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In eukaryotic cells, aberrant mRNPs with processing and packaging defects are targeted co- transcriptionally by a surveillance system that triggers their nuclear retention and ultimately the degradation of their mRNA component by the 3’-5’ activity of the exosome-associated exoribonuclease Rrp6 along with its associated factors Rrp47 and Mpp6. This mRNP quality control process is stimulated by the NNS complex (Nrd1-Nab3-Sen1), which otherwise mediates termination, processing and decay of ncRNAs and the process involves additionally the exosome co- activator TRAMP complex (Trf4-Air2-Mtr4). Here, we describe a genome-wide approach to visualize the dynamic recruitment of these quality control components over the yeast chromosomes upon global perturbation of mRNP biogenesis by the RNAdependent helicase/translocase activity of the bacterial Rho factor. The method provides valuable information on how the surveillance system coordinates with the transcription machinery in order to detect faulty events during perturbation of mRNP biogenesis. In addition, our overview shows that the assembly of the quality control components for the affected mRNA genes takes place at the expense of their commitment to be recruited to ncRNAs genomic features, which in turn leads to termination and processing defects of ncRNAs.

Over the past decade, advances in high-throughput RNA-sequencing methods have enabled deep insight into complexity of Saccharomyces cerevisiae transcriptome, revealing a great number of RNA transcripts, that are not translated into proteins and are therefore referred to as long non-coding RNAs (lncRNAs). In yeast, based on their sensitivity to different RNA- degradation pathways or conditions in which they are transcribed, this class of RNAs includes stable unannotated transcripts (SUTs), cryptic unstable transcripts (CUTs), meiotic unannotated transcripts (MUTs) and Xrn1-sensitive unstable transcripts (XUTs). Several classes of lncRNAs are targeted by the conserved 5’-3’ exoribonuclease Rrp6. This enzyme is associated with the exosome, a multi-subunit complex important for RNA processing and stability. Some lncRNAs are known to be implicated in various cellular processes such as gene expression, nuclear organization and chromatin structure, however vast majority of these transcripts is still unexplored. High-throughput RNA-sequencing methods discovered a plethora of antisense transcripts which are transcribed across genes encoding extracellular proteins, such as genes related to cell wall biogenesis and phosphate metabolism. We are taking advantage of high-throughput RNA-sequencing methods, as well as transcript-specific methods, to gain insight into how the cell wall transcriptome is modulated to support different life cycle phases and tolerate various environmental perturbations. Our preliminary results suggest inclusion of lncRNAs in these processes, acting via modulation of chromatin structure, promoter interference and/or overlapping sense/antisense pairs. The implications of these observations for the expression and stability for lncRNAs, mRNAs and proteins will be discussed.

International audience; This article corrects: The conserved histone deacetylase Rpd3 and the DNA binding regulator Ume6 repress BOI1's meiotic transcript isoform during vegetative growth in Saccharomyces cerevisiae. Vol. 96, Issue 4, 861–874, Article first published online: 21 MAR 2015

The sizes of many transcripts change when yeast cells exit mitosis and enter the meiotic differentiation pathway. These differences were attributed to length variations of the 5’ untranslated regions (UTR) of mRNAs. The function of UTRs in protein translation is well established. However, the mechanism controlling the expression of distinct transcript isoforms during mitotic growth and meiotic development is unknown. Based on tilling array data, we order developmentally regulated transcript isoforms according to their expression at specific stages during meiosis and gametogenesis, as compared to vegetative growth and starvation. We employ regulatory motif prediction, in vivo protein-DNA binding assays, genetic analyses and monitoring of epigenetic amino acid modification patterns to identify a novel role for Rpd3 and Ume6, two components of a histone deacetylase complex already known to repress early meiosis-specific genes in dividing cells, in mitotic repression of meiosis-specific transcript isoforms. Our findings classify developmental stage-specific early, middle and late meiotic transcript isoforms, and they point to a novel HDAC- dependent control mechanism for flexible transcript architecture during cell growth and differentiation. Since Rpd3 is highly conserved and ubiquitously expressed in many tissues, our results are likely relevant for development and disease in higher eukaryotes. The implications of these observations for the expression and stability for lncRNAs, mRNAs and proteins during meiotic development will be discussed.

