Context-dependent utilization of Notch activity in Drosophila glial determination

During Drosophila neurogenesis, glial differentiation depends on the expression of glial cells missing (gcm). Understanding how glial fate is achieved thus requires knowledge of the temporal and spatial control mechanisms directing gcm expression. A recent report showed that in the adult bristle lineage, gcm expression is negatively regulated by Notch signaling ( Van De Bor, V. and Giangrande, A. (2001). Development 128, 1381-1390). Here we show that the effect of Notch activation on gliogenesis is context-dependent. In the dorsal bipolar dendritic (dbd) sensory lineage in the embryonic peripheral nervous system (PNS), asymmetric cell division of the dbd precursor produces a neuron and a glial cell, where gcm expression is activated in the glial daughter. Within the dbd lineage, Notch is specifically activated in one of the daughter cells and is required for gcm expression and a glial fate. Thus Notch activity has opposite consequences on gcm expression in two PNS lineages. Ectopic Notch activation can direct gliogenesis in a subset of embryonic PNS lineages, suggesting that Notch-dependent gliogenesis is supported in certain developmental contexts. We present evidence that POU-domain protein Nubbin/PDM-1 is one of the factors that provide such context.     

Asymmetric cell division of thoracic neuroblast 6-4 to bifurcate glial and neuronal lineage in Drosophila

In the development of the Drosophila central nervous system, some of the neuroblasts designated as neuroglioblasts generate both glia and neurons. Little is known about how neuroglioblasts produce these different cell types. NB6-4 in the thoracic segment (NB6-4T) is a neuroglioblast, although the corresponding cell in the abdominal segment (NB6-4A) produces only glia. Here, we describe the cell divisions in the NB6-4T lineage, following changes in cell number and cell arrangement. We also examined successive changes in the expression of glial cells missing (gcm) mRNA and protein, activity of which is known to direct glial fate from the neuronal default state. The first cell division of NB6-4T occurred in the medial-lateral orientation, and was found to bifurcate the glial and neuronal lineage. After division, the medial daughter cell expressed GCM protein to produce three glial cells, while the lateral daughter cell with no GCM expression produced ganglion mother cells, secondary precursors of neurons. Although gcm mRNA was present evenly in the cytoplasm of NB6-4T before the first cell division, it became detected asymmetrically in the cell during mitosis and eventually only in the medial daughter cell. In contrast, NB6-4A showed a symmetrical distribution of gcm mRNA and GCM protein through division. Our observations suggest that mechanisms regulating gcm mRNA expression and its translation play an important role in glial and neuronal lineage bifurcation that results from asymmetric cell division.     

Notch signaling represses the glial fate in fly PNS

By using gain-of-function mutations it has been proposed that vertebrate Notch promotes the glial fate. We show in vivo that glial cells are produced at the expense of neurons in the peripheral nervous system of flies lacking Notch and that constitutively activated Notch produces the opposite phenotype. Notch acts as a genetic switch between neuronal and glial fates by negatively regulating glial cell deficient/glial cells missing, the gene required in the glial precursor to induce gliogenesis. Moreover, Notch represses neurogenesis or gliogenesis, depending on the sensory organ type. Numb, which is asymmetrically localized in the multipotent cell that produces the glial precursor, induces glial cells at the expense of neurons. Thus, a cell-autonomous mechanism inhibits Notch signaling.     

A novel mode of asymmetric division identifies the fly neuroglioblast 6-4T.

