A stereodivergent asymmetric approach to difluorinated aldonic acids

A (bromodifluoromethyl)alkyne has been deployed in a stereoselective route to difluorinated aldonic acid analogues, in which a Sharpless asymmetric dihydroxylation reaction and diastereoisomer separation set the stage for phenyl group oxidation

Lack of maternal Heat Shock Factor 1 results in multiple cellular and developmental defects, including mitochondrial damage and altered redox homeostasis, and leads to reduced survival of mammalian oocytes and embryos

AbstractHeat Shock Factor 1 (HSF1) is a transcription factor whose loss of function results in the inability of Hsf1−/− females to produce viable embryos, as a consequence of early developmental arrest. We previously demonstrated that maternal HSF1 is required in oocytes to regulate expression of chaperones, in particular Hsp90α, and is essential for the progression of meiotic maturation. In the present work, we used comparative morphological and biochemical analytic approaches to better understand how Hsf1−/− oocytes undergo irreversible cell death. We found that the metaphase II arrest in mature oocytes, cortical granule exocytosis and formation of pronuclei in zygotes were all impaired in Hsf1−/− mutants. Although oogenesis generated fully grown oocytes in follicles, intra-ovarian Hsf1−/− oocytes displayed ultrastructural abnormalities and contained dysfunctional mitochondria as well as elevated oxidant load. Finally, the apoptotic effector, caspase-3, was activated in most mutant oocytes and embryos, reflecting their commitment to apoptosis. In conclusion, our study shows that early post-ovulation events are particularly sensitive to oxidant insult, which abrogates the developmental competence of HSF1-depleted oocytes. They also reveal that Hsf1 knock-out mice constitute a genetic model that can be used to evaluate the importance of redox homeostasis in oocytes

Eph:ephrin-B1 forward signaling controls fasciculation of sensory and motor axons

AbstractAxon fasciculation is one of the processes controlling topographic innervation during embryonic development. While axon guidance steers extending axons in the accurate direction, axon fasciculation allows sets of co-extending axons to grow in tight bundles. The Eph:ephrin family has been involved both in axon guidance and fasciculation, yet it remains unclear how these two distinct types of responses are elicited. Herein we have characterized the role of ephrin-B1, a member of the ephrinB family in sensory and motor innervation of the limb. We show that ephrin-B1 is expressed in sensory axons and in the limb bud mesenchyme while EphB2 is expressed in motor and sensory axons. Loss of ephrin-B1 had no impact on the accurate dorso-ventral innervation of the limb by motor axons, yet EfnB1 mutants exhibited decreased fasciculation of peripheral motor and sensory nerves. Using tissue-specific excision of EfnB1 and in vitro experiments, we demonstrate that ephrin-B1 controls fasciculation of axons via a surround repulsion mechanism involving growth cone collapse of EphB2-expressing axons. Altogether, our results highlight the complex role of Eph:ephrin signaling in the development of the sensory-motor circuit innervating the limb

Genome restructuring in mouse embryos during reprogramming and early development

AbstractAlthough a growing number of studies investigates functional genome organization in somatic cell nuclei, it is largely unknown how mammalian genome organization is established during embryogenesis. To address this question, we investigated chromocenter formation and the peculiar arrangements of chromosome domains in early mouse embryos. At the one-cell stage, we observed characteristic arrangements of chromosomes and chromocenter components. Subsequently, starting with the burst of zygotic genome transcription major rearrangements led to the establishment of somatic type chromocenters with a defined spatio-temporal organization. These processes appeared to be completed at the blastocyst stage with the onset of cell differentiation. During the same developmental period, a fraction of pericentric heterochromatin that was late replicating in the first cycle underwent switches in replication timing, spatial organization and epigenetic marks. Cloning experiments revealed that the genome organization typical for more advanced stages was quickly reverted into the one-cell stage-specific form after nuclear transfer, supporting the idea that reprogramming associated genome remodeling in normal and cloned embryos is determined by cytoplasmic factors. Together, the results suggest that distinct but characteristic forms of nuclear genome organization are required for genome reprogramming in early embryos and for proper regulation of differential gene expression patterns at later stages

Oocyte–Targeted Deletion Reveals That Hsp90b1 Is Needed for the Completion of First Mitosis in Mouse Zygotes

Hsp90b1 is an endoplasmic reticulum (ER) chaperone (also named Grp94, ERp99, gp96,Targ2, Tra-1, Tra1, Hspc4) (MGI:98817) contributing with Hspa5 (also named Grp78, BIP) (MGI:95835) to protein folding in ER compartment. Besides its high protein expression in mouse oocytes, little is known about Hsp90b1 during the transition from oocyte-to-embryo. Because the constitutive knockout of Hsp90b1 is responsible for peri-implantation embryonic lethality, it was not yet known whether Hsp90b1 is a functionally important maternal factor.To circumvent embryonic lethality, we established an oocyte-specific conditional knockout line taking advantage of the more recently created floxed Hsp90b1 line (Hsp90b1(flox), MGI:3700023) in combination with the transgenic mouse line expressing the cre recombinase under the control of zona pellucida 3 (ZP3) promoter (Zp3-cre, MGI:2176187). Altered expression of Hsp90b1 in growing oocytes provoked a limited, albeit significant reduction of the zona pellucida thickness but no obvious anomalies in follicular growth, meiotic maturation or fertilization. Interestingly, mutant zygotes obtained from oocytes lacking Hsp90b1 were unable to reach the 2-cell stage. They exhibited either a G2/M block or, more frequently an abnormal mitotic spindle leading to developmental arrest. Despite the fact that Hspa5 displayed a similar profile of expression as Hsp90b1, we found that HSPA5 and HSP90B1 did not fully colocalize in zygotes suggesting distinct function for the two chaperones. Consequently, even if HSPA5 was overexpressed in Hsp90b1 mutant embryos, it did not compensate for HSP90B1 deficiency. Finally, further characterization of ER compartment and cytoskeleton revealed a defective organization of the cytoplasmic region surrounding the mutant zygotic spindle.Our findings demonstrate that the maternal contribution of Hsp90b1 is critical for the development of murine zygotes. All together our data indicate that Hsp90b1 is involved in unique and specific aspects of the first mitosis, which brings together the maternal and paternal genomes on a single spindle

