Antiprion drugs 6-aminophenanthridine and guanabenz reduce PABPN1 toxicity and aggregation in oculopharyngeal muscular dystrophy

Oculopharyngeal muscular dystrophy (OPMD) is an adult-onset syndrome characterized by progressive degeneration of specific muscles. OPMD is caused by extension of a polyalanine tract in poly(A) binding protein nuclear 1 (PABPN1). Insoluble nuclear inclusions form in diseased muscles. We have generated a Drosophila model of OPMD that recapitulates the features of the disorder. Here, we show that the antiprion drugs 6-aminophenanthridine (6AP) and guanabenz acetate (GA), which prevent formation of amyloid fibers by prion proteins in cell models, alleviate OPMD phenotypes in Drosophila, including muscle degeneration and nuclear inclusion formation. The large ribosomal RNA and its activity in protein folding were recently identified as a specific cellular target of 6AP and GA. We show that deletions of the ribosomal DNA locus reduce OPMD phenotypes and act synergistically with sub-effective doses of 6AP. In a complementary approach, we demonstrate that ribosomal RNA accelerates in vitro fibril formation of PABPN1 N-terminal domain. These results reveal the conserved role of ribosomal RNA in different protein aggregation disorders and identify 6AP and GA as general anti-aggregation molecules

Étude de la synthèse et de la maturation des précurseurs dicistroniques ARNt-snoARN chez Arabidopsis thaliana

Les C/D snoARN sont des petits ARN nucléolaires présents chez tous les eucaryotes et les archae qui guident la méthylation de nucléotides sur les ARNr ou d autres ARN cellulaires. In vivo les snoARN sont complexés avec quatre protéines dont la fibrillarine responsable de la méthylation. Chez les plantes les tsnoARN sont un nouveau type d organisation génomique de snoARN, dont l expression génère un précurseur dicistronique ARNt-snoARN. Ceci suggère un nouveau mode de maturation de snoARN qui impliquerait des facteurs de la biogenèse des ARNt. Le but de ce travail est d identifier l endonucléase responsable de la maturation du prétsnoARN et de rechercher d autres facteurs impliqués dans la synthèse et la maturation des tsnoARN. Lors de ce travail, nous avons montré que les tsnoARN sont répandus chez les plantes et que leurs transcriptions dépendaient de l ARN polymérase III qui synthétise les ARNt. In vivo des mutations de l ARNt du tsnoARN comme l inactivation des gènes de la tRNASEZ et d un homologue de RNT1P n affectent pas l expression du snoARN. Pour contourner les problèmes éventuels de redondance génique, nous avons donc créé un système in vitro qui reproduit la maturation du prétsnoARN basé sur l utilisation d extraits nucléaires d inflorescences de choux fleur. Nous avons alors montré que la maturation du prétsnoARN s effectue par deux voies distinctes dont une seule implique la tRNaseZ. Enfin l étude de prétsnoARN mutés dans les boîtes C ou D impliqués dans la reconnaissance des protéines du complexe snoRNP suggère que le système in vitro mis au point reproduit l assemblage de ces particulesSmall nucleolar RNAs of class C/D (C/D snoRNAs) represent a large class of small non coding RNAs guiding methylation of ribosomal RNAs and other cellular RNAs. In plants more than a hundred genes have been identified in Arabiodopsis thaliana. Most of them are organised in clusters and expressed as polycistronic precursors that must be processed by endonuclease and exonuclease to liberate the snoRNA. A variation to these, unique to plants, are the dicistronic glycine tRNA-C/D snoRNA genes.In this work, we searched to identify the endonuclease responsible for tsnoRNA maturation and more factors involved in the tsnoRNA synthesis and maturation.To study the transcription and maturation pathway in vivo, we created transgenic plants expressing mutants of these tsnoRNAs. We show that dicistronic tRNA-snoRNA precursor is synthesised by RNA polymerase III system and is further processed to produce the tRNA and the snoRNA separated products. To study this step, we developed nuclear extracts from cauliflower inflorescence that accurately process the dicistronic tsnoRNA precursor in vitro. In addition we have evidence that these extracts allows assembly of the small nucleolar ribonucleoprotein (snoRNP) that is essential for snoRNA stability and activity.PERPIGNAN-BU Sciences (661362101) / SudocSudocFranceF

A Pre-mRNA Degradation Pathway that Selectively Targets Intron-Containing Genes Requires the Nuclear Poly(A)-Binding Protein

