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16 p.-7 fig., Virus-induced plant diseases in cultivated plants cause important damages in yield. Although the mechanisms of virus infection are intensely studied at the cell biology level, only little is known about the molecular dialog between the invading virus and the host genome. Here we describe a combinatorial genome-wide approach to identify networks of sRNAs-guided posttranscriptional regulation within local Turnip mosaic virus (TuMV) infection sites in Brassica napus leaves. We show that the induction of host-encoded, virus-activated small interfering RNAs (vasiRNAs) observed in virus-infected tissues is accompanied by site-specific cleavage events on both viral and host RNAs that recalls the activity of small RNA-induced silencing complexes (RISC). Cleavage events also involve virus-derived siRNA (vsiRNA)–directed cleavage of target host transcripts as well as cleavage of viral RNA by both host vasiRNAs and vsiRNAs. Furthermore, certain coding genes act as virus-activated regulatory hubs to produce vasiRNAs for the targeting of other host genes. The observations draw an advanced model of plant-virus interactions and provide insights into the complex regulatory networking at the plant-virus interface within cells undergoing early stages of infection., This work has been subject to the trinational PLANT-KBBE 2012 project GAMAVIR. We acknowledge funding from the French Agence National de la Recherche (grant ANR-13-KBBE-0005-01), the Swiss National Science Foundation (SNF, grant 31003A_140694), and the Région Alsace (PhD fellowship for N.P.) to M.H., from the Spanish Ministerio de Economia y Competitividad (MINECO, PCIN-213-064 and BIO2015-70752-R) to C.L., and the Deutsche Forschungsgemeinschaft (DFG, grant 031A324) to M.W.

The RNA exosome is a key 3’−5’ exoribonuclease with an evolutionarily conserved structure and function. Its cytosolic functions require the co-factors SKI7 and the Ski complex. Here we demonstrate by co-purification experiments that the ARM-repeat protein RESURRECTION1 (RST1) and RST1 INTERACTING PROTEIN (RIPR) connect the cytosolic Arabidopsis RNA exosome to the Ski complex. rst1 and ripr mutants accumulate RNA quality control siRNAs (rqc-siRNAs) produced by the post-transcriptional gene silencing (PTGS) machinery when mRNA degradation is compromised. The small RNA populations observed in rst1 and ripr mutants are also detected in mutants lacking the RRP45B/CER7 core exosome subunit. Thus, molecular and genetic evidence supports a physical and functional link between RST1, RIPR and the RNA exosome. Our data reveal the existence of additional cytosolic exosome co-factors besides the known Ski subunits. RST1 is not restricted to plants, as homologues with a similar domain architecture but unknown function exist in animals, including humans., Cytosolic RNA degradation by the RNA exosome requires the Ski complex. Here the authors show that the proteins RST1 and RIPR assist the RNA exosome and the Ski complex in RNA degradation, thereby preventing the production of secondary siRNAs from endogenous mRNAs.

Uridylation is a widespread modification destabilizing eukaryotic mRNAs. Yet, molecular mechanisms underlying TUTase-mediated mRNA degradation remain mostly unresolved. Here, we report that the Arabidopsis TUTase URT1 participates in a molecular network connecting several translational repressors/decapping activators. URT1 directly interacts with DECAPPING 5 (DCP5), the Arabidopsis ortholog of human LSM14 and yeast Scd6, and this interaction connects URT1 to additional decay factors like DDX6/Dhh1-like RNA helicases. Nanopore direct RNA sequencing reveals a global role of URT1 in shaping poly(A) tail length, notably by preventing the accumulation of excessively deadenylated mRNAs. Based on in vitro and in planta data, we propose a model that explains how URT1 could reduce the accumulation of oligo(A)-tailed mRNAs both by favoring their degradation and because 3’ terminal uridines intrinsically hinder deadenylation. Importantly, preventing the accumulation of excessively deadenylated mRNAs avoids the biogenesis of illegitimate siRNAs that silence endogenous mRNAs and perturb Arabidopsis growth and development., TUTase mediated uridylation of mRNA promotes degradation. Here, Scheer, de Almeida et al. show that Arabidopsis TUTase URT1 interacts directly with the translation inhibitor and decay factor DECAPPING5 and suppresses siRNA biogenesis by preventing accumulation of deadenylated mRNAs

Beyond their key role in translation, cytosolic transfer RNAs (tRNAs) are involved in a wide range of other biological processes. Nuclear tRNA genes (tDNAs) are transcribed by the RNA polymerase III (RNAP III) and cis-elements, trans-factors as well as genomic features are known to influence their expression. In Arabidopsis, besides a predominant population of dispersed tDNAs spread along the 5 chromosomes, some clustered tDNAs have been identified. Here, we demonstrate that these tDNA clusters are transcriptionally silent and that pathways involved in the maintenance of DNA methylation play a predominant role in their repression. Moreover, we show that clustered tDNAs exhibit repressive chromatin features whilst their dispersed counterparts contain permissive euchromatic marks. This work demonstrates that both genomic and epigenomic contexts are key players in the regulation of tDNAs transcription. The conservation of most of these regulatory processes suggests that this pioneering work in Arabidopsis can provide new insights into the regulation of RNA Pol III transcription in other organisms, including vertebrates.

Beyond their key role in translation, cytosolic transfer RNAs (tRNAs) are involved in a wide range of other biological processes. Nuclear tRNA genes (tDNAs) are transcribed by the RNA polymerase III (RNAP III) andcis-elements,trans-factors as well as genomic features are known to influence their expression. In Arabidopsis, besides a predominant population of dispersed tDNAs spread along the 5 chromosomes, some clustered tDNAs have been identified. Here, we demonstrate that these tDNA clusters are transcriptionally silent and that pathways involved in the maintenance of DNA methylation play a predominant role in their repression. Moreover, we show that clustered tDNAs exhibit repressive chromatin features whilst their dispersed counterparts contain permissive euchromatic marks. Our data highlight that the combination of both genomic environment and epigenomic landscape contribute to fine tune the differential expression of dispersed versus clustered tDNAs in Arabidopsis.

