Françoise Trousse

Induction of oligodendrocyte progenitors in the trunk neural tube by ventralizing signals: effects of notochord and floor plate grafts, and of sonic hedgehog.

Recent evidence indicates that oligodendrocytes originate initially from the ventral neural tube. We have documented in chick embryos the effect of early ventralization of the dorsal neural tube on oligodendrocyte differentiation. Notochord or floor plate grafted at stage 10 in dorsal position induced the development of oligodendrocyte precursors in the dorsal spinal cord. In vitro, oligodendrocytes differentiated from medial but not intermediate neural plate explants, suggesting that the ventral restriction of oligodendrogenesis is established early. Furthermore, quail fibroblasts overexpressing the ventralizing signal Sonic Hedgehog induced oligodendrocyte differentiation in both the intermediate neural plate and the E4 dorsal spinal cord. These results strongly suggest that the emergence of the oligodendrocyte lineage is related to the establishment of the dorso-ventral polarity of the neural tube.

SFRP1 regulates the growth of retinal ganglion cell axons through the Fz2 receptor

Axon growth is governed by the ability of growth cones to interpret attractive and repulsive guidance cues. Recent studies have shown that secreted signaling molecules known as morphogens can also act as axon guidance cues. Of the large family of Wnt signaling components, only Wnt4 and Wnt5 seem to participate directly in axon guidance. Here we show that secreted Frizzled-related protein 1 (SFRP1), a proposed Wnt signaling inhibitor, can directly modify and reorient the growth of chick and Xenopus laevis retinal ganglion cell axons. This activity does not require Wnt inhibition and is modulated by extracellular matrix molecules. Intracellularly, SFRP1 function requires Gα protein activation, protein synthesis and degradation, and it is modulated by cyclic nucleotide levels. Because SFRP1 interacts with Frizzled-2 (Fz2) and interference with Fz2 expression abolishes growth cone responses to SFRP1, we propose a previously unknown function for this molecule: the ability to guide growth cone movement via the Fz2 receptor.

Proper patterning of the optic fissure requires the sequential activity of BMP7 and SHH

The optic disc develops at the interface between optic stalk and retina, and enables both the exit of visual fibres and the entrance of mesenchymal cells that will form the hyaloid artery. In spite of the importance of the optic disc for eye function, little is known about the mechanisms that control its development. Here, we show that in mouse embryos, retinal fissure precursors can be recognised by the expression of netrin 1 and the overlapping distribution of both optic stalk (Pax2, Vax1) and ventral neural retina markers (Vax2, Raldh3). We also show that in the absence of Bmp7, fissure formation is not initiated. This absence is associated with a reduced cell proliferation and apoptosis in the proximoventral quadrant of the optic cup, lack of the hyaloid artery, optic nerve aplasia, and intra-retinal misrouting of RGC axons. BMP7 addition to organotypic cultures of optic vesicles from Bmp7-/- embryos rescues Pax2 expression in the ventral region, while follistatin, a BMP7 antagonist, prevents it in early, but not in late, optic vesicle cultures from wild-type embryos. The presence of Pax2-positive cells in late optic cup is instead abolished by interfering with Shh signalling. Furthermore, SHH addition re-establishes Pax2 expression in late optic cups derived from ocular retardation (or) embryos, where optic disc development is impaired owing to the near absence of SHH-producing RGC. Collectively, these data indicate that BMP7 is required for retinal fissure formation and that its activity is needed, before SHH signalling, for the generation of PAX2-positive cells at the optic disc.

SFRP1 modulates retina cell differentiation through a beta-catenin-independent mechanism.

Secreted frizzled related proteins (SFRPs) are soluble molecules capable of binding WNTS and preventing the activation of their canonical signalling cascade. Here we show that Sfrp1 contributes to chick retina differentiation with a mechanism that does not involve modifications in the transcriptional activity of β-catenin. Thus, addition of SFRP1 to dissociated retinal cultures or retroviral mediated overexpression of the molecule consistently promoted retinal ganglion and cone photoreceptor cell generation, while decreasing the number of amacrine cells. Measure of the activity of the β-catenin-responsive Tcf-binding site coupled to a luciferase reporter in transiently transfected retinal cells showed that Sfrp1 was unable to modify the basal β-catenin transcriptional activity of the retina cells. Interestingly, a dominant-negative form of GSK3β gave similar results to those of Sfrp1, and a phosphorylation-dependent inhibition of GSK3β activity followed SFRP1 treatment of retina cells. Furthermore, retroviral mediated expression of a dominant-negative form of GSK3β induced a retina phenotype similar to that observed after Sfrp1 overexpression, suggesting a possible involvement of this kinase in SFRP1 function.

