Horizontal Transfers (HTs) refers to the movement of genetic material between distantly related species by mechanisms other than sexual reproduction. In prokaryotes, HTs are well known to be a source of new adaptive traits such as the spreading of genes implicated in antibiotic resistance. Over the last decade, it has become increasingly obvious that in eukaryotes too such as plants, HTs can also lead to major evolutionary leaps and very fast adaptation to new ecological niches that would not be possible only by standard genetic mutations. However, little is known about their rate, evolutionary significance and the biotic interaction promoting HT between plants. While host-parasite relationships appear to be a major pathway for horizontal transfers, an increasing number of HT cases reported in recent years involve species that do not share intimate cell-to-cell contact. This, raises the question alternative HT pathway between species not involved in tight physical contact and the possible involvement of vectors. With the advent of next and third generation sequencing technologies and the substantial increase of genomic data in plants, it becomes feasible to assess the extent and routes of HT at an unprecedented larger-scale. While these sequencing technologies have enabled a massive increase in genomic data from plants and other organisms, new challenges must be addressed not only in terms of bioinformatics analysis of large datasets, but also for the automatic and accurate HT characterization. To date, no studies have been conducted on a genome-wide scale and for a large number of plant species. Indeed, most previous HT studies have focused on specific genes or only on certain transposable elements (TEs) classes and on a limited number of plant species. The EXOTICA project seeks to tackle those challenges by providing innovative comparative genomic approaches for HTs characterization that could handle big genomic data in order to determine the extent, possible routes and the nature of biological interactions promoting HTs in plants. We have recently developed novel bioinformatics approaches that seek to characterize HTs at the whole genome scale and without any prior knowledge on genome annotation. Thanks to these tools, we will characterize HTs on an unprecedented scale by comparing thousands of publicly available genomes and by de novo sequencing of several plant species from a natural ecosystem: the Massane forest located in southern France. EXOTICA will set up the basis for a full and comprehensive understanding of HT routes and the type of biotic interactions promoting HT between non-parasitic plants. We will also test the hypothesis of the implication of vector such as bacteria, fungi or insect in plant-to-plant DNA transfer.

High ambient temperatures due to climate change impact plant growth and survival. Recent data indicate that chromatin modification is an essential process of gene expression reprogramming during plant response to elevated growth temperature. Histone deacetylases (HDAC) that regulate histone acetylation levels were shown to play important roles in plant adaptation to environment. In addition, HDACs are also involved in deacetylation of non-histone proteins such as metabolic enzymes and transcription factors to control their activity. Our preliminary data indicate that Arabidopsis HDAC members play distinct roles in plant response to high ambient temperature. In addition, we found that high ambient temperature alters plant cell redox environment and that cellular redox environment regulates HDAC subcellular localization and deacetylase activity. The objective of this proposal is to elucidate the molecular mechanisms of interplay between cellular redox and chromatin modification that regulate plant response to high ambient temperature. In particular, the project aims to elucidate how cellular redox environment regulates HDAC activity to control lysine acetylation of histone and non-histone proteins involved in either epigenetic regulation of gene expression or metabolism and/or signaling during plant response to high ambient temperature, and to identify and study implicated redox regulators. We propose to first identify redox post-translational modifications of Arabidopsis HDACs under normal and high ambient temperature conditions by using biochemical and mass spectrometry approaches. Then, we will examine effects of redox modifications on HDAC enzymatic activity, subcellular localization, and function in plant response to high ambient temperature thanks to plants expressing tagged HDAC proteins in wild-type and mutated versions of redox-sensitive residues. Then, we plan to identify and validate redox regulators involved in modifications of the HDACs by both biochemical and genetic approaches and to study the role of redox regulators in plant response to high temperature. Next, we will study the effects of redox modifications on HDAC epigenetic function in terms of chromatin structure, genome-wide histone modifications, DNA methylation and gene expression by high throughput sequencing and cell biology approaches. Finally, we will investigate the effect of redox modifications on HDAC functions in regulating lysine acetylation of non-histone proteins in order to identify HDAC-regulated key metabolic enzymes and signaling proteins involved in plant response to high ambient temperature. This project aiming to elucidate redox-epigenetics-metabolism networks in plants will deepen current understanding of how plants adapt or resist to a changing environment. More specifically, the project will decipher the molecular basis of HDAC regulation in response to stress and how thiol modifications modulate their functions. Noteworthy, this study will reveal the function of HDAC-dependent lysine acetylation in regulating activity of non-histone proteins which is largely unknown at present time. We believe that the results obtained from this project will lead to establish a general link and reveal the molecular mechanisms of interplay between redox signaling, epigenetic regulation and plant adaptation to environment. The REPHARE project assembles complementary expertise in the fields of epigenetic regulation and redox signaling from the two partners who have already built a solid basis which will lead the project to success.

