Search

Plant responses to salt exposure involve large reconfigurations of hormonal pathways that orchestrate physiological changes towards tolerance. Jasmonate (JA) hormones are essential to withstand biotic and abiotic assaults, but their roles in salt tolerance remain unclear. Here we describe the dynamics of JA metabolism and signaling in root and leaf tissue of rice, a plant species that is highly exposed and sensitive to salt. Roots activate the JA pathway in an early pulse, while the second leaf displays a biphasic JA response with peaks at 1 h and 3 d post-exposure. Based on higher salt tolerance of a rice JA-deficient mutant (aoc), we examined, through kinetic transcriptome and physiological analysis, the salt-triggered processes that are under JA control. Profound genotype-differential features emerged that could underlie the observed phenotypes. Abscisic acid (ABA) content and ABA-dependent water deprivation responses were impaired in aoc shoots. Moreover, aoc accumulated more Na+ in roots, and less in leaves, with reduced ion translocation correlating with root derepression of the HAK4 Na+ transporter gene. Distinct reactive oxygen species scavengers were also stronger in aoc leaves, along with reduced senescence and chlorophyll catabolism markers. Collectively, our results identify contrasted contributions of JA signaling to different sectors of the salt stress response in rice.

PlantRNA (http://plantrna.ibmp.cnrs.fr/) is a comprehensive database of transfer RNA (tRNA) gene sequences retrieved from fully annotated nuclear, plastidial and mitochondrial genomes of photosynthetic organisms. In the first release (PlantRNA 1.0), tRNA genes from 11 organisms were annotated. In this second version, the annotation was implemented to 51 photosynthetic species covering the whole phylogenetic tree of photosynthetic organisms, from the most basal group of Archeplastida, the glaucophyte Cyanophora paradoxa, to various land plants. Transfer RNA genes from lower photosynthetic organisms such as streptophyte algae or lycophytes as well as extremophile photosynthetic species such as Eutrema parvulum were incorporated in the database. As a whole, circa 37 000 tRNA genes were accurately annotated. In the frame of the tRNA genes annotation from the genome of the Rhodophyte Chondrus crispus, non-canonical splicing sites in the D- or T- regions of tRNA molecules were identified and experimentally validated. As for PlantRNA 1.0, comprehensive biological information including 5’- and 3’-flanking sequences, A and B box sequences, region of transcription initiation and poly(T) transcription termination stretches, tRNA intron sequences and tRNA mitochondrial import are included.

Specialized metabolite (SM) diversification is a core process to plants’ adaptation to diverse ecological niches. Here we implemented a computational mass spectrometry (MS)-based metabolomics approach to explore SM diversification in tissues of 20 species covering Nicotiana phylogenetics sections. To drastically increase metabolite annotation, we created a large in silico fragmentation database, comprising more than 1 million structures, and scripts for connecting class prediction to consensus substructures. Altogether, the approach provides an unprecedented cartography of SM diversity and section-specific innovations in this genus. As a case-study, and in combination with NMR and MS imaging, we explored the distribution of N-acyl nornicotines, alkaloids predicted to be specific to Repandae allopolyploids, and revealed their prevalence in the genus, albeit at much lower magnitude, as well as a greater structural diversity than previously thought. Altogether, the novel data integration approaches provided here should act as a resource for future research in plant SM evolution.TeaserComputational metabolomics delineates main trends in the diversification of specialized metabolism in the genus Nicotiana

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.

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.

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 Lachancea kluyveri species, we expanded our knowledge on how genetic characteristics of large introgression events can affect the phenotypic landscape.

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.

Hybrid incompatibility resulting from deleterious gene combinations is thought to be an important step towards reproductive isolation and speciation. Here we demonstrate involvement of a silent epiallele in hybrid incompatibility. InArabidopsis thalianastrain Col-0, one of the two copies of a duplicated histidine biosynthesis gene,HISN6Bis not expressed, for reasons that have been unclear, making its paralog,HISN6Aessential. By contrast, in strain Cvi-0,HISN6Bis essential becauseHISN6Ais mutated. As a result of these differences, Cvi-0 × Col-0 hybrid progeny that are homozygous for both Col-0HISN6Band Cvi-0HISN6Ado not survive. We show thatHISN6Bis not a defective pseudogene in the Col-0 strain, but a stably silenced epiallele. MutatingHISTONE DEACETYLASE 6 (HDA6)or the cytosine methyltransferase genes,MET1orCMT3erasesHISN6B’ssilent locus identity in Col-0, reanimating the gene such thathisn6alethality and hybrid incompatibility are circumvented. These results show thatHISN6-dependent hybrid lethality is a revertible epigenetic phenomenon and provide additional evidence that epigenetic variation has the potential to limit gene flow between diverging populations of a species.Significance statementDeleterious mutations in different copies of a duplicated gene pair have the potential to cause hybrid incompatibility between diverging subpopulations, contributing to reproductive isolation and speciation. This study demonstrates a case of epigenetic gene silencing, rather than pseudogene creation by mutation, contributing to a lethal gene combination upon hybridization of two strains ofArabidopsis thaliana. The findings provide direct evidence that naturally occurring epigenetic variation can contribute to incompatible hybrid genotypes, reducing gene flow between strains of the same species.

SummaryMendelian traits are considered to be at the lower end of the complexity spectrum of heritable phenotypes. However, more than a century after the rediscovery of Mendel’s law, the global landscape of monogenic variants, as well as their effects and inheritance patterns within natural populations, is still not well understood. Using the yeast Saccharomyces cerevisiae, we performed a species-wide survey of Mendelian traits across a large population of isolates. We generated offspring from 41 unique parental pairs and analyzed 1,105 cross/trait combinations. We found that 8.9% of the cases were Mendelian. Further tracing of causal variants revealed background-specific expressivity and modified inheritances, gradually transitioning from Mendelian to complex traits in 30% of the cases. In fact, when taking into account the natural population diversity, the hidden complexity of traits could be substantial, confounding phenotypic predictability even for simple Mendelian traits.

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.

RNA-guided surveillance systems constrain the activity of transposable elements (TEs) in host genomes. In plants, RNA polymerase IV (Pol IV) transcribes TEs into primary transcripts from which RDR2 synthesizes double-stranded RNA precursors for small interfering RNAs (siRNAs) that guide TE methylation and silencing. How the core subunits of Pol IV, homologs of RNA polymerase II subunits, diverged to support siRNA biogenesis in a TE-rich, repressive chromatin context is not well understood. Here we studied the N-terminus of Pol IV’s largest subunit, NRPD1. Arabidopsis lines harboring missense mutations in this N-terminus produce wild-type (WT) levels of NRPD1, which co-purifies with other Pol IV subunits and RDR2. Our in vitro transcription and genomic analyses reveal that the NRPD1 N-terminus is critical for robust Pol IV-dependent transcription, siRNA production and DNA methylation. However, residual RNA-directed DNA methylation observed in one mutant genotype indicates that Pol IV can operate uncoupled from the high siRNA levels typically observed in WT plants. This mutation disrupts a motif uniquely conserved in Pol IV, crippling the enzyme's ability to inhibit retrotransposon mobilization. We propose that the NRPD1 N-terminus motif evolved to regulate Pol IV function in genome surveillance.