Conception, synthèse et évaluation d'une nouvelle famille d'agents de fluoration électrophile énantiosélective issue des alcaloïdes du Quinquina

Ce travail focalise sur l'obtention de composés carbonylés a-fluorés énantiomériquement enrichis par fluoration électrophile énantiosélective. D'abord, nous avons conçu et synthétisé une nouvelle famille d'agents de fluoration électrophile énantiosélective dérivés des alcaloïdes du quinquina. Leur synthèse rapide et efficace permet de les obtenir sous forme d'ammoniums quaternaires N-fluorés avec de bons rendements par réaction de transfluoration. Cette synthèse mise au point, nous avons tiré profit de la structure polyfonctionnelle des alcaloïdes pour synthétiser de nombreux dérivés afin d'avoir une large panoplie de structures. Ces nouveaux agents de fluoration ont été engagés dans la fluoration d'énolates et d'énoxysilanes. Le produits fluorés correspondants ont été obtenus avec des rendements quantitatifs et des énantiosélections intéressantes. En effet, on a pu découvrir que la chimie des énolates donnaient des énantiosélections moyennes de l'ordre de 50 %, alors que les énoxysilanes ont permis d'obtenir les a-fluoro-cétones désirées avec de hautes énantiosélections (jusqu'à 87 %. Puis, grâce aux différents substrats étudiés sous forme d'énolates ou d'énoxysilanes, et grâce aux différents agents de fluoration synthétisés, nous avons réalisé une étude Relation Structure-Enantiosélectivité. Ainsi on a élaboré un modèle de reconnaissance moléculaire et pu comprendre les énantiosélections observées pour les substrats étudiés. Enfin notre travail a consisté en l'application de notre méthodologie à la synthèse d'une molécule fluorée d'intérêt thérapeutique. Après la synthèse de l'intermédiaire clef désiré, nos agents de fluoration ont servi pour la fluoration électrophile énantiosélective tardive de cet intermédiaire. La molécule fluorée a ainsi été obtenue avec un excès énantiomérique de 76 %.This work is focused on the synthesis of chiral a-fluoro-carbonyl compounds by electrophilic enantioselective fluorination. We designed and synthesized a novel class of electrophilic enantioselective fluorinating agents derived from cinchona alkaloïds. Their straightforward synthesis allow us to obtain them as N-fluoro ammonium salts in high yields by means of transfer-fluorination. Then, we took advantage of the polyfonctionnal structures of the alkaloïds in order to synthesize numerous derivatives providing a wide range of differents structures. These new fluorinating agents were used in the fluorination of enolates and silyl enol ethers. The corresponding fluorinated compounds are obtained in quantitative yields and interesting enantiomeric excesses. Indeed, we demonstrated that enolates give intermediates enantioselections (ca. 50 %), but silyl enol ethers give very good enantiomeric excesses (up to 87 %). Then, we realised a Structure-Enantioselectivity Relationship study which allows us to elaborate a model of molecular recognition, and to understand the enantioselection observed in the enantioselective fluorination. Finally, we apply our methodology to the synthesis of a fluorinated molecule of biological interest. Our fluorinating agents were used in the late electrophilic enantioselective fluorination and the target fluorinated product was obtained with an enantiomeric excess of 76 %.ROUEN-BU Sciences (764512102) / SudocSudocFranceF

Enantioselective electrophilic fluorination in ionic liquids

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Efnb2 haploinsufficiency induces early gap junction plaque disassembly and endocytosis in the cochlea.

Chromosome 13q deletions encompassing EFNB2, which encodes the transmembrane protein ephrin-B2, are likely to cause syndromic forms of sensorineural hearing loss of unclear origin. Thus, unravelling the pathogenic mechanisms could help to improve therapeutic strategies. In the cochlea, adjacent non-sensory epithelial cells are connected via gap junction channels, the activity of which is critical to maintain cochlear homeostasis. Here we show that ephrin-B2 promotes the assembly of connexin 30 (Cx30) gap junction plaques (GJPs) between adjacent non-sensory Deiters' cells. An in situ proximity ligation assay revealed that ephrin-B2 preferentially interacts with Cx30 in the periphery of the GJPs, i.e. where newly synthesized connexin hemichannels accrue to the GJP. Moreover, we observed that heterozygous mice encoding an Efnb2 null allele display excessive clathrin-mediated internalization of Cx30 GJPs in early postnatal stages. Finally, an in vitro organotypic assay revealed that ectopic activation of ephrin-B2 reverse signalling promotes the internalization of Cx30 GJPs. These data argue in favor of a cell-autonomous, Eph receptor-independent role of ephrin-B2 in the assembly of Cx30 GJPs. According to recent observations, early GJP degradation could certainly play a role in the pathogenic process leading to progressive sensorineural hearing loss due to Efnb2/EFNB2 haploinsufficiency

A direct and rapid route to α,α-difluoroacylsilanes from trifluoroethanol

α,α-Difluoroacylsilanes were synthesised directly from a range of allyl ethers of trifluoroethanol via dehydrofluorination/metallation procedures, followed by thermal [3,3]-sigmatropic rearrangement of the intermediate vinylmetals. The scope and limitations of ether synthesis and dehydrofluorination/metallation are described