Forcing neural progenitor cells to cycle is insufficient to alter cell-fate decision and timing of neuronal differentiation in the spinal cord

BackgroundDuring the development of the nervous system, neural progenitor cells can either stay in the pool of proliferating undifferentiated cells or exit the cell cycle and differentiate. Two main factors will determine the fate of a neural progenitor cell: its position within the neuroepithelium and the time at which the cell initiates differentiation. In this paper we investigated the importance of the timing of cell cycle exit on cell-fate decision by forcing neural progenitors to cycle and studying the consequences on specification and differentiation programs.ResultsAs a model, we chose the spinal progenitors of motor neurons (pMNs), which switch cell-fate from motor neurons to oligodendrocytes with time. To keep pMNs in the cell cycle, we forced the expression of G1-phase regulators, the D-type cyclins. We observed that keeping neural progenitor cells cycling is not sufficient to retain them in the progenitor domain (ventricular zone); transgenic cells instead migrate to the differentiating field (mantle zone) regardless of cell cycle exit. Cycling cells located in the mantle zone do not retain markers of neural progenitor cells such as Sox2 or Olig2 but upregulate transcription factors involved in motor neuron specification, including MNR2 and Islet1/2. These cycling cells also progress through neuronal differentiation to axonal extension. We also observed mitotic cells displaying all the features of differentiating motor neurons, including axonal projection via the ventral root. However, the rapid decrease observed in the proliferation rate of the transgenic motor neuron population suggests that they undergo only a limited number of divisions. Finally, quantification of the incidence of the phenotype in young and more mature neuroepithelium has allowed us to propose that once the transcriptional program assigning neural progenitor cells to a subtype of neurons is set up, transgenic cells progress in their program of differentiation regardless of cell cycle exit.ConclusionOur findings indicate that maintaining neural progenitor cells in proliferation is insufficient to prevent differentiation or alter cell-fate choice. Furthermore, our results indicate that the programs of neuronal specification and differentiation are controlled independently of cell cycle exit.

Neurogenic role of Gcm transcription factors is conserved in chicken spinal cord.

Although glial cells missing (gcm) genes are known as glial determinants in the fly embryo, the role of vertebrate orthologs in the central nervous system is still under debate. Here we show for the first time that the chicken ortholog of fly gcm (herein referred to as c-Gcm1), is expressed in early neuronal lineages of the developing spinal cord and is required for neural progenitors to differentiate as neurons. Moreover, c-Gcm1 overexpression is sufficient to trigger cell cycle exit and neuronal differentiation in neural progenitors. Thus, c-Gcm1 expression constitutes a crucial step in the developmental cascade that prompts progenitors to generate neurons: c-Gcm1 acts downstream of proneural (neurogenin) and progenitor (Sox1-3) factors and upstream of NeuroM neuronal differentiation factor. Strikingly, this neurogenic role is not specific to the vertebrate gene, as fly gcm and gcm2 are also sufficient to induce the expression of neuronal markers. Interestingly, the neurogenic role is restricted to post-embryonic stages and we identify two novel brain neuronal lineages expressing and requiring gcm genes. Finally, we show that fly gcm and the chick and mouse orthologs induce expression of neural markers in HeLa cells. These data, which demonstrate a conserved neurogenic role for Gcm transcription factors, call for a re-evaluation of the mode of action of these genes during evolution.

Regenerating Islet-derived 1α (Reg-1α) Protein Is New Neuronal Secreted Factor That Stimulates Neurite Outgrowth via Exostosin Tumor-like 3 (EXTL3) Receptor

Background: Reg-1α is a small secretory protein overexpressed during the early stages of Alzheimer disease. Results: Secreted Reg-1α stimulates axon outgrowth, and this paracrine effect is mediated by its receptor EXTL3. Conclusion: Reg-1α emerges as an important actor in brain plasticity and the regenerative process. Significance: Learning how Reg-1α regulates the nerve cells is important for understanding its implications in early stages of Alzheimer disease. Regenerating islet-derived 1α (Reg-1α)/lithostathine, a member of a family of secreted proteins containing a C-type lectin domain, is expressed in various organs and plays a role in proliferation, differentiation, inflammation, and carcinogenesis of cells of the digestive system. We previously reported that Reg-1α is overexpressed during the very early stages of Alzheimer disease, and Reg-1α deposits were detected in the brain of patients with Alzheimer disease. However, the physiological function of Reg-1α in neural cells remains unknown. Here, we show that Reg-1α is expressed in neuronal cell lines (PC12 and Neuro-2a) and in rat primary hippocampal neurons (E17.5). Reg-1α is mainly localized around the nucleus and at the membrane of cell bodies and neurites. Transient overexpression of Reg-1α or addition of recombinant Reg-1α significantly increases the number of cells with longer neurites by stimulating neurite outgrowth. These effects are abolished upon down-regulation of Reg-1α by siRNA and following inhibition of secreted Reg-1α by antibodies. Moreover, Reg-1α colocalizes with exostosin tumor-like 3 (EXTL3), its putative receptor, at the membrane of these cells. Overexpression of EXTL3 increases the effect of recombinant Reg-1α on neurite outgrowth, and Reg-1α is not effective when EXTL3 overexpression is down-regulated by shRNA. Our findings indicate that Reg-1α regulates neurite outgrowth and suggest that this effect is mediated by its receptor EXTL3.