General discard pathways eliminate unprocessed and irregular pre-mRNAs to control the quality of gene expression. In contrast to such general prem-RNA decay, we describe here a nuclear pre-mRNA degradation pathway that controls the expression of select intron-containing genes. We show that the fission yeast nuclear poly(A)-binding protein, Pab2, and the nuclear exosome subunit, Rrp6, are the main factors involved in this polyadenylation-dependent pre-mRNA degradation pathway. Transcriptome analysis and intron swapping experiments revealed that inefficient splicing is important to dictate susceptibility to Pab2-dependent pre-mRNA decay. We also show that negative splicing regulation can promote the poor splicing efficiency required for this pre-mRNA decay pathway, and in doing so, we identified a mechanism of cross-regulation between paralogous ribosomal proteins through nuclear pre-mRNA decay. Our findings unveil a layer of regulation in the nucleus in which the turnover of specific pre-mRNAs, besides the turnover of mature mRNAs, is used to control gene expression

Early exposure to food contaminants reshapes maturation of the human brain-gut-microbiota axis

International audienc

Food Contaminants Effects on an In Vitro Model of Human Intestinal Epithelium

International audienc

Complexification of In Vitro Models of Intestinal Barriers, A True Challenge for a More Accurate Alternative Approach

The use of cell models is common to mimic cellular and molecular events in interaction with their environment. In the case of the gut, the existing models are of particular interest to evaluate food, toxicants, or drug effects on the mucosa. To have the most accurate model, cell diversity and the complexity of the interactions must be considered. Existing models range from single-cell cultures of absorptive cells to more complex combinations of two or more cell types. This work describes the existing solutions and the challenges that remain to be solved

Arabidopsis Encodes Four tRNase Z Enzymes1[W][OA]

Functional transfer RNA (tRNA) molecules are a prerequisite for protein biosynthesis. Several processing steps are required to generate the mature functional tRNA from precursor molecules. Two of the early processing steps involve cleavage at the tRNA 5′ end and the tRNA 3′ end. While processing at the tRNA 5′ end is performed by RNase P, cleavage at the 3′ end is catalyzed by the endonuclease tRNase Z. In eukaryotes, tRNase Z enzymes are found in two versions: a short form of about 250 to 300 amino acids and a long form of about 700 to 900 amino acids. All eukaryotic genomes analyzed to date encode at least one long tRNase Z protein. Of those, Arabidopsis (Arabidopsis thaliana) is the only organism that encodes four tRNase Z proteins, two short forms and two long forms. We show here that the four proteins are distributed to different subcellular compartments in the plant cell: the nucleus, the cytoplasm, the mitochondrion, and the chloroplast. One tRNase Z is present only in the cytoplasm, one protein is found exclusively in mitochondria, while the third one has dual locations: nucleus and mitochondria. None of these three tRNase Z proteins is essential. The fourth tRNase Z protein is present in chloroplasts, and deletion of its gene results in an embryo-lethal phenotype. In vitro analysis with the recombinant proteins showed that all four tRNase Z enzymes have tRNA 3′ processing activity. In addition, the mitochondrial tRNase Z proteins cleave tRNA-like elements that serve as processing signals in mitochondrial mRNA maturation

Prevention of Adult Colitis by Oral Ferric Iron in Juvenile Mice Is Associated with the Inhibition of the Tbet Promoter Hypomethylation and Gene Overexpression

International audienceIron is an essential nutrient needed for physiological functions, particularly during the developmental period of the early childhood of at-risk populations. The purpose of this study was to investigate, in an experimental colitis, the consequences of daily oral iron ingestion in the early period on the inflammatory response, the spleen T helper (Th) profiles and the associated molecular mechanisms. Juvenile mice orally received microencapsulated ferric iron or water for 6 weeks. On adult mice, we induced a sham or experimental trinitrobenzene sulfonic acid (TNBS) moderate colitis during the last week of the experiment before sacrificing the animals 7 days later. The severity of the gut inflammation was assessed by macroscopic damage scores (MDS) and the myeloperoxidase activity (MPO). Th profiles were evaluated by the examination of the splenic gene expression of key transcription factors of the Th differentiation (Tbet, Gata3, Foxp3 and RORγ) and the methylation of their respective promoter. While TNBS-induced colitis was associated with a change of the Th profile (notably an increase in the Tbet/Gata3 ratio in the spleen), the colitis-inhibition induced by ferric iron was associated with a limitation of the splenic Th profiles perturbation. The inhibition of the splenic Tbet gene overexpression was associated with an inhibition of promoter hypomethylation. In summary, mice treated by long-term oral ferric iron in the early period of life exhibited an inhibition of colitis associated with the inhibition of the splenic Tbet promoter hypomethylation and gene overexpression