Plant adaptation to challenging environmental conditions around the world has made root growth and development an important research area for plant breeders and scientists. Despite its importance, the genes and molecular mechanisms that govern root growth and development in changing conditions remain largely unknown. ROXYs constitute a plant-specific family of Glutaredoxins and a growing body of evidence indicates that they are involved in numerous cellular pathways to regulate different aspects of plant development such as root development in response to nutrients. Despite a large amount of data, the function of ROXYs at the molecular level remains largely unknown. In most studies, ROXYs were suggested to mediate their biological effects through protein-protein interactions with one or more members of the TGA transcription factor family. The interaction between glutaredoxins and TGA proteins may cause the transcriptional activity changes of TGA proteins caused by Cys residue modification. We recently identified 2 ROXYs, ROXY18 and ROXY19, involved in root development in response to nitrate deficiency and drought. We postulate that the Arabidopsis ROXY18 and ROXY19 are key regulators of root system architecture and are probably essential for its proper adaptation to changing growth conditions. We aim to understand how these ROXYs control root architecture adaptation to environment, through a combination of molecular genetics, physiology and cell biology. We will 1- analyze the impact of nitrate avaibility and drought on ROXYs and TGAs transcriptional and post-transcriptional regulation; 2- study the functions and the molecular mechanisms of ROXY18 and 19 and candidates TGAs involved in modulating root development; 3- determine how these ROXY/TGA complexes regulate root development in response to nitrate and drought.

Global climate change is a reality, day-to-day impacting our capacity to ensure food security worldwide. The timely reprogramming of gene expression in response to internal and external cues is essential for plant development and acclimation, especially in harsh environmental conditions. Understanding how plants do this is essential to design the next generation of stress-resistant crops. Recently, mRNA chemical (i.e epitranscriptomic) modifications were shown to play a key role in this process. In contrast, tRNA chemical modifications, although critical for optimal function of the translational apparatus, and much more diverse and quantitatively important compared to mRNAs modifications, were until recently considered as mainly static chemical decorations. The main objective of this project is, using several genome-wide approaches followed by functional validations, to test the hypothesis that plant tRNA epitranscriptomic marks can vary between different developmental stages and stress conditions and that these variations are not only passive consequences of different cellular conditions but can fine-tune translation to adapt plant responses to internal and external cues. It also aims at identifying the most critical tRNA marks needed to efficiency translate key developmental and stress responsive genes and to provide mechanistic evidence that proper codon decoding is central to this regulatory network. The project will attempt to establish tRNA epitranscriptomic variations as a new layer of plant gene regulation important for development and stress responses. It also aims at reveling direct links between tRNA marks, tRNA steady state levels, mRNAs translation efficiency and protein production at genome-wide level. This project will identify critical developmental and stress-responsive genes whose expression is affected by the loss of key tRNA marks. It will also reveal new gene targets for improving plant growth and its capacity to survive abiotic stresses.

Ribosome assembly and regulation of translation activity is crucial for cell growth and response to external cues or environmental conditions in all eukaryotic organisms. Ribosomal RNA (rRNA) are the linchpins of ribosome assembly and function. The rRNAs 25S, 5.8S, 5S and 18S form, respectively, the large (LSU) and the small (SSU) subunits of the ribosome. The LSU acts as a ribozyme, catalyzing peptide bond formation, while SSU harbors the decoding center which monitors the complementarity of tRNA anticodon with mRNA codon during translation. In all eukaryotic cells, rRNA are subjects of two major types of nucleotide modifications: methylation of sugars (2’-O-ribose) and conversion of uridine to pseudouridine. Over 90% of the rRNA methylations are 2’-O-ribose methylations which are guided by antisense small nucleolar RNA (snoRNA) referred as C/D-box snoRNA. The C/D-box snoRNA associates to four nucleolar proteins called the C/D core proteins (Nop56p/p62, Nop58p, snu13/15.5K and the methyltransferase Nop1p/Fibrillarin) to form the C/D-box snoRNP. The extent and positions of rRNA 2’-O-Me fluctuate between species, however the majority of modifications occurs in functionally conserved regions. Remarkably, modulation of rRNA 2'-O-methylation affects ribosome translation fidelity and is linked to human diseases. In plants, only limited information is available on rRNA 2'-O-methylation profile, mechanism of modification and role(s) of modified nucleotides in ribosome activity. In the plant model A. thaliana, ~100 C/D-box snoRNA have been identified by computational screening of genomic sequences. Later, analysis of small RNA-seq data and targeted sequencing of nucleolar RNAs expanded the list of known C/D-box snoRNA to ~200 species. Notably, studies in Arabidopsis demonstrated that the knockout of a single C/D-box snoRNA triggers strong developmental and growth defects, and gene disruption of NUFIP, a C/D-box snoRNP assembly factor, also leads to severe developmental phenotypes and inhibited 2’-O-methylation at specific rRNA sites. We have recently performed RiboMethSeq analysis of Arabidopsis thaliana rRNA and identified both conserved and plant-specific rRNA 2’-O-Me sites. Remarkably; our data suggest that some rRNA 2’-O-Me sites do not have matching snoRNAs, suggesting 2’-O-Me through a non-canonical snoRNA and/or another guiding mechanism. Furthermore, we also observed alterations of rRNA 2’-O-Me level in both conserved and plant-specific rRNA positions in response to specific developmental conditions. Furthermore, RNA-seq analysis of Arabidopsis C/D-box snoRNPs identified a subset of non-coding RNA, other than known C/D-box snoRNA, that might be implicated in 2’-O-methylation of rRNA and /or other RNAs. In this project, we will perform deep investigation of the mechanisms controlling 2’-O-methylation of rRNA and determine connections with other rRNA modifications and translation activity of plant ribosomes. We will determine how 2’-O-ribose methylation of rRNA contributes to the production of heterogeneous ‘specialized’ ribosomes in plants and impact plant resistance and/or adaptation to heat stress conditions. The MetRibo project is intended to contribute substantially into the intense efforts currently devoted to decipher how ribosome modifications contributes to regulatory pathways and how is tuned during adaptive responses of multicellular organisms to external cues, a field to which plants have much to offer.