Asymmetric cell divisions and segregation of fate determinants are crucial events in the generation of cell diversity. Fly neuroblasts, the precursors that self-reproduce and generate neurons, represent a clear example of asymmetrically dividing cells. Less is known about how neurons and glial cells are generated by multipotent precursors. Flies provide the ideal model system to study this process. Indeed, neuroglioblasts (NGBs) can be specifically identified and have been shown to require the glide/gcm fate determinant to produce glial cells, which otherwise would become neurons. Here, we follow the division of a specific NGB (NGB6-4T), which produces a neuroblast (NB) and a glioblast (GB). We show that, to generate the glioblast, glide/gcm RNA becomes progressively unequally distributed during NGB division and preferentially segregates. Subsequently, a GB-specific factor is required to maintain glide/gcm expression. Both processes are necessary for gliogenesis, showing that the glial vs. neuronal fate choice is a two-step process. This feature, together with glide/gcm subcellular RNA distribution and the behavior of the NGB mitotic apparatus identify a novel type of division generating cell diversity.     

glide/gcmIs Expressed and Required in the Scavenger Cell Lineage

Glial cell differentiation inDrosophila melanogasterrequires the activity ofglide/gcm(glial cell deficient/glial cell missing). The role of this gene is to direct the cell fate switch between neurons and glial cells by activating the glial developmental program in multipotent precursor cells of the nervous system. In this paper, we show thatglide/gcmis also expressed and required in the lineage of hemocytes/macrophages, scavenger cells that phagocytose cells undergoing programmed cell death. In addition, we show that, as for glial cells,glide/gcmplays an instructive role in hemocyte differentiation. Interestingly, it has been shown that in the development of the fly adult nervous system the role of scavenger cells is played by glial cells. These data and our findings on the dual role ofglide/gcmindicate that glial cells and hemocytes/macrophages are functionally and molecularly related.     

A critical role for cyclin E in cell fate determination in the central nervous system of Drosophila melanogaster

We have examined the process by which cell diversity is generated in neuroblast (NB) lineages in the central nervous system of Drosophila melanogaster. Thoracic NB6-4 (NB6-4t) generates both neurons and glial cells, whereas NB6-4a generates only glial cells in abdominal segments. This is attributed to an asymmetric first division of NB6-4t, localizing prospero (pros) and glial cell missing (gcm) only to the glial precursor cell, and a symmetric division of NB6-4a, where both daughter cells express pros and gcm. Here we show that the NB6-4t lineage represents the ground state, which does not require the input of any homeotic gene, whereas the NB6-4a lineage is specified by the homeotic genes abd-A and Abd-B. They specify the NB6-4a lineage by down-regulating levels of the G1 cyclin, DmCycE (CycE). CycE, which is asymmetrically expressed after the first division of NB6-4t, functions upstream of pros and gcm to specify the neuronal sublineage. Loss of CycE function causes homeotic transformation of NB6-4t to NB6-4a, whereas ectopic CycE induces reverse transformations. However, other components of the cell cycle seem to have a minor role in this process, suggesting a critical role for CycE in regulating cell fate in segment-specific neural lineages.     

Wasp, the Drosophila Wiskott-Aldrich syndrome gene homologue, is required for cell fate decisions mediated by Notch signaling

Wiskott-Aldrich syndrome proteins, encoded by the Wiskott-Aldrich syndrome gene family, bridge signal transduction pathways and the microfilament-based cytoskeleton. Mutations in the Drosophila homologue, Wasp (Wsp), reveal an essential requirement for this gene in implementation of cell fate decisions during adult and embryonic sensory organ development. Phenotypic analysis of Wsp mutant animals demonstrates a bias towards neuronal differentiation, at the expense of other cell types, resulting from improper execution of the program of asymmetric cell divisions which underlie sensory organ development. Generation of two similar daughter cells after division of the sensory organ precursor cell constitutes a prominent defect in the Wsp sensory organ lineage. The asymmetric segregation of key elements such as Numb is unaffected during this division, despite the misassignment of cell fates. The requirement for Wsp extends to additional cell fate decisions in lineages of the embryonic central nervous system and mesoderm. The nature of the Wsp mutant phenotypes, coupled with genetic interaction studies, identifies an essential role for Wsp in lineage decisions mediated by the Notch signaling pathway.     

