Exploration bioinformatique des interactions pollen–pistil chez Solanum chacoense

Rassemblant 107 espèces distribuées dans toute l’Amérique latine, les pommes de terre sauvages (Solanum sect. Petota) forment un réservoir de germoplasme important pour l’amélioration de leur cousine cultivée (S. tuberosum). Pourtant, bien qu’elles vivent en sympatrie et qu’elles aient la capacité physiologique de s’hybrider avec de proches parentes, ces espèces sauvages présentent de fortes barrières d’isolement reproductif. Les résultats préliminaires sur lesquels se fondent cette thèse, présentés au chapitre 1, montrent que les interactions pollen-pistil sont au cœur de cet isolement: dans un pistil hétérospécifique, les tubes polliniques (TP) sont ralentis et leur réceptivité aux signaux chimioattractifs de l’ovule est compromise. Comme nous le verrons au chapitre 2, l’examen de l’abondante littérature scientifique existante indique qu’un grand nombre de protéines exprimées dans le pistil et le TP ont été caractérisées chez d’autres espèces et que plusieurs contribuent, par leur divergence interspécifique, à l’établissement de points de contrôle précis tout le long du trajet du TP dans le pistil. Parmi elles, les protéines chimioattractives LURE contrôlant le guidage directionnel du TP chez Torenia et Arabidopsis sont un exemple remarquable de médiateurs moléculaires de l’isolement reproductif. L’objectif de cette thèse est d’isoler des gènes candidats jouant des fonctions analogues chez les Solanacées tubéreuses. C’est dans ce souci que nous avons entrepris d’explorer à grande échelle, dans une approche mêlant bioinformatique, protéomique et transcriptomique, la diversité des gènes exprimés dans les ovules de S. chacoense. Au chapitre 3, nous présentons un nouvel outil de recherche de séquences, KAPPA, spécifiquement dédié à la recherche de peptides riches en cystéines (CRP), une catégorie de protéines particulière dont font partie les chimioattractants ovulaires connus, ainsi que plusieurs autres interactants pollen-pistil. Cet outil a facilité l’identification de nouvelles CRP dans les chapitres suivants. Nous en détaillons l’algorithme, en présentons les principales fonctions et fournissons des résultats démontrant ses performances sur des jeux de données calibrés. Dans le chapitre 4, nous décrivons une étude du sécrétome ovulaire de S. chacoense comparant le contenu protéique des exsudats d’ovules matures (qui sont capables d’attirer le TP) et d’ovules immatures (qui en sont incapables). Nous présentons la méthode tissue-free gravity-extraction method (tf-GEM) utilisée pour collecter les exsudats et détaillons l’analyse bioinformatique effectuée à partir des résultats de spectrométrie de masse. Nous passons en revue les 305 protéines sécrétées identifiées dans les exsudats d’ovules, dont environ la moitié sont régulées entre les deux conditions à l’étude, analysons les catégories fonctionnelles associées, et discutons des applications possibles pour la découverte de gènes candidats. Le chapitre 5 présente les résultats d’une analyse par biopuce de la réponse transcriptionnelle des ovaires de S. chacoense à différents types de pollinisation : conspécifique compatible, conspécifique incompatible, et hétérospécifique compatible. Nous montrons que des gènes ovulaires sont activés ou réprimés à distance dès les premières heures suivant la pollinisation et que cette réponse s’amplifie et devient spécifique à chaque génotype mâle au fur et à mesure de la progression des TP dans le style jusqu’à la fécondation. Nous discutons du rôle de différents médiateurs, notamment l’éthylène, dans la transmission à longue distance du signal de pollinisation. Au chapitre 6, nous présentons les résultats d’une analyse transcriptomique de plus grande profondeur réalisée par séquençage d’ARN sur des ovules de S. chacoense capables (ovules sauvages matures) ou incapables (ovules sauvages immatures, ovules du mutant frk1 dépourvus de sac embryonnaire) d’attirer les TP. Nous montrons comment notre méthode hybride, recourant à la fois à des assemblages de novo et ab initio contre le génome récemment publié de S. chacoense, nous a conduits à obtenir un jeu de 4353 transcrits enrichis dans l’ovule. Nous détaillons les améliorations portées aux annotations existantes du génome, avec notamment la découverte de nouveaux loci codant pour des CRP reproductives. Nous présentons une analyse du profil d’expression des gènes dans l’ovule, discutons des catégories fonctionnelles associées, et terminons avec l’identification d’une liste de chimioattractants candidats. Enfin, le chapitre 7 présente en détail ces chimioattractants candidats, les