells

International audienceIn this article, we report the synthesis of sydnonimines fromsydnones and their use as dipoles for fast click-and-releasereactions. The process relies on nucleophilic aromaticsubstitution of aliphatic and aromatic amines with triflatedsydnones. This new methodology allowed the preparation offunctionalised sydnonimines probes otherwise difficult toprepare. Those probes were then used to release a drug and afluorescent aromatic isocyanate inside living cell

Linkage and association mapping for two major traits used in the maritime pine breeding program : Height growth and stem straightness

Background Increasing our understanding of the genetic architecture of complex traits, through analyses of genotype-phenotype associations and of the genes/polymorphisms accounting for trait variation, is crucial, to improve the integration of molecular markers into forest tree breeding. In this study, two full-sib families and one breeding population of maritime pine were used to identify quantitative trait loci (QTLs) for height growth and stem straightness, through linkage analysis (LA) and linkage disequilibrium (LD) mapping approaches. Results The populations used for LA consisted of two unrelated three-generation full-sib families (n = 197 and n = 477). These populations were assessed for height growth or stem straightness and genotyped for 248 and 217 markers, respectively. The population used for LD mapping consisted of 661 founders of the first and second generations of the breeding program. This population was phenotyped for the same traits and genotyped for 2,498 single-nucleotide polymorphism (SNP) markers corresponding to 1,652 gene loci. The gene-based reference genetic map of maritime pine was used to localize and compare the QTLs detected by the two approaches, for both traits. LA identified three QTLs for stem straightness and two QTLs for height growth. The LD study yielded seven significant associations (P ô 0.001): four for stem straightness and three for height growth. No colocalisation was found between QTLs identified by LA and SNPs detected by LD mapping for the same trait. Conclusions This study provides the first comparison of LA and LD mapping approaches in maritime pine, highlighting the complementary nature of these two approaches for deciphering the genetic architecture of two mandatory traits of the breeding program.</p