Etude du rôle du complexe de polyadénylation dans la formation de l'extrémité 3' de tous les transcrits de l'ARN polymérase II chez saccharomyces cerevisiae (caractérisation de trois nouvelles sous-unités, les protéines Mpelp, Ptilp et Ref2p)

L'ARN polymérase II eucaryote transcrit les précurseurs des ARN messagers et des petits ARN non-codants snRNA et snoRNA. La formation de l'extrémité 3' des ARNpm consiste en un clivage et de la polyadénylation de cette extremité. Les sn et snoRNA sont également clivés en 3' mais ne sont pas polydénylés. Chez la levure, un complexe multiprotéique composé des facteurs de clivage et de polyadénylation CFI (Clevage Factor I), CFC (Clevage and Polyadenylation Factor) et la protéine Nab2p permet de reconstituer in vitro la maturation 3' des ARNpm. Il aété montré que des mutations affectant des sous-unités du CFI généraient un début de maturation 3' des pr-snRNA et pre-snoRNA transcrits indépendamment. Aucun des mutants du CPF testés jusqu'à présent ne montrait ce défaut, suggérant que le complexe CPF ne participerait pas à la maturation 3' des petits ARN non-codants. Afin de caractériser de nouveaux facteurs interagissant avec le complexe de polyadenylation, nous avons recherché des suppresseurs multicopie et extragéniques de la thermosensibilité du mutant du CFI pcf11-2, isolant ainsi deux nouveaux gènes : MPEI (Mutant pcfII extragenic suppresseur 1) et PTII (PtaI interacting protein I). Le gène REF2 a été sélectionné comme codant une protéine interagissant avec la protéine PtiIp en double hybride. Nous avons montré que ces trois gènes essentiels codent des sous-unités du CPF. La protéine Mpe1p est essentielle à la reconstitution in vitro de la maturation 3' des ARNpm et serait impliquée dans le coopérativité des complexes CFI et CPF requise pour ce processus. Les protéines Pti1p et Ref2p sont nécessaires à la maturation 3' des snoRNA transcrits indépendamment, suggérant que le complexe CPF participerait aussi à ce processus, au cours duquel Pti1p et Ref2p seraient impliquées dans le découplage du clivage et de la polyadénylation. Tous les trancrits de l'ARN pol II seraient donc maturés par un même processus grâce à des facteurs commun recrutés par l'ARN pol II.VERSAILLES-BU Sciences et IUT (786462101) / SudocSudocFranceF

Persistent left cranial vena cava causing oesophageal obstruction and consequent megaoesophagus in a dog

International audienceA 2-month-old Brittany spaniel dog was presented for persistent regurgitation, first observed soon after weaning. Clinical examination and diagnostic imaging suggested megaoesophagus associated with a vascular ring anomaly. The normal location of the trachea on the X-ray was not consistent with a persistent right aortic arch. Postmortem examination revealed a persistent left cranial vena cava that formed a non-elastic fibrous band enclosing the oesophagus and trachea, and causing constriction of the oesophagus. This uncommon congenital vascular defect has never previously been associated with megaoesophagus in the dog

Pti1p and Ref2p found in association with the mRNA 3′ end formation complex direct snoRNA maturation

Eukaryotic RNA polymerase II transcribes precursors of mRNAs and of non-protein-coding RNAs such as snRNAs and snoRNAs. These RNAs have to be processed at their 3′ ends to be functional. mRNAs are matured by cleavage and polyadenylation that require a well-characterized protein complex. Small RNAs are also subject to 3′ end cleavage but are not polyadenylated. Here we show that two newly identified proteins, Pti1p and Ref2p, although they were found associated with the pre-mRNA 3′ end processing complex, are essential for yeast snoRNA 3′ end maturation. We also provide evidence that Pti1p probably acts by uncoupling cleavage and polyadenylation, and functions in coordination with the Nrd1p-dependent pathway for 3′ end formation of non-polyadenylated transcripts

Bidirectional promoters generate pervasive transcription in yeast

Genome-wide pervasive transcription has been reported in many eukaryotic organisms, revealing a highly interleaved transcriptome organization that involves hundreds of previously unknown non-coding RNAs. These recently identified transcripts either exist stably in cells (stable unannotated transcripts, SUTs) or are rapidly degraded by the RNA surveillance pathway (cryptic unstable transcripts, CUTs). One characteristic of pervasive transcription is the extensive overlap of SUTs and CUTs with previously annotated features, which prompts questions regarding how these transcripts are generated, and whether they exert function. Single-gene studies have shown that transcription of SUTs and CUTs can be functional, through mechanisms involving the generated RNAs or their generation itself. So far, a complete transcriptome architecture including SUTs and CUTs has not been described in any organism. Knowledge about the position and genome-wide arrangement of these transcripts will be instrumental in understanding their function. Here we provide a comprehensive analysis of these transcripts in the context of multiple conditions, a mutant of the exosome machinery and different strain backgrounds of Saccharomyces cerevisiae. We show that both SUTs and CUTs display distinct patterns of distribution at specific locations. Most of the newly identified transcripts initiate from nucleosome-free regions (NFRs) associated with the promoters of other transcripts (mostly protein-coding genes), or from NFRs at the 3' ends of protein-coding genes. Likewise, about half of all coding transcripts initiate from NFRs associated with promoters of other transcripts. These data change our view of how a genome is transcribed, indicating that bidirectionality is an inherent feature of promoters. Such an arrangement of divergent and overlapping transcripts may provide a mechanism for local spreading of regulatory signals-that is, coupling the transcriptional regulation of neighbouring genes by means of transcriptional interference or histone modification