To explore the origin of the diversity observed in natural populations, many studies have investigated the relationship between genotype and phenotype. In yeast species, especially in Saccharomyces cerevisiae, these studies are mainly conducted using recombinant offspring derived from two genetically diverse isolates, allowing to define the phenotypic effect of genetic variants. However, large genomic variants such as interspecies introgressions are usually overlooked even if they are known to modify the genotype-phenotype relationship. To have a better insight into the overall phenotypic impact of introgressions, we took advantage of the presence of a 1-Mb introgressed region, which lacks recombination and contains the mating-type determinant in the Lachancea kluyveri budding yeast. By performing linkage mapping analyses in this species, we identified a total of 89 loci affecting growth fitness in a large number of conditions and 2,187 loci affecting gene expression mostly grouped into two major hotspots, one being the introgressed region carrying the mating-type locus. Because of the absence of recombination, our results highlight the presence of a sexual dimorphism in a budding yeast for the first time. Overall, by describing the phenotype-genotype relationship in the L. kluyveri species, we expanded our knowledge on how genetic characteristics of large introgression events can affect the phenotypic landscape.

International audience; Cells have sophisticated RNA-directed mechanisms to regulate genes, destroy viruses, or silence transposable elements (TEs). In terrestrial plants, a specialized non-coding RNA machinery involving RNA polymerase IV (Pol IV) and small interfering RNAs (siRNAs) targets DNA methylation and silencing to TEs. Here, we present a bioinformatics protocol for annotating and quantifying siRNAs that derive from long terminal repeat (LTR) retrotransposons. The approach was validated using small RNA northern blot analyses, comparing the species Arabidopsis thaliana and Brachypodium distachyon. To assist hybridization probe design, we configured a genome browser to show small RNA-seq mappings in distinct colors and shades according to their nucleotide lengths and abundances, respectively. Samples from wild-type and pol IV mutant plants, cross-species negative controls, and a conserved microRNA control validated the detected siRNA signals, confirming their origin from specific TEs and their Pol IV-dependent biogenesis. Moreover, an optimized labeling method yielded probes that could detect low-abundance siRNAs from B. distachyon TEs. The integration of de novo TE annotation, small RNA-seq profiling, and northern blotting, as outlined here, will facilitate the comparative genomic analysis of RNA silencing in crop plants and non-model species.

In the natural environment, organisms have to cope with a large number of stresses. Comparison of the gene expression response to different stresses across different yeast species shows that transcriptomic stress response can be linked to the lifestyle of the studied species., Defining how organisms respond to environmental change has always been an important step toward understanding their adaptive capacity and physiology. Variation in transcription during stress has been widely described in model species, especially in the yeast Saccharomyces cerevisiae, which helped to shape general rules regarding how cells cope with environmental constraints, as well as to decipher the functions of many genes. Comparison of the environmental stress response (ESR) across species is essential to obtaining better insight into the common and species-specific features of stress defense. In this context, we explored the transcriptional landscape of the yeast Lachancea kluyveri (formerly Saccharomyces kluyveri) in response to diverse stresses, using RNA sequencing. We investigated variation in gene expression and observed a link between genetic plasticity and environmental sensitivity. We identified the ESR genes in this species and compared them to those already found in S. cerevisiae. We observed common features between the two species, as well as divergence in the regulatory networks involved. Of interest, some changes were related to differences in species lifestyle. Thus we were able to decipher how adaptation to stress has evolved among different yeast species. Finally, by analyzing patterns of coexpression, we were able to propose potential biological functions for 42% of genes and also annotate 301 genes for which no function could be assigned by homology. This large data set allowed for the characterization of the evolution of gene regulation and provides an efficient tool for assessing gene function.

Meiotic recombination is a major factor of genome evolution, deeply characterized in only a few model species, notably the yeast Saccharomyces cerevisiae. Consequently, little is known about variations of its properties across species. In this respect, we explored the recombination landscape of Lachancea kluyveri, a protoploid yeast species that diverged from the Saccharomyces genus more than 100 million years ago and we found striking differences with S. cerevisiae. These variations include a lower recombination rate, a higher frequency of chromosomes segregating without any crossover and the absence of recombination on the chromosome arm containing the sex locus. In addition, although well conserved within the Saccharomyces clade, the S. cerevisiae recombination hotspots are not conserved over a broader evolutionary distance. Finally and strikingly, we found evidence of frequent reversal of commitment to meiosis, resulting in return to mitotic growth after allele shuffling. Identification of this major but underestimated evolutionary phenomenon illustrates the relevance of exploring non-model species., Author summary Meiotic recombination promotes accurate chromosome segregation and genetic diversity. To date, the mechanisms and rules lying behind recombination were dissected using model organisms such as the budding yeast Saccharomyces cerevisiae. To assess the conservation and variation of this process over a broad evolutionary distance, we explored the meiotic recombination landscape in Lachancea kluyveri, a budding yeast species that diverged from S. cerevisiae more than 100 million years ago. The meiotic recombination map we generated revealed that the meiotic recombination landscape and properties significantly vary across distantly related yeast species, raising the yet to confirm possibility that recombination hotspots conservation across yeast species depends on synteny conservation. Finally, the frequent meiotic reversions we observed led us to re-evaluate their evolutionary importance.