Asthma and cardiovascular (CV) diseases are two common conditions with important public health and economic burden. Despite growing evidence that asthma is associated with increased risk of major CV events, the mechanisms by which asthma may affect the risk for CV events remain poorly understood. In particular, whether asthma and CV diseases share common etiological processes (such as anthropometric, lifestyle, social, environmental and/or genetic factors), or whether CV diseases are a consequence of some asthma characteristics (such as asthma treatments or systemic inflammation) remains unknown. The limited knowledge on this mechanism hampers preventive intervention. We aim to disentangle the complex association between asthma and early markers of CV risk, in order to provide new directions in clinical management of patients with asthma and in preventive intervention to prevent CV comorbidities in asthma. To reach this objective, specific aims are: (1) WP1: To collect new data, the 4th follow-up at 30 years of the EGEA (Epidemiological study on the Genetics and Environment of Asthma) cohort to accurately assess: - anthropometric, lifestyle, social and environmental factors through questionnaire, smartphone application, validated 24-h dietary records, passive sampler for individual NO2 assessment, geographical information system-based models; - treatments, hospitalizations, causes of death and long-term illnesses through data linkage with health administrative databases (SNDS); - phenotypic information through detailed questionnaires and clinical examination of the participants including anthropometric measures, lung function, 6-minute walk test, blood pressure and markers of cardiovascular risk (aortic pulse wave velocity [aPWV], Coronary Calcium Score (CAC) assessed by computed tomographic scanner, and biomarkers providing prognostic information on CV risk that will be measured longitudinally (hypersensitive CRP, IL6, hypersensitive Troponine (I), NT-proBNP, and soluble ST2)); (2) WP2: To characterize the longitudinal association of asthma and asthma specific phenotypes with markers of CV risk (aPWV, CAC and biomarkers of CV risk); (3) WP3: To clarify the causal association between asthma and markers of CV risk. Two alternative explanations will be investigated: 1) how much the co-occurrence of asthma and markers of CV risk might be explained by the presence of shared anthropometric, lifestyle, social environmental and genetic common causes; and 2) the role of asthma on markers of CV risk through causal direct and indirect effects (e.g. mediated by asthma treatments or inflammatory pathways). One major asset of the EGEA_30years program is the new follow-up at 30 years of a unique existing cohort including a group of asthma cases recruited in chest clinics, their first-degree relatives and a group of controls (total n=2120), particularly well characterized across the different follow-ups (clinical examination, biomarkers, lifestyle, social, environment, genetic (GWAS), epigenetic (methylome) and metabolomic data) and underpinned by a rich biobank. Given the high follow-up rates achieved in previous EGEA follow-ups, the family and multicentric design of the study, we anticipate that ~1300 individuals will complete the 30-year follow-up questionnaire including ~1000 participants with a clinical examination. Further strengths relate to the expertise of the consortium in setting-up and coordinating cohorts, in asthma and CV research, in analysing large scale omic and exposome data and in applying advanced statistical techniques including cluster and mediation analyses. Our multidisciplinary program will provide tools for identifying and prioritizing determinants of CV risk in asthma and feed into risk prediction, new directions in clinical management as well as development of preventive interventions in asthma. EGEA_30years may unravel actionable levers of life-threatening CV comorbidities in asthma.

