Mice carrying a complete deletion of the talin2 coding sequence are viable and fertile

AbstractMice homozygous for several Tln2 gene targeted alleles are viable and fertile. Here we show that although the expression of talin2 protein is drastically reduced in muscle from these mice, other tissues continue to express talin2 albeit at reduced levels. We therefore generated a Tln2 allele lacking the entire coding sequence (Tln2cd). Tln2cd/cd mice were viable and fertile, and the genotypes of Tln2cd/+ intercrosses were at the expected Mendelian ratio. Tln2cd/cd mice showed no major difference in body mass or the weight of the major organs compared to wild-type, although they displayed a mildly dystrophic phenotype. Moreover, Tln2cd/cd mouse embryo fibroblasts showed no obvious defects in cell adhesion, migration or proliferation. However, the number of Tln2cd/cd pups surviving to adulthood was variable suggesting that such mice have an underlying defect

An intergenic non-coding RNA promoter required for histone modifications in the human ß-globin chromatin domain

Transcriptome analyses show a surprisingly large proportion of the mammalian genome is transcribed; much more than can be accounted for by genes and introns alone. Most of this transcription is non-coding in nature and arises from intergenic regions, often overlapping known protein-coding genes in sense or antisense orientation. The functional relevance of this widespread transcription is unknown. Here we characterize a promoter responsible for initiation of an intergenic transcript located approximately 3.3 kb and 10.7 kb upstream of the adult-specific human ß-globin genes. Mutational analyses in ß-YAC transgenic mice show that alteration of intergenic promoter activity results in ablation of H3K4 di- and tri-methylation and H3 hyperacetylation extending over a 30 kb region immediately downstream of the initiation site, containing the adult [delta]- and ß-globin genes. This results in dramatically decreased expression of the adult genes through position effect variegation in which the vast majority of definitive erythroid cells harbor inactive adult globin genes. In contrast, expression of the neighboring [epsilon]- and [gamma]-globin genes is completely normal in embryonic erythroid cells, indicating a developmentally specific variegation of the adult domain. Our results demonstrate a role for intergenic non-coding RNA transcription in the propagation of histone modifications over chromatin domains and epigenetic control of ß-like globin gene transcription during development

CARACTERISATION DU CENTRE D'INACTIVATION DU CHROMOSOME X (CARTOGRAPHIE PHYSIQUE COMPAREE ET ANALYSE FONCTIONNELLE PAR TRANSGENESE DE YACS)

L'INACTIVATION DU CHROMOSOME X, PROCESSUS DE COMPENSATION DE DOSE RENCONTRE CHEZ LES MAMMIFERES, CONSISTE EN UNE EXTINCTION TRANSCRIPTIONNELLE DE LA PLUPART DES GENES DE L'UN DES DEUX CHROMOSOMES X AU HASARD CHEZ LA FEMELLE. SA MISE EN PLACE REQUIERT LA PRESENCE EN CIS DU CENTRE D'INACTIVATION, XIC (X INACTIVATION CENTER). LE GENE XIST (X INACTIVE SPECIFIC TRANSCRIPT) LOCALISE DANS LE XIC PRODUIT UN GRAND ARN NON CODANT COUVRANT ENTIEREMENT LE CHROMOSOME X INACTIF DANS LES TISSUS SOMATIQUES DE LA FEMELLE. CE GENE EST ESSENTIEL A L'INITIATION DE L'INACTIVATION. NEANMOINS, UNE DELETION DE 65KB EN 6 DE XIST A EGALEMENT SUGGERE L'IMPORTANTE FONCTION DE SEQUENCES AVALES DANS LA REGULATION DE L'EXPRESSION DE XIST. L'ORGANISATION DE LA REGION XIC SEMBLE COMPLEXE PUISQUE L'ORIENTATION DU GENE XIST ET DE DEUX GENES SITUES EN 3 EST INVERSEE PAR RAPPORT A L'ORDRE GENERAL DE LA REGION DE SYNTENIE ENTRE L'HOMME ET LA SOURIS. LE PREMIER OBJECTIF DE CE TRAVAIL A ETE L'ASSEMBLAGE D'UN CONTIG DE YACS COUVRANT ENVIRON 2 MB AUTOUR DE XIST. LA PRESENCE SUR LE CONTIG DU GENE XPCT (X-LINKED PEST-CONTAINING TRANSMEMBRANE TRANSPORTER) ET LA COMPARAISON AVEC LA SITUATION DEJA CONNUE CHEZ L'HOMME, A PERMIS DE MONTRER QUE LA REGION XPCT-XIST A SUBI DES REMANIEMENTS COMPLEXES AU COURS DE L'EVOLUTION. CETTE ANALYSE STRUCTURALE A CONDUIT A DISCUTER LA DEFINITION DE LA REGION CANDIDATE POUR LE XIC MURIN ET LES LIMITES DE L'APPROCHE DE CARTOGRAPHIE COMPAREE DANS L'ETUDE DES LOCI COMPLEXES. LA DEUXIEME PARTIE DE CE TRAVAIL A CONSISTE EN UNE ETUDE FONCTIONNELLE, PAR DELETION ET TRANSGENESE DE YACS, DANS DES CELLULES SOUCHES EMBRYONNAIRES MURINES, D'UNE SEQUENCE CANDIDATE POTENTIELLEMENT IMPLIQUEE DANS LES FONCTIONS DU XIC. L'ENSEMBLE DES RESULTATS OBTENUS A PERMIS D'ETABLIR QUE DXPAS34, ELEMENT REPETE SITUE 15 KB EN 3 DE XIST, JOUE UN ROLE FONDAMENTAL DANS LA REGULATION DE L'ACTIVITE TRANSCRIPTIONNELLE D'UNE REGION DE 50 KB INCLUANT XIST, ET DANS LES PROCESSUS DE COMPTAGE ET/OU DE CHOIX, LIES A L'INITIATION DE L'INACTIVATION. LE ROLE DE CET ELEMENT DANS LA REGULATION DE LA STRUCTURE CHROMATINIENNE DU LOCUS XIST AINSI QUE DANS LES FONCTIONS D'INACTIVATION IN VIVO, A ETE ABORDE PAR IMMUNOPRECIPITATION DE LA CHROMATINE ET CREATION DE SOURIS CHIMERES.PARIS-BIUSJ-Thèses (751052125) / SudocCentre Technique Livre Ens. Sup. (774682301) / SudocPARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

