Blanc SVSE 3 - Blanc - SVSE 3 - Microbiologie, immunologie, infectiologie

The cellular arm of innate immunity: functional characterisation of the Drosophila adult haematopoietic system. – ADHAEMOS

The cellular arm of innate immunity: functional characterisation of the Drosophila adult haematopoietic system.

All multicellular organisms are confronted with pathogenic microorganisms and tissue damages. A front-line of defence against these threats relies on cells of the haematopoietic system. Drosophila has emerged as a potent model to study haematopoietic cell development and features. While much attention has been paid to embryonic and larval blood cells, the origin, fate, roles and regulation of adult haemocytes are poorly described.

Functional characterisation of the Drosophila adult haematopoietic system.

Like their mammalian counterpart of the monocyte/macrophage system, adult haemocytes participate in the cellular immune response. Yet, the limited description of the adult haematopoietic system hampers the study of these cells function in immune response or in other processes. <br />The main objective of this research project is to obtain an integrated view of Drosophila adult haematopoietic system that encompasses its ontogeny, its functions and its regulation. Accordingly, we will seek to establish the diversity and origin of the blood cells present in adult flies. We will determine whether active haematopoiesis (blood cell proliferation and differentiation) takes place during adulthood. We will study the dynamic and regulation of adult blood cells in response to different infections and during aging. Finally, we will identify key genes regulating blood cell development or function and characterize their mode of action.

This project heavily relies on the analysis of the expression of blood cell markers (antibodies; GFP/RFP transgenic reporter lines, available or generated for the project) by epifluorescent and confocal imaging in vivo, on tissue sections or on isolated blood cells.
An other part of the project requires the prospective isolation of different blood cell populations by FACS followed by gene expression profiling by RNAseq.
We will use the genetic toolkit available in Drosophila to perform cell lineage analysis, assess hemocyte cell cycle status and study the function of key genes in the development and/or function of adult blood cells (e.g.: using UAS-dsRNA or mutant lines).
Finally, we will infect Drosophila with different pathogens to study adult blood cells reaction to immune challenges.

Our results show that distinct population of hemocytes are present in adult Drosophila. Notably, we observed different population of macrophages as well as some crystal cells. These cells are derived from the embryonic and larval wave of hematopoiesis and undistinctively in circulation, as isolated sessile cells under the epidermis or concentrate in patches along the heart in the abdomen,
Up to now, all the hemocyte population that we have analyzed tend to decrease during aging. Interestingly though, our data suggest that active haematopoiesis can take place during adulthood as we observed differentiation and found that an immune challenge can elicit some blood cell proliferation.
Finally, we have identified a small population of hemocytes that may represent a stem or progenitor blood cell pool.

As a follow up of our encouraging results, we will focus on the characterization of the prospective hematopoietic progenitors. In parallel, we will pursue the analysis of the response of the adult blood cell to different immune challenges and we will try to assess the respective function of different blood cell populations in the cellular immune response. Finally, we will try to define at the molecular level the identity and specificity of embryo versus larval derived blood cells. We will then elicit the functional analysis of genes potentially regulating adult blood cell fate or function.

Oral (2) and poster (2) communications at conferences.

All multicellular organisms are confronted with pathogenic microorganisms and tissue damages. A front-line of defence against these threats relies on cells of the haematopoietic system. These specialised cells arise from common haematopoietic progenitors specified during early development. They are maintained throughout adulthood to perform essential functions such as establishing the proper response against invading pathogens, removing cancerous and apoptotic cells, or producing clotting factors. In humans, deregulation of their functions or development is the cause of several disorders including cancers, increased susceptibility to infections or inflammatory diseases. Hence, unravelling how their development, homeostasis and functions are regulated is of paramount importance.
Model organisms have greatly contributed to our understanding of the development and regulation of the human immune system. Notably, seminal studies of Drosophila melanogaster humoral immune response have paved the way for the characterisation of the inflammatory response pathways in mammals. More recently, works from different labs including ours have shown that several aspects of blood cell development and functions have been conserved from Drosophila to human. Indeed Drosophila possesses three myeloid-like cell types (haemocytes) and many genes employed in mammalian blood cells have similar functions in fly. Hence, Drosophila has emerged as a potent model to study haematopoietic cell development and features. While much attention has been paid to embryonic and larval blood cells, the origin, fate, roles and regulation of adult haemocytes are poorly described. Like their mammalian counterpart of the monocyte/macrophage system, adult haemocytes participate in the cellular immune response notably by phagocyting microorganisms and some results suggest that they participate in tolerance to infection or mediate immune priming. Yet, these studies have been hampered by the limited description of the adult haematopoietic system. For instance, while adult haemocytes were believed to consist of a single cell type (macrophages), recent evidence shows that this population is heterogeneous. Moreover, our unpublished observations indicate that there is a greater diversity of blood cell types and that progenitor blood cells may be present in the adult. In contrast to the highly dynamic embryonic and larval stages that span a relatively short time, fly adulthood offers a post-developmental set-up for analysing various aspects of immune cell biology such as stem/progenitor cell maintenance, immune senescence/aging or inflammation. Here, we wish to establish the adult fly as a model system for studying blood cells and characterising the innate cellular immune response.
Besides the conservation of the gene networks employed in immune cells, Drosophila provides a powerful collection of genetic and genomic tools as well as cell lineage analysis techniques that are perfectly suited for studying adult blood cells in vivo. By combining complementary approaches (intravital imaging, lineage tracing, transcriptomics and functional genomics) and different physiological contexts, we will aim at characterising Drosophila adult haematopoietic system. The first goal of this project will be to assess the origin and diversity of the adult blood cells. Second, we will explore the mechanisms underlying blood cells maintenance and immunosenescence during adulthood. Third we will investigate the dynamics and regulation of adult haemocytes in response to different infections. Finally, we will identify key genes that regulate adult blood cell behaviour and characterise their functions. All together, these experiments should increase our understanding of the molecular mechanisms underlying the development and functions of myeloid-like cells in vivo and accelerate the discovery of genetic networks that control the cells of the innate immune system in other organisms, including humans.

Project coordinator

Monsieur Lucas Waltzer (Centre de Biologie du Développement) – lucas.waltzer@uca.fr

The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.

Partner

CBD Centre de Biologie du Développement

Help of the ANR 289,948 euros
Beginning and duration of the scientific project: January 2014 - 42 Months

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