SCIENTIFIC CONTEXT. Unicellular organisms need to adapt to rapid and unanticipated changes in their environment. Spores ensure their survival by encapsulating the genome in a protective configuration while awaiting optimal growth conditions. Indeed, yeast spores enter into a quiescent state with minimal metabolic activity and are surrounded by a thick protective wall. Yet, protecting the integrity of the genome in these conditions involves not only a dramatic decrease in transcriptional activity, an extreme nuclear compaction, but also the capacity to completely revert these processes to allow germination. The mechanisms involved in the establishment of genomic quiescence in spores, the protection of their genome and its reactivation, remain unknown. OBJECTIVE. Several lines of evidence show that chromatin is highly compacted in spores. In addition, histone H4 is hyperacetylated and this modification is essential for spore viability. This observation seems counter-intuitive because H4 acetylation (H4ac) is usually associated with transcription activation and open chromatin. Therefore, the general objectives of this project are to understand (i) the mechanisms by which H4 is hyperacetylated in spores, (ii) how H4ac is compatible with quiescence in spores and their chromatin compaction, (iii) whether and how H4ac prepares genome reactivation observed during early germination. IMPACT. Through EpiSpores, we will improve our general knowledge on H4ac signalling pathways, chromatin organisation and transcription regulation. Furthermore, yeast spores provide an alternative model system to investigate the molecular mechanisms of quiescence entry, maintenance and exit in all eukaryotic cells. Finally, our previous work on yeast spores has been translated to the treatment of fungal infections, collectively responsible for 1.5 millions of deaths per year. Our future work exploring chromatin signalling pathways in Candida albicans and their functional role in the virulence of this pathogenic yeast will be based on the technological development of this proposal. CONSORTIUM. This project brings together young researchers, by academic standards, with collective and synergetic expertise in yeast biology, genetics, biochemistry, interactomics, high-throughput genetics, super-resolution microscopy and epigenomic approaches.

Photoacoustic imaging (PAI) is a biomedical imaging technique that provides optical contrasts at depth through the generation of acoustic waves with light. Handheld systems can be made for 3D navigation to image the vasculature and its oxygenation. However, these systems are impacted by limited view artifacts that hide some structures and by low contrast-to-noise when using a sparse array. Photoacoustic Fluctuation imaging (PAFI) enables to solve these two issues and we will develop further this technique towards quantitative view-full SO2 imaging. One of the challenges is to correct the spectral coloring effects due to the tissue surrounding the vessels. To this end, we will employ a highly novel multi-modal combination. Beyond these advances concerning PAFI, this technique still suffers from its low temporal resolution. Relying on deep neural networks (DNN) image enhancement abilities, we will improve the frame rate towards the ultimate limit of SO2 images obtained from single shot multispectral images. A main challenge for DNN is to provide quantitative predictions. To address this issue, FULBOX proposes novel experimental approaches to enable real-time full-view imaging of blood oxygenation, which, coupled to state-of-the-art Ultrasound Doppler will enrich the diagnosis for several pathologies.

In confined 3D microenvironments’ where metazoan cells move, their nuclear genome scaffolding and dynamic functions are preserved through the nucleus elastic properties conferred by components partitioning between the dense nucleus matrix and the cytoplasm-located skeletons. Specifically the intrinsic high viscosity of the nucleus as compared to other organelles is provided by the chromatin nuclear content together with the lamin-composed intermediate filament meshwork. These elements concur to structure the nuclear inner membrane and maintain the nuclear forces. To cope with the physical constraints the nucleus senses during migration, cells have evolved distinct strategies: several leucocytes migrating through tight spaces show intrinsically flexible nuclei often characterized by their lower lamin content as compared to fibroblasts’ or cancer cells’ nuclei. Nucleus deformability however can also be increased through actin-based forces applied onto the nucleus envelope that promote transient rupture of the lamin shell preferentially at the highest curvature site. Inversely in some cancer cells, lamins A/C were shown to protect the nuclear envelope against curvature-induced rupture, hence preventing loss of DNA repair factors in response either to external probing forces or to contractile acto-myosin forces generated at adhesion sites. The nucleus of Toxoplasma- a protozoan parasite that belongs to the Apicomplexa phylum- significantly deforms when sequentially (a) trafficking in extra-cellular confined matrices (b) then invading metazoan host cells on which relies its fitness. How the parasite proceeds within these various and non-uniform confined microenvironments and in particular how it prevents its nucleus from the threat of mechanical-induced injuries - a prerequisite to progeny production - remains fully elusive. Very little is actually known on the nucleus structure and function and there is yet no evidence of cytoskeleton element connecting the nucleus and the plasma membrane . While nucleus deformation is also prominent over the developmental program of the parasite in its obligate hosting cell. the concept of mechanotransduction has not yet been investigated in any Apicomplexa parasites. The TOXONUC proposal primarily aims at filling key gaps of knowledge on the nuclear mechanics at work during physiologically relevant and confined motions of the protozoan Toxoplasma. To this end, the three member-consortium has been built around a true complementary expertise offering acute expertise in the molecular and cell biology – in particular high resolution and super resolution live imaging - of this single-celled eukaryote but also providing the advantage of combining with the long-standing expertise in plugin design dedicated to high-content 3 and 4D cell imaging – development of the ICY platform- . Adding biophysics concepts and tools with force microscopy in conjunction with nanotechnology and microfluidics at the heart of this program will contribute to give a multi-scale understanding of the integrated molecular structure of the nucleus within the cellular organization, in particular during sensing and response to cellular mechanical forces. In conclusion actions taken by the consortium will allow decoding how the Toxoplasma nuclear genome scaffolding and functions could swiftly operate and be preserved over displacement and deformation but will also pave the way for better knowledge on the nuclear mechanotransduction mechanisms much beyond the field of Toxoplasma.