techniques utilisées pour exprimer et purifier ces protéines en bactéries et en levures, ainsi que les dispositifs microfluidiques élaborés pour en vérifier la capacité à attirer le tube pollinique. Nous incluons les résultats préliminaires obtenus avec cette méthode et concluons sur les développements possibles de notre projet.With 107 species distributed throughout Latin America, wild potatoes (Solanum sect. Petota) form an important germplasm reservoir for the breeding of their cultivated cousin (S. tuberosum). Yet, although they live in sympatry and have the physiological ability to hybridize with close relatives, those wild species exhibit strong reproductive isolation barriers. The preliminary results on which this thesis is based, presented in chapter 1, show that pollen-pistil interactions are at the center of this isolation: in a heterospecific pistil, pollen tubes (PTs) are slowed down and their receptivity to ovular chemoattractive signals is compromised. As we will see in chapter 2, the review of the abundant existing scientific literature indicates that a large number of proteins expressed in pistils and PTs have been characterized in other species and that several contribute, through their interspecific divergence, to the establishment of specific checkpoints along the entire path of PTs in pistil. Among them, the LURE chemoattractive proteins controlling the directional guidance of PTs in Torenia and Arabidopsis are an outstanding example of molecular mediators of reproductive isolation. The objective of this thesis is to isolate candidate genes playing similar functions in tuber-bearing Solanaceous species. It is with this in mind that we have undertaken a large-scale exploration of the diversity of genes expressed in the ovules of S. chacoense, in an approach combining bioinformatics, proteomics and transcriptomics. In chapter 3, we present a new sequence search tool, KAPPA, specifically dedicated to the detection of cystein-rich peptides (CRPs), a particular class of proteins that includes known ovular chemoattractants, as well as several other pollen–pistil interactants. This tool has facilitated the identification of new CRPs in the following chapters. We detail the algorithm, present its main functions and provide results demonstrating its performance on calibrated datasets. In chapter 4, we describe a study of the ovule secretome of S. chacoense comparing the protein content of exudates from mature ovules (which are able to attract PTs) and immature ovules (which are unable to do so). We present the tissue-free gravity-extraction method (tf-GEM) used to collect exudates and detail the bioinformatics analysis performed on mass spectrometry results. We review the 305 secreted proteins identified in ovule exudates, about half of which are regulated between the two conditions under study, analyze the associated functional categories, and discuss possible applications for the discovery of candidate genes. Chapter 5 presents the results of a microarray analysis of the transcriptional response of S. chacoense ovaries to different pollination types: conspecific compatible, conspecific incompatible, and heterospecific compatible. We show that ovarian genes are activated or repressed at a distance as early as the first few hours after pollination and that this response is amplified and becomes specific to each male genotype as the PTs progress in the style until fertilization. We discuss the role of different mediators, including ethylene, in the long-range transmission of the pollination signal. In chapter 6, we present the results of a deeper transcriptomic analysis performed by sequencing RNAs from PT guidance-competent (mature wild-type) ovules and PT guidance-incompetent (immature wild-type and embryo sac-devoid frk1 mutant) ovules in S. chacoense. We show how our hybrid method, relying on both de novo and ab initio assemblies against the S. chacoense genome published recently, led us to obtain a set of 4353 ovule-enriched transcripts. We detail the improvements made to existing genome annotations, including the discovery of new loci encoding reproductive CRPs. We present an analysis of the gene expression profile in the ovule, discuss associated functional categories, and conclude with the identification of a list of candidate chemoattractants. Finally, chapter 7 details these candidate chemoattractants, the techniques used to express and purify them in bacteria and yeasts, and the microfluidic devices developed to verify their ability to attract the pollen tube. We include preliminary results obtained with this method and conclude on possible developments of our project

