Transposable elements (TEs) make up half of our genome and induce DNA damages when active. In recent years, an adaptive immune system against TE transposition has been uncovered. In the germline, a dedicated pathway based on small non-coding piRNAs targets TEs at the transcriptional and post-transcriptional levels through base-pair complementarity. The piRNAs are produced by discrete genomic loci called piRNA clusters, which are made of multiple and intermingled TE fragments and constitute the memory of this immune system. Our global objective is to understand how this defense system is generated and how it evolves and adapts to novel TE invasions and external stresses, without triggering an “autoimmune” response against endogenous genes. Based on findings from a previous ANR project, our main hypothesis is that DNA loci encoding for tRNA (tDNA) and subtelomeric sequences play major roles in the genesis of such genome defenses both at short and long term. We will focus on Drosophila oogenesis, in particular on one of its periods of which we have previously discovered the importance for the control of TEs, the "pilp" (piwiless pocket). Our first objective is to test whether clustered transfer RNA (tRNA) loci (tDNA clusters) and subtelomeric regions of chromosomes can induce piRNA cluster formation by promoting multiple TEs insertions in their vicinity. Our second objective is to identify the cis- and trans-acting factors required for the activation and maintenance of these nascent or dormant piRNA clusters. In particular, we will analyze maternally inherited piRNAs and their involvement in the initiation and/or maintenance of the activity of piRNA clusters. On the other hand, a Histone DeMethylase (HDM) recently discovered in the laboratory for its role in maintaining the extinct or dormant state of piRNA clusters will be characterized in detail in this biopiC project using multi-omic approaches. Finally, we have a unique tool that allows us to turn a piRNA cluster ON or OFF without affecting the levels of its primary transcripts. We wish to investigate the effect of RNA methylation on the quality/stability of these primary transcripts and establish a molecular mechanism linking the activity or inactivity of these piRNA clusters as recently suggested by our preliminary data. Our project is both original and ambitious. In the past decade, many studies have sought to describe the biogenesis and functions of piRNAs in the repression of TEs. Here, we rather focus on studying the genomic plasticity of this “immune system” and the complementary functions of TEs and piRNAs in the birth and activation/maintenance of piRNA clusters, respectively. Our consortium is built with the partners that participated in ANR “PlasTiSiPi” (PRC-2013). Under this program, members of the consortium published and co-signed 20 peer-reviewed articles or reviews in high impact journals. The success of the consortium is based on the complementarity of expertises between the labs in the fields of genetics, epigenetics, cell biology and bioinformatics tools development and analysis as well as on frequent meetings between partners. The complementarity between our labs allows us to propose an integrated project from biochemistry to bioinformatics. Our consortium is among the pioneers in the field, and our tools, generated in a previous program, give us a strong competitive advantage. Defending against TEs and adapting to external stresses are problems that almost all organisms face. Thus, our results will benefit basic research as well as reproductive medicine and oncology.

Cocaine Use Disorder (CUD) is a major public health problem. CUD affects 3% of the general population over a lifetime and is associated with a high mortality rate. Cocaine is the most rapidly addictive substance. Its use, including crack cocaine, has been on the rise in Europe over the last ten years. This led to an increase in admissions for cocaine detoxification in specialized clinics. Unfortunately, neither biomarkers nor validated specific medication are available for CUD. A recent model proposes that CUD can be due to compulsive drug use caused by an imbalance between goal-directed behaviors (GDB) and habits (HB). The GDB/HB balance is finely tuned by dopaminergic and acetylcholine nicotinic transmissions in the caudate nucleus and in the putamen. However, human brain imaging of the striatum in CUD produced inconsistent results because of difficulties to finely analyze these striatal areas. Furthermore, several clinical trials conducted with cholinergic drugs yielded provided inconsistent results. The interpretation of these clinical studies is limited because of their short durations, very small sample sizes, and the paucity of analyzed clinical data. Our recent findings in transgenic rodents (self-administration, brain imaging, pharmacology) as well as in patients (clinical genetics, case/control brain imaging) confirms the pivotal involvement of the nicotinic cholinergic regulation of dopamine signaling in CUD. We recently identified a genetic variant of the alpha5 subunit of the nicotinic receptor (alpaha5SNP) in a population of patients with substance use disorders. We obtained a rat line expressing the alpha5SNP. The central objective of our proposal is to better characterize the impact of the a5SNP variant on compulsive cocaine-self administration, focusing on the striatal function. We will determine the morphology (structure and connectivity) of striatal subregions and on acetylcholine/dopamine balance in the caudate and the putamen of our unique humanized rat model. Based on recent results from the consortium we will assess a novel treatment of CUD in rat model with the cholinesterase inhibitor donepezil. We will also expand our investigations to patients hospitalized for severe CUD. To parallel rodent experiments, we will characterize the structure and resting-state functional connectivity of different regions of the striatum as a function of alpha5SNP genotypes and CUD severity in 68 patients. Finally, we will also test the effect of a donepezil in a long-term (6-month) randomized controlled trial in 120 CUD patients. The phenotypic characterization of the two cohorts will also parallel rodent experiments. It includes extensive clinical data about compulsive cocaine use, the trajectory of acquisition of CUD, and diagnostic criteria for CUD corresponding to addiction-like behavior. The feasibility of this ambitious translational research program is excellent, thanks to the complementary expertise and long-standing collaboration of the consortium. We already obtained solid preliminary preclinical and partly acquired clinical human data supporting the project. This project could open the way to increased understanding of CUD and consequently lead to the development of novel therapeutical avenues. The expected results are far-reaching, numerous possibilities for valorization in neuroscience and patient care.