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Background Environmental factors such as pesticides can cause phenotypic changes in various organisms, including mammals. We studied the effects of the widely used herbicide atrazine (ATZ) on meiosis, a key step of gametogenesis, in male mice. Methods Gene expression pattern was analysed by Gene–Chip array. Genome-wide mapping of H3K4me3 marks distribution was done by ChIP-sequencing of testis tissue using Illumina technologies. RT-qPCR was used to validate differentially expressed genes or differential peaks. Results We demonstrate that exposure to ATZ reduces testosterone levels and the number of spermatozoa in the epididymis and delays meiosis. Using Gene-Chip and ChIP-Seq analysis of H3K4me3 marks, we found that a broad range of cellular functions, including GTPase activity, mitochondrial function and steroid-hormone metabolism, are affected by ATZ. Furthermore, treated mice display enriched histone H3K4me3 marks in regions of strong recombination (double-strand break sites), within very large genes and reduced marks in the pseudoautosomal region of X chromosome. Conclusions Our data demonstrate that atrazine exposure interferes with normal meiosis, which affects spermatozoa production. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-2095-y) contains supplementary material, which is available to authorized users.

5-Fluorouracil (5-FU) has been widely used to treat solid tumors. The drug is known to inhibit enzymes involved in DNA replication (TK) and RNA processing/modification (Rrp6/EXOSC10). However, the molecular mechanism of 5-FU's anti-proliferative activity is not fully understood and 5-FU resistance is a serious clinical problem. We hypothesize that (i) long non- coding RNAs regulated by Rrp6/EXOSC10 are important for 5-FU toxicity and that (ii) elevated levels of the enzyme may confer 5-FU resistance in cancer cells. We compared the transcriptomes of a wild type yeast strain treated with 5-FU to an rrp6 deletion mutant and identified transcripts that respond to the drug, the RRP6 deletion or both. Furthermore, we discovered that RRP6 overexpression confers 5-FU resistance during vegetative growth. Our data offer interesting leads for the discovery of novel protein-coding and non-coding RNAs that may be involved in 5-FU toxicity, and they indicate that elevated levels of Rrp6 compromise 5-FU activity. These results are potentially important for improving 5-FU based chemotherapy.

International audience; In eukaryotic cells, the co-transcriptional mRNA processing and packaging reactions that lead to the formation of export competent messenger ribonucleoprotein particles (mRNPs) are under the surveillance of quality control (QC) steps. Aberrant mRNPs resulting from faulty events are detected by the QC apparatus and retained in the nucleus with ensuing elimination of their mRNA component by the RNA degradation machinery. A decade of biochemical and genetic experiments in yeast allowed the identification of the nuclear degradation machinery including the core exosome and its two associated catalytic subunits Rrp6p and Rrp44p, its cofactors Rrp47p and Mpp6p as well as the activator complex TRAMP. Similarly, studies of the THO-Sub2 complex of the mRNP assembly and export apparatus have provided valuable information on the nuclear retention and degradation of a particular class of aberrant mRNPs. However, a unifying mechanism of action underlying the QC process remains elusive. Here, we review the implementation of a new experimental approach whereby the production of aberrant mRNPs is massively increased upon heterologous expression of the bacterial Rho helicase in yeast. Using this methodology, we have shown that the QC process is coordinated by Nrd1p (a component of the early termination complex) whose increased co-transcriptional recruitment promotes the attachment of the 3’-5’ exonuclease Rrp6p along with the co-factors Rrp47p and Mpp6p. Interestingly, we established that Rrp6p functions independently from the core exosome, yet is stimulated by two forms of the TRAMP complex that include Trf4p or Trf5p and Air2p but not Air1p. The results suggest that specific substrates could be primed for decay via various QC pathways owing to the versatility of the mRNA degradation apparatus. In this context, the bacterial Rho helicase provides a valuable tool to decipher the QC molecular process in yeast and possibly the homologous process in mammalian cells.

International audience; The co-transcriptional mRNA processing and packaging reactions that lead to the formation of export-competent mRNPs are under the surveillance of quality control steps. Aberrant mRNPs resulting from faulty events are retained in the nucleus with ensuing elimination of their mRNA component. The molecular mechanisms by which the surveillance system recognizes defective mRNPs and stimulates their destruction by the RNA degradation machinery are still not completely elucidated. Using an experimental approach in which mRNP formation in yeast is disturbed by the action of the bacterial Rho helicase, we have shown previously that the targeting of Rho-induced aberrant mRNPs is mediated by Rrp6p which is recruited co-transcriptionally in association with Nrd1p following Rho action. Here, we investigated the specific involvement in this quality control process of different cofactors associated with the nuclear RNA degradation machinery. We show that, in addition to the main hydrolytic action of the exonuclease Rrp6p, the cofactors Rrp47p and Mpp6p as well as the TRAMP components Trf4p, Trf5p and Air2p contribute significantly by stimulating the degradation process upon their co-transcriptional recruitment. Trf4p and Trf5p are apparently recruited in two distinct TRAMP complexes that both contain Air2p as component. Surprisingly, Rrp47p appears to play an important role in mutual protein stabilization with Rrp6p which highlights a close association between the two partners. Together, our results provide an integrated view of how different cofactors of the RNA degradation machinery cooperate to target and eliminate aberrant mRNPs.

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