La PF Transcriptome et Epigenome développe des projets de séquençage à haut débit (collaboration et service) avec des équipes du Campus. Ceux-ci couvrent l'ensemble des thématiques du campus ainsi qu'une large gamme d'organismes (des virus aux mammifères). La plate-forme exerce des activités de biologie humide (construction des librairies et séquençage) et de biologie sèche (analyse bioinformatiques et statistiques). La personne mise a disposition interagira étroitement avec les autres bioinformaticiens du pôle BioMics et du Hub. Ses activités concerneront notamment: - La participation à la conception et à la mise en place des projets avec les équipes demandeuses, la prise en charge des analyses et le reporting aux utilisateurs - La mise en place d'un workflow d'analyse bioinformatique des données de transcriptome /épigénome en étroite collaboration avec le C3BI, la DSI et les autres bioinformaticiens du pole. Ce workflow permettra le contrôle qualité des données, leur prétraitement, le mapping des séquences sur les génomes/transcriptomes de réference, et le comptage des reads pour les différents éléments de l'annotation - L'adaptation du workflow d'analyse aux questions biologiques et aux organismes étudiés dans le cadre des activités de la PF - L'activité de veille technologique et bibliographique (test et validation de nouveaux outils d'analyse, updates d'outils existants...) - La mise en place et le développement d'outils d'analyse adaptés aux futurs projets de la PF: single cell RNAseq, métatranscriptome, ChIPseq, analyse des isoformes de splicing.. Ceci se fera notamment via la réalisation d'analyses dédiées avec certains utilisateurs. Les outils mis en place et validés dans ce cadre seront ensuite utilisés pour l'ensemble des projets. - L'activité de communication et de formation (participation aux réunions du consortium France Génomique,formation permanente à l' Institut Pasteur… - la participation a d autres projets du Pole BioMics (selon disponibilité) Bernd Jagla, qui était le bioinformaticien de la plateforme a rejoint le Hub au 1er janvier 2016. Rachel Legendre est mise a disposition depuis le 2 novembre 2015 et remplace Bernd Jagla. Je souhaite que Rachel Legendre soit mise à disposition de la plateforme pour une durée d'au moins 2 ans.

In Plasmodium falciparum a virulence gene family with 60 var genes codes for the PfEMP1 surface proteins, which undergo antigenic variation. This epigenetically controlled mechanism promotes immune evasion of the parasite. As ncRNAs are emerging as relevant regulators of gene expression we are investigating a GC-rich ncRNA gene family consisting of 15 highly homologous members all positioned next to var gene clusters. We recently found that GC-rich ncRNA transcript associates with the distinct nuclear expression site in which a single var gene is transcribed. We hypothesize that GC-rich ncRNAs interact with the nascent mRNA of var virulence genes. We intend to predict potential RNA-RNA interaction with thermodynamic calculation provided by the RNAup software. In silico prediction of potential binding sites and strength of interaction will help to generate hypotheses and inform our further experimental design.

We study the regulation of type I interferon (IFN) response in humans and in particular the functioning of a key negative feedback regulator, USP18. A recent article reported on a predicted 897nt-long LincRNA (long intergenic non coding RNA) that may target USP18. We have collected information for this LincRNA (genomic locus, putative transcript variants, sequence similarities with USP18 RNA, predicted ORFs, homologies with other genes, etc). To draw a guideline for wet lab experiments, we need to mine databases on this LincRNA. We expect to find it up-regulated in conditions of inflammation or in specific immune cell subsets, like macrophages. We wish to interrogate existing RNA-seq and Chip-seq human databases to obtain information on expression, transcriptional regulation, function, tissue-specificity or relation with pathological conditions of this LincRNA.

Entamoeba histolytica is a protozoan parasite and an amitochondriate pathogenic amoeba, which causes amoebiasis (dysentery and liver abscess) in humans. In addition to E. histolytica several species infect the human intestine although these do not cause disease and include in most of cases E. dispar and ocassionnally E. moshkovskii. A phylogenetically close Entamoeba, E. invadens infecting snails, is used as cellular model for Entamoeba cyst formation.