Adversarial autoencoders for novelty detection

In this paper, we address the problem of novelty detection, i.e recognizing at test time if a data item comes from the training data distribution or not. We focus on Adversarial autoencoders (AAE) that have the advantage to explicitly control the distribution of the known data in the feature space. We show that when they are trained in a (semi-)supervised way, they provide consistent novelty detection improvements compared to a classical autoencoder. We further improve their performance by introducing an explicit rejection class in the prior distribution coupled with random input images to the autoencoder

On the analysis of perfectly matched layers for a class of dispersive media and application to negative index metamaterials

This work deals with Perfectly Matched Layers (PMLs) in the context of dispersive media, and in particular for Negative Index Metamaterials (NIMs). We first present some properties of dispersive isotropic Maxwell equations that include NIMs. We then demonstrate numerically the inherent instabilities of the classical PMLs applied to NIMs. We propose and analyse the stability of very general PMLs for a large class of dispersive systems using a new change of variable. We give necessary criteria for the stability of such models. For dispersive isotropic Maxwell equations, this analysis is completed by giving necessary and sufficient conditions of stability. Finally, we propose new PMLs that satisfy these criteria and demonstrate numerically their efficiency

Shared nearest neighbors match kernel for bird songs identification -LifeCLEF 2015 challenge

International audienceThis paper presents a new fine-grained audio classification technique designed and experimented in the context of the LifeCLEF 2015 bird species identification challenge. Inspired by recent works on fine-grained image classification, we introduce a new match kernel based on the shared nearest neighbors of the low level audio features extracted at the frame level. To make such strategy scalable to the tens of millions of MFCC features extracted from the tens of thousands audio recordings of the training set, we used high-dimensional hashing techniques coupled with an efficient approximate nearest neighbors search algorithm with controlled quality. Further improvements are obtained by (i) using a sliding window for the temporal pooling of the raw matches (ii) weighting each low level feature according to the semantic coherence of its nearest neighbors. Results show the effectiveness of the proposed technique which ranked 2nd among the 7 research groups participating to the LifeCLEF bird challenge

Spatially Localized Visual Dictionary Learning

International audienceThis paper addresses the challenge of devising new representation learning algorithms that overcome the lack of interpretability of classical visual models. Therefore, it introduces a new recursive visual patch selection technique built on top of a Shared Nearest Neighbors embedding method. The main contribution of the paper is to drastically reduce the high-dimensionality of such over-complete representation thanks to a recursive feature elimination method. We show that the number of spatial atoms of the representation can be reduced by up to two orders of magnitude without much degrading the encoded information. The resulting representations are shown to provide competitive image classification performance with the state-of-the-art while enabling to learn highly interpretable visual models

Recognizing Thousands of Legal Entities through Instance-based Visual Classification

International audienceThis paper considers the problem of recognizing legal en-tities in visual contents in a similar way to named-entity recognizers for text documents. Whereas previous works were restricted to the recognition of a few tens of logotypes, we generalize the problem to the recognition of thousands of legal persons, each being modeled by a rich corporate identity automatically built from web images. We intro-duce a new geometrically-consistent instance-based classifi-cation method that is shown to outperform state-of-the-art techniques on several challenging datasets while being much more scalable. Further experiments performed on an au-tomatic web crawl of 5,824 legal entities demonstrates the scalability of the approach