Asymmetric localization and function of cell-fate determinants: a fly's view

One mechanism to generate daughter cells with distinct fates is the asymmetric inheritance of regulatory proteins, leading to differential gene regulation in the daughter cells. This mode of cell division is termed 'asymmetric cell division.' The nervous system of the fly employs asymmetric cell division, both in the central nervous system, to generate neural precursors, neurons and glial cells; and in the peripheral nervous system, to create sensory organs that are composed of multiple cell types. These cell lineages are excellent models to examine the gene expression program that leads to fate acquisition, the cell-fate determinants that control these programs and how these determinants, in turn, are distributed through cell polarity machinery.     

Prospero distinguishes sibling cell fate without asymmetric localization in the Drosophila adult external sense organ lineage

The adult external sense organ precursor (SOP) lineage is a model system for studying asymmetric cell division. Adult SOPs divide asymmetrically to produce IIa and IIb daughter cells; IIa generates the external socket (tormogen) and hair (trichogen) cells, while IIb generates the internal neuron and sheath (thecogen) cells. Here we investigate the expression and function of prospero in the adult SOP lineage. Although Prospero is asymmetrically localized in embryonic SOP lineage, this is not observed in the adult SOP lineage: Prospero is first detected in the IIb nucleus and, during IIb division, it is cytoplasmic and inherited by both neuron and sheath cells. Subsequently, Prospero is downregulated in the neuron but maintained in the sheath cell. Loss of prospero function leads to 'double bristle' sense organs (reflecting a IIb-to-IIa transformation) or 'single bristle' sense organs with abnormal neuronal differentiation (reflecting defective IIb development). Conversely, ectopic prospero expression results in duplicate neurons and sheath cells and a complete absence of hair/socket cells (reflecting a IIa-to-IIb transformation). We conclude that (1) despite the absence of asymmetric protein localization, prospero expression is restricted to the IIb cell but not its IIa sibling, (2) prospero promotes IIb cell fate and inhibits IIa cell fate, and (3) prospero is required for proper axon and dendrite morphology of the neuron derived from the IIb cell. Thus, prospero plays a fundamental role in establishing binary IIa/IIb sibling cell fates without being asymmetrically localized during SOP division. Finally, in contrast to previous studies, we find that the IIb cell divides prior to the IIa cell in the SOP lineage.     

Terminal tendon cell differentiation requires the glide/gcm complex

Locomotion relies on stable attachment of muscle fibres to their target sites, a process that allows for muscle contraction to generate movement. Here, we show that glide/gcm and glide2/gcm2, the fly glial cell determinants, are expressed in a subpopulation of embryonic tendon cells and required for their terminal differentiation. By using loss-of-function approaches, we show that in the absence of both genes, muscle attachment to tendon cells is altered, even though the molecular cascade induced by stripe, the tendon cell determinant, is normal. Moreover, we show that glide/gcm activates a new tendon cell gene independently of stripe. Finally, we show that segment polarity genes control the epidermal expression of glide/gcm and determine, within the segment, whether it induces glial or tendon cell-specific markers. Thus, under the control of positional cues, glide/gcm triggers a new molecular pathway involved in terminal tendon cell differentiation, which allows the establishment of functional muscle attachment sites and locomotion.     

Mechanism of glia-neuron cell-fate switch in the Drosophila thoracic neuroblast 6-4 lineage

During development of the Drosophila central nervous system, neuroblast 6-4 in the thoracic segment (NB6-4T) divides asymmetrically into a medially located glial precursor cell and a laterally located neuronal precursor cell. In this study, to understand the molecular basis for this glia-neuron cell-fate decision, we examined the effects of some known mutations on the NB6-4T lineage. First, we found that prospero (pros) mutations led to a loss of expression of Glial cells missing, which is essential to trigger glial differentiation, in the NB6-4T lineage. In wild-type embryos, Pros protein was localized at the medial cell cortex of dividing NB6-4T and segregated to the nucleus of the glial precursor cell. miranda and inscuteable mutations altered the behavior of Pros, resulting in failure to correctly switch the glial and neuronal fates. Our results suggested that NB6-4T used the same molecular machinery in the asymmetric cell division as other neuroblasts in cell divisions producing ganglion mother cells. Furthermore, we showed that outside the NB6-4T lineage most glial cells appeared independently of Pros.