CXCR7 Receptor Controls the Maintenance of Subpial Positioning of Cajal–Retzius Cells

Cajal-Retzius (CR) cells are essential for cortical development and lamination. These pioneer neurons arise from distinct progenitor sources, including the cortical hem and the ventral pallium at pallium-subpallium boundary (PSB). CXCR4, the canonical receptor for the chemokine CXCL12, controls the superficial location of hem-derived CR cells. However, recent studies showed that CXCR7, a second CXCL12 receptor, is also expressed in CR cells at early developmental stages. We thus investigated the role of CXCR7 during CR cell development using multiple loss-of-function approaches. Cxcr7 gene inactivation led to aberrant localization of Reelin-positive cells within the pallium. In addition, Cxcr7(-/-) mice were characterized by significant accumulation of ectopic CR cells in the lateral part of the dorsal pallium compared with Cxcr4 knockout mice. Loss-of-function approaches, using either gene targeting or pharmacological receptor inhibition, reveal that CXCR7 and CXCR4 act both in CR positioning. Finally, conditional Cxcr7 deletion in cells derived from Dbx1-expressing progenitors indicates an essential role of CXCR7 in controlling the positioning of a subpopulation of PSB-derived CR cells. Our data demonstrate that CXCR7 has a role in the positioning of hem and PSB-derived CR cells, CXCL12 regulating CR cell subpial localization through the combined action of CXCR4 and CXCR7.

Determination of Glial Lineages During Early Central Nervous System Development

The rapidly cycling neuroepithelial cells, located in the ventricular and subventricular zones of the neural tube, give rise to most of the neurons and macroglial cells, astrocytes and oligodendrocytes, in the vertebrate central nervous system (CNS). In most CNS areas, neurons are the first to develop, followed by astrocytes, and at later stages by oligodendrocytes. An important issue in the study of early CNS development is to understand the lineage relationships of the various CNS cell types and the mechanisms by which these lineages segregate and differentiate. One aspect of this question is to define when and how precursor cells become committed to a specific differentiation pathway. For example, neuroepithelial cells could initially all be endowed with equivalent differentiation capabilities. The specification of these multipotential precursor cells towards a defined phenotype could be controlled by instructive and selective cues arising progressively from their immediate environment. Alternatively, neuroepithelial progenitors could be, from early stages in nervous system ontogeny, already segregated into subpopulations with differing potentialities. In this case, environmental cues would be less critical than intrinsic developmental programs in regulating phenotypic choices.

Neuron/Glia Lineages During Early Nervous System Development in Amphibian and Chicken Embryos

Neurons and macroglial cells of the vertebrate central nervous system (CNS) arise from neuroepithelial precursor cells that proliferate rapidly in the ventricular and subventricular zones and present similar morphological characteristics. In most CNS regions, neurons develop first, together with a subpopulation of immature glial cells, the radial glia. The other glial cell types, astrocytes and oligodendrocytes, develop later on according to spatio-temporal schemes specific for each central structure. This sequence of differentiation events raises several key questions regarding the lineage relationships of the various CNS cell types and the mechanisms by which these lineages segregate and differentiate. One important aspect of these questions is to define when and how precursor cells become irreversibly committed to a specific differentiation pathway. Neuroepithelial cells initially could be all endowed with equivalent differentiation capabilities. The specification of these multipotential precursor cells toward a defined phenotype could occur progressively, resulting, as development proceeds, in the formation of discrete families of determined progenitors. Alternatively, they could be restricted in their developmental fate only when they undergo their final round of division, just before differentiating. In either of these two alternatives, the environment should play an important role in specifying the ultimate cell phenotype. In another possible scheme, neuroepithelial progenitors could be already segregated into subpopuladons with differing potentialities at early stages in nervous system ontogeny. In this case, environmental cues would be less critical than intrinsic developmental programs in regulating phenotypic choices.