The ancient phylum Apicomplexa includes many of the world’s pre-eminent protozoan pathogens. Most deadly to humans is Plasmodium, the agent of malaria, which kills around a million people annually. As obligate intracellular parasites, they establish intimate interactions with their hosts. Toxoplasma gondii is an extreme example of this adaptation, able to replicate within nearly every cell type in any warm-blooded host. Not only the developmental program of this parasite in wildlife and livestock animals can result in potentially negative socio-economic impact, but remarkably, in about a third of the human population Toxoplasma also experiences prolonged quasi-silent persistence in tissues such as brain and retina. While the asymptomatic parasitism that proceeds typically offers life-long equilibrium and protection in immune competent hosts, sustained immune dysfunction is known to break parasite dormancy, promoting bradyzoite to tachyzoite transition and further T. gondii tachyzoite population expansion, these combined processes eventually resulting in encephalitis and meningitis as major damages. The strategy of T. gondii as a parasite is based on a quest for avirulence, a capacity to attenuate but not to fully counteract the immune defense of the host, thus securing the permanent residence required to await transmission. HostQuest focuses on elucidating the molecular mechanisms by which T. gondii is orchestrating immune evasion and lifelong persistence in hosts. Once intracellular, parasites actively reprogram gene expression of the immune cells they infect by subverting host transcription factors activity or by modulating the epigenetic status of target genes. Secreted effectors are involved. Those are singularly exported beyond the vacuole-containing parasites and reach the host cell nucleus to reshape the host genetic program. The discovery of new exported Toxoplasma effectors and the characterization of their activities, or the mechanisms by which they are recognized by the host immune system, continues to gather pace. Much has been learnt in recent years but we have only been chipping at the tip of the iceberg. We aim to study the modus operandi of these effectors and particularly their possible implications in immune evasion and parasite persistence. These effectors may adopt at least three alternative, although not mutually exclusive, strategies to subvert host gene expression. They may (i) modulate upstream signaling pathways (ii) directly target host transcription factor protein levels/activity and/or (iii) affect histone packing and chromatin configuration. HostQuest is an interdisciplinary project that aims to: i) Determine the full repertoire of GRA effectors and the magnitude of the changes they are eliciting in the infected cell; ii) Explore their synergistic and/or antagonist effects on gene regulation; iii) Decipher the extent to which they contribute to immune evasion and/or sustained parasitism; iv) Gain knowledge of the three-dimensional structure of effectors in complex with host cell factors in order to understand the protein function or to guide further experiments to investigate function. Studies of effectors also continue to offer opportunities for the development of tools to probe host cell biology in the absence of disease. In this respect, HostQuest is also poised to exploit Toxoplasma molecular intelligence developed over million years of co-evolution with its hosts to learn new lessons on the mechanisms regulating cell homeostasis and their alterations in host cells, including cancer cells.