Supported by the National Agency for Research (ANR-10-GENM-0011) we developed a project to firstly study the transcriptional landscape of pathogenic E. histolytica. Among the results we discovered that 60% of ORFs present anti-sense RNAs (NATs) that map to the 3‘ end of genes. Their regulation is modified upon environmental changes. The regulation of NATs is basically governed by genomic sequences within the very short intragenic region of the amoeba genome. Secondly, we have started to conduct comparative genomics and transcriptomics approaches to understand phenotypic differences between Entamoeba species, in particular with respect to virulence.

This research project focuses on the characterization of the role of small RNAs and their associated Argonaute proteins in transcriptional and post-transcriptional regulation of germline gene expression. Using the nematode C. elegans, we have recently showed that one of the germline-expressed Argonaute protein, CSR-1, promotes germline transcription. However, CSR-1 also possess an endonucleolytic activity that might participate in post-transcriptional silencing. Therefore, two possible functions of the protein might regulate the germline transcriptome. 1) CSR-1 promotes specific germline transcription programs in the nucleus, and 2) negatively regulates expression of target transcripts in the cytoplasm. To gain mechanistic insights into these two functions, we aim to use RNA-seq, sRNA-seq, ChIP-seq, GRO-seq, Ribo-seq, RIP-seq, iCLIP in wild type worms, knock out and catalytic inactive mutants of CSR-1 protein at different times of germline development.

The canonical (“textbook”) process of DNA homology search and recognition is initiated by DNA double-strand breaks and is mediated by the universally conserved recombinases of the RecA family. Using the phenomenon “Repeat Induced Point mutation” (RIP) in N. crassa as a model system, we have previously revealed the existence of another way to search for DNA homology, which does not require RecA proteins and which apparently operates on intact DNA double helices. This pathway can be extremely efficient, as it allows some fungi to detect the presence of only two gene-sized DNA repeats in the genome. Our current work on Meiotic Silencing by Unpaired DNA (MSUD) has shown that the same recombination-independent pathway may also be involved in the early steps of homologous chromosome pairing in meiosis, thus emerging as a conserved, perhaps fundamental mechanism of DNA homology search and recognition. We are now interested in further investigating this mechanism, using RIP and MSUD as two complementary recombination-independent processes. Specifically for this project, we are interested in identifying molecular genetic features that associate with (or even trigger) the production of small RNAs during MSUD in N. crassa.

Despite the worldwide importance of malaria due to Plasmodium vivax, there is currently almost no data on the molecular responses of the Anopheles mosquito vectors to this parasite species. Understanding these responses will contribute to identify relevant strategies to interrupt the transmission of P. vivax by targeting the mosquito vector. Such approaches are urgently needed, as P. vivax is difficult to target on the long term in humans as a consequence of the hypnozoite stage that is responsible for relapses. This project will investigate the molecular response of Anopheles arabiensis, member of the Anopheles gambiae complex, to P. vivax from experimental infections performed in Madagascar where both mosquito and parasite are present. This is a unique situation that will capitalize on the strong knowledge and tools available for An. gambiae sensu lato. The response will be compared to the one triggered by P. falciparum, also present in Madagascar. From mosquitoes infected in a field setting, a transcriptomic (RNAseq) approach will be used to identify common and unique pathways to both parasite species. These analyses will further contribute to identify targets for interrupting transmission of each or both parasites simultaneously. However, a pilot study performed with the PF transcriptomics & epigenomics revealed that the current release of the An. arabiensis genome is poorly annotated. Therefore, to be able to make sens of the RNAseq analyses per se, a strong support from the C3BI Hub for bioinformatics and biostatistics is critical.

Pathogen leptospires are responsible for the zoonotic disease leptospirosis. During infection, Leptospira are confronted with dramatic adverse environmental changes such as deadly reactive oxygen species (ROS). Defenses against ROS, e.g. peroxidase activity, are crucial for Leptospira virulence and the adaptive response to ROS is controlled by PerR regulators. We aim at studying how small non-coding RNA participate in the adaptive response to oxidative stress in pathogen Leptospira.