A Non-Invasive Droplet Digital PCR (ddPCR) Assay to Detect Paternal CFTR Mutations in the Cell-Free Fetal DNA (cffDNA) of Three Pregnancies at Risk of Cystic Fibrosis via Compound Heterozygosity.

Non-invasive prenatal diagnosis (NIPD) makes use of cell-free fetal DNA (cffDNA) in the mother's bloodstream as an alternative to invasive sampling methods such as amniocentesis or CVS, which carry a 0.5-1% risk of fetal loss. We describe a droplet digital PCR (ddPCR) assay designed to inform the testing options for couples whose offspring are at risk of suffering from cystic fibrosis via compound heterozygosity. By detecting the presence or absence of the paternal mutation in the cffDNA, it is possible to predict whether the fetus will be an unaffected carrier (absence) or whether further invasive testing is indicated (presence).We selected a family in which the parents were known to carry different mutated CFTR alleles as our test system. NIPD was performed for three of their pregnancies during the first trimester (at around 11-12 weeks of gestation). Taqman probes were designed against an amplicon in exon 11 of the CFTR gene, to quantify the proportion of mutant (ΔF508-MUT; FAM) and normal (ΔF508-NOR; VIC) alleles at position c.1521_1523 of the CFTR gene.The assay correctly and unambiguously recognized the ΔF508-MUT CFTR allele in the cffDNA of all three proband fetuses and none of the six unaffected control fetuses. In conclusion, the Bio-Rad QX100 was found to be a cost-effective and technically undemanding platform for designing bespoke NIPD assays

Example ddPCR-NIPD assay results for male proband (PM1), male control (CM3) and female proband (PF3).

<p>Three replicates (rep1, rep2 and rep3) are shown for each sample, separated by dashed vertical lines. Each event represents a separate droplet. ZFX and ΔF508-NOR were labelled with VIC and detected at 535–555 nm (positive droplets are shown in green). ZFY and ΔF508-MUT were labelled with FAM and detected at 510–530 nm (positive droplets are shown in blue). Grey points represent negative droplets. As expected, ZFY positive droplets are almost absent in the female sample and ΔF508-MUT positive droplets are absent in the healthy control.</p

Pedigree of a family likely to benefit from non-invasive prenatal diagnosis.

<p>The parents each carried a different mutation, putting offspring at risk of compound heterozygosity. Three proband pregnancies (P) were tested, as well as three male and three female unrelated control pregnancies (not shown). Non-invasive testing was performed at around week 11–12 of gestation.</p

Proportions of ΔF508-MUT (A) and male (B) DNA found in cffDNA.

<p>Results are shown for two male probands (PM1, PM3), a female proband (PF2), three control male pregnancies (CM1, CM2, CM3) and three control female pregnancies (CF4, CF5, CF6). Male and female samples are labelled with the (♂) and (♀) symbols respectively. The suggested threshold between a positive and negative result is represented by a grey dashed line at 0.25x the mean of the known positive samples.</p

PLATELETS ACTIVITY OF A NEW ENGINEERED RECOMBINANT COLLAGEN PEPTIDE

International audienceCoupled transfers of aroma compounds and water vapour were investigated by varying the relative humidity gradient of storage (50% or 90%) of two paper packaging at 25 degrees C. These papers differed in their coating surface: both were identically impregnated then supercalendered, and only one was twice coated on both sides with a synthetic barrier substance. Permeability and solubility coefficients were determined. The coating treatment was more effective to decrease the permeabilities of water vapour and ethyl ester than the effect of RH. On the contrary, the RH modified the water content of the treated papers and affected more strongly their permeability and solubility to cis-3-hexenol and benzaldehyde. The cis-3-hexenol transfer through the non-coated paper and the benzaldehyde transfer through the coated paper decreased due to a probable competition with sorbed water although it increases for the cis-3-hexenol of a plasticisation phenomenon