Projects
The genetic control of the development of the vertebrate nervous system is the principal focus of the group headed by Patrick Charnay. Two major orientations are addressed:
Hindbrain patterning
Hindbrain morphogenesis involves a segmentation process that leads to the formation of successive, homologous territories along the anterior-posterior (AP) axis. This developmental process is highly conserved during vertebrate evolution, allowing us to use mouse, chick or zebrafish as model systems.
To unravel the molecular and cellular mechanisms governing the establishment of specific territories within the developing embryo, we have been concentrating on one gene, Krox20. Krox20 encodes a transcription factor, which delimits and specifies hindbrain segments 3 and 5, controlling the expression of numerous other regulatory genes, including Hox genes. More specifically, to understand the basis of the delimitation of segmental territories, we study the control of the expression of Krox20 itself. We have deciphered the regulatory network responsible for the formation of segment 3: Hox genes of the paralogous group 1, through both direct activating and indirect repressive functions, grossly pattern the activity of one of the Krox20 enhancers along the AP axis; this activity is then tuned in a spatially uniform manner by FGF signalisation; the final positioning of the limits of Krox20 expression requires the interpretation of this initial pattern by Krox20 positive feedback loop, orchestrated by a distinct enhancer, and acting as a binary molecular switch. Altogether, this study offers a comprehensive view of the delimitation of a territory and shows how positional information provided by different patterning mechanisms is integrated through a gene regulatory network involving several cis-acting elements operating on the same gene.
Current work is designed to characterize the molecular mechanisms responsible for novel, in cis, genetic interactions that we have recently observed between two Krox20 enhancers. For this purpose, we introduce directed mutations in the zebrafish using the CRISP technology and combine this approach with chromatin immunoprecipitation (ChIP) and chromosome conformation capture (3C).
Peripheral stem cells
We study a cell population, boundary cap (BC) cells, located at the CNS/PNS interface, at the entry and exit points of peripheral nerves. These cells derive from the neural crest and we have shown that they are themselves able to migrate within the embryo, to give rise to different cell types, including stem cells. We have identified several genes specifically expressed in BC cells and developed genetic tools that allow to trace BC cell derivatives in the mouse. BC cells give rise to the entire Schwann cell component of nerve roots, as well as part of the glial satellite cells, and nociceptive and proprioceptive neurons in dorsal root ganglia (DRG). However, we have also observed that a part of the motor root BC cell derivatives further migrate along peripheral nerves, to reach the skin. Here they give rise to Schwann cells, mainly non-myelinating, localised along the hypodermis nerves and at the level of nerve termini in the dermis. These observations show a dual origin for Schwann cells: the neural crest for those located along the major part of the peripheral nerves, and the BCs for those located at the proximal and distal extremities of the nerves.
We have also demonstrated that a part of BC cell derivatives located in the skin at birth possess stem cell properties. In vitro, these cells can generate spheres that can be propagated over multiple passages and give rise to various cell types (neurons, Schwann cells, myofibroblasts and adipocytes). In vivo, after transplantation into adult DRGs, the skin BC derivatives give rise to glial cells as well as numerous sensory neurons. In conclusion, this work shows the existence in the skin of a population of easily accessible stem cells, with a very interesting regenerative potential Our tools should allow their purification and detailed characterization.
Our present projects precisely concern the further characterization of this novel stem cell population in the skin, with the search for specific molecular markers, the exploration of their survival and differentiation potential in the CNS and the analysis of their possible implication in the development of type 1 Neurofibromatosis, where tumours frequently arise at the level of proximal and distal extremities of peripheral nerves, precisely where BC cell derivatives are found.
