From neural precursors to glial cells – Epinest

Nervous system, which constitutes the most complex tissue in our entire body, allows us to respond and adjust to the environment. Its sophisticated and stereotypic architecture relies on the differentiation of neurons and glia at specific times and positions from neural stem cells, a source and a reservoir of multipotent precursors that can be reprogrammed and persist throughout the entire life. Neural stem cells have also recently drawn much attention in medical science for their potential in curing neurodegenerative diseases. Their interest in cell replacement therapies, however, heavily relies on our understanding of the mechanisms triggering their differentiation in specific cell types. Recent data show that transcriptional regulation, responsible for the establishment and maintenance of cell identities, is accompanied by chromatin modifications. The current view is that pluripotency of stem cells is associated with an unfettered and plastic genome, accessible to transcriptional regulation and thus poised to differentiate into different cell types. As pluripotent cells proliferate and are specified into tissue lineages, they selectively activate specific subsets of genes. Regions of the genome that are not involved in the emerging cell lineage become silenced and thereafter remain inaccessible to transcriptional regulation. A number of histone modifications, including acetylation, methylation, phosphorylation, ubiquitination, sumoylation and ADP-ribosylation, regulate gene expression and chromatin structure. Histone acetylation represents a signature of an active transcriptional state, its regulation accompanying vertebrate neuronal and glial differentiation in vitro. Simple tissue and genome organization, together with the evolutionary conservation of basic processes and the available genetic tools make Drosophila an organism of choice for studying neural development. Drosophila nervous system originates from stereotyped, invariant, lineages that can be easily identified. Fly neural precursors (NPs) self renew and undergo a series of asymmetric divisions to produce neurons and glia, two clear features of stemness. We have previously identified and characterized all glial producing stem cells as well as the Drosophila glial determinant, Glide/Gcm (for the sake of simplicity, Gcm, throughout the text). Gcm codes for a transcription factor that induces gliogenesis at the expense of neuronal differentiation. In the embryo, lack of Gcm leads to loss of most lateral glia, while ectopic Gcm leads to the differentiation of supernumerary glia. Thus, Gcm acts as a binary genetic switch, as its loss leads glia to become neurons and its overexpression is sufficient to induce expression of glial markers and to repress that of neuronal markers. The Gcm gain and loss of function phenotypes indicate a high degree of cell plasticity and the ability of a single transcription factor to trigger a specific developmental pathway. These data allow us to ask a fundamental question: how does Gcm act onto the fate choice' How does it intearct with the epigenetic machinery controling cell identity' We have put forward the following hypotheses: 1) NPs and differentiated cells display specific histone acetylation states. 2) The transcriptional program induced by Gcm involves epigenetic changes (H3K9ac). 3) Gcm induces supernumerary glial differentiation upon NP reprogramming. To test these hypotheses, we have identified four major objectives that make the object of the present proposal. 1) To produce a Gcm specific marker by a novel transgenic approach (Tagged Gcm BAC). This will allow us to follow the cell in which Gcm induces the fate choice between glia and neurons. A first transgenic line has been obtained. 2) To establish the H3K9 acetylation profile during neural development. We will be able to define the epigenetic state of distinct cell types. Preliminary data suggest a dynamic profile of H3K9ac. 3) To follow the H3K9ac and the NP fate upon Gcm overexpression. This will allow us to define the identity of the challenged cells. Our recent data strongly suggest that Gcm reprograms NPs and that it interacts with an enzyme that triggers histone acetylation (HAT). 4) To identify the direct Gcm targets that are necessary in the fate choice between glia and neurons. This will open novel perspectives and allow us to identify the molecular pathway involved in gliogenesis and cell reprogramming. These data will contribute to understand the mechanisms controlling cell differentiation and plasticity, two key processes during development and in pathological conditions.