H3.1K27me1 maintains transcriptional silencing and genome stability by preventing GCN5-mediated histone acetylation

Epigenetic mechanisms play diverse roles in the regulation of genome stability in eukaryotes. In Arabidopsis thaliana, genome stability is maintained during DNA replication by the H3.1K27 methyltransferases ARABIDOPSIS TRITHORAX-RELATED PROTEIN 5 (ATXR5) and ATXR6, which catalyze the deposition of K27me1 on replication-dependent H3.1 variants. The loss of H3.1K27me1 in atxr5 atxr6 double mutants leads to heterochromatin defects, including transcriptional de-repression and genomic instability, but the molecular mechanisms involved remain largely unknown. In this study, we identified the transcriptional co-activator and conserved histone acetyltransferase GCN5 as a mediator of transcriptional de-repression and genomic instability in the absence of H3.1K27me1. GCN5 is part of a SAGA-like complex in plants that requires the GCN5-interacting protein ADA2b and the chromatin remodeler CHR6 to mediate the heterochromatic defects in atxr5 atxr6 mutants. Our results also indicate that Arabidopsis GCN5 acetylates multiple lysine residues on H3.1 variants, but H3.1K27 and H3.1K36 play essential functions in inducing genomic instability in the absence of H3.1K27me1. Finally, we show that H3.1K36 acetylation by GCN5 is negatively regulated by H3.1K27me1 in vitro. Overall, this work reveals a key molecular role for H3.1K27me1 in maintaining transcriptional silencing and genome stability in heterochromatin by restricting GCN5-mediated histone acetylation in plants

The histone H3.1 variant regulates TONSOKU-mediated DNA repair during replication

The tail of replication-dependent histone H3.1 varies from that of replication-independent H3.3 at the amino acid located at position 31 in plants and animals, but no function has been assigned to this residue to demonstrate a unique and conserved role for H3.1 during replication. Here, we show that TONSOKU (TSK/TONSL), which rescues broken replication forks, specifically interacts with H3.1 via recognition of alanine 31 by its tetratricopeptide repeat domain. Our results indicate that genomic instability in the absence of ATXR5/ATXR6-catalyzed H3K27me1 in plants depends on H3.1, TSK and DNA polymerase theta (Pol θ). Overall, this work reveals an H3.1-specific function during replication and the common strategy used in multicellular eukaryotes for regulating post-replicative chromatin maturation and TSK, which relies on histone mono-methyltransferases and reading the H3.1 variant

Innovative Solution for Building Integrated Photovoltaics

Among the main challenges of our century, the climate change and the need of diversification of the energy sources are of most importance. Renewable energies undoubtedly have an important role to play, photovoltaic (PV) electricity being especially well suited to face these energy challenges. However, the current integration of PV panels often comes without architectural consideration. In this context, the Archinsolar project [1] aims to develop a new generation of photovoltaic elements based on silicon thin films technologies (amorphous and micromorph), ultra-reliable and manufacturable at a very low cost, allowing a unique architectural integration, respectful of the bui lt environment and overall landscape. Here we will present our new developments on innovative PV elements including colored PV panels and a solar tile using a composite back-structure

Armadillo 1.1: An Original Workflow Platform for Designing and Conducting Phylogenetic Analysis and Simulations

In this paper we introduce Armadillo v1.1, a novel workflow platform dedicated to designing and conducting phylogenetic studies, including comprehensive simulations. A number of important phylogenetic and general bioinformatics tools have been included in the first software release. As Armadillo is an open-source project, it allows scientists to develop their own modules as well as to integrate existing computer applications. Using our workflow platform, different complex phylogenetic tasks can be modeled and presented in a single workflow without any prior knowledge of programming techniques. The first version of Armadillo was successfully used by professors of bioinformatics at Université du Quebec à Montreal during graduate computational biology courses taught in 2010–11. The program and its source code are freely available at: <http://www.bioinfo.uqam.ca/armadillo>