Alterations of the eyes during ontogenesis inAporrhais pespelecani (Mollusca,Caenogastropoda)

The cerebrally innervated eyes of metamorphically competent larvae, newly metamorphosed larvae, and adults ofAporrhais pespelecani are ultrastructurally investigated and compared. The eyes are composed of a lens, a cornea, and an everse retina. In adults, a humour is located behind the lens. The retina consists of two different types of cells: sensory cells and supportive cells. The present study confirms earlier results and demonstrates that the distal part of the sensory cells is altered during ontogenesis. In metamorphically competent larvae, the sensory cells are exclusively ciliary. In newly metamorphosed larvae and in adults, however, the sensory cells are of the mixed type, bearing both cilia and microvilli. Furthermore, the findings confirm that both the supportive and corneal cells, as well as the distal supportive cell processes which are restricted to the eyes of adults are involved in lens formation.     

A late role for a subset of neurogenic genes to limit sensory precursor recruitments in Drosophila embryos

Summary In Drosophila, mutations in a class of genes, the neurogenic genes, produce an excess of neurons. This neural hyperplasia has been attributed to the formation of more than the normal number of neuronal precursor cells at the expense of epidermal cells. In order to find out whether the neurogenic genes only act at this intial step of neurogenesis, we studied the replication pattern of the sensory organ precursor cells by monitoring BrdU incorporation in embryos mutant for Notch (N), Delta (Dl), mastermind (mam), almondex (amx), neuralized (neu), big brain (bib) and the Enhancer of split-Complex (E(spl)-C). Using temperature sensitive alleles of two of the neurogenic genes, DI and N, we also induced an acute increase of replicating sensory precursors by shifting briefly to the restricted temperature. We have found that the loss of function of all the seven neurogenic loci that were tested causes an increase in replicating sensory precursor cells, consistent with the model that these neurogenic genes normally participate in the process of restricting the number of neuronal precursors. Whereas the temporal pattern of replication appeared normal in mutants of five of the seven neurogenic loci, in N and mam embryos replicating PNS cells are present beyond the time when they normally undergo replication. Experiments with colchicine suggest that many of these late replicating cells may be newly emerging precursors and probably not additional cell divisions of already recruited precursors. Thus, different neurogenic genes may be required over different periods of time for the specification of sensory precursor cells. Correspondence to: R. Bodmer     

The big brain gene of Drosophila functions to control the number of neuronal precursors in the peripheral nervous system.

big brain (bib) is one of the six known zygotic neurogenic genes involved in the decision of an ectodermal cell to take on the neurogenic or the epidermogenic cell fate. Previous studies suggest that bib functions in a pathway separate from the one involving Notch and other known neurogenic genes. For a better understanding of the bib function, it is essential first to characterize the mutant phenotype in detail. Our mutant analyses show that loss of bib function approximately doubles the number of neuronal precursors and their progeny cells in the embryonic peripheral nervous system. Mosaic studies reveal a hypertrophy of sensory bristles in bib mutant patches in adult flies. Our observations are compatible with a function of bib in specifying neuronal precursors of both the embryonic and adult sensory nervous system. This is in contrast to the function of Notch, which continues to be required at multiple stages of neural development subsequent to this initial determination event.     

A gain-of-function screen for genes that affect the development of the Drosophila adult external sensory organ

The Drosophila adult external sensory organ, comprising a neuron and its support cells, is derived from a single precursor cell via several asymmetric cell divisions. To identify molecules involved in sensory organ development, we conducted a tissue-specific gain-of-function screen. We screened 2293 independent P-element lines established by P. Rorth and identified 105 lines, carrying insertions at 78 distinct loci, that produced misexpression phenotypes with changes in number, fate, or morphology of cells of the adult external sensory organ. On the basis of the gain-of-function phenotypes of both internal and external support cells, we subdivided the candidate lines into three classes. The first class (52 lines, 40 loci) exhibits partial or complete loss of adult external sensory organs. The second class (38 lines, 28 loci) is associated with increased numbers of entire adult external sensory organs or subsets of sensory organ cells. The third class (15 lines, 10 loci) results in potential cell fate transformations. Genetic and molecular characterization of these candidate lines reveals that some loci identified in this screen correspond to genes known to function in the formation of the peripheral nervous system, such as big brain, extra macrochaetae, and numb. Also emerging from the screen are a large group of previously uncharacterized genes and several known genes that have not yet been implicated in the development of the peripheral nervous system.     

The emergence of sense organs in the wing disc of Drosophila

We have examined the origin of a set of precisely located sense organs in the notum and wing of Drosophila, in transformant flies where lacZ is expressed in the progenitor cells of the sense organs (the sensory mother cells) and in their progeny. Here we describe the temporal pattern of appearance and divisions of the sensory mother cells that will form the eleven macrochaetes and the two trichoid sensilla of the notum, and five campaniform sensilla on the wing blade. The complete pattern of sensory mother cells develops in a strict sequence that extends over most of the third larval instar and the first 10 h after puparium formation. The delay between the onset of lacZ expression and the first differentiative division ranges from 30 h, in the case of the earliest mother cells, to 2 h for the latest mother cells. The first division shows a preferential orientation which is also specific for each sensory mother cell. Up to this stage, there is no marked difference between the three types of mechanosensory organs.     

Revisiting cell fate specification in the inner ear

Generating the diversity of cell types in the inner ear may require an interplay between regional compartmentalization and local cellular interactions. Recent evidence has come from gene targeting, lineage analysis, fate mapping and gene expression studies. Notch signaling and neurogenic gene regulation are involved in patterning or specification of sensory organs, ganglion cells and hair cell mechanoreceptors.     

The function of the neurogenic genes during epithelial development in the Drosophila embryo.

The complex embryonic phenotype of the six neurogenic mutations Notch, mastermind, big brain, Delta, Enhancer of split and neuralized was analyzed by using different antibodies and PlacZ markers, which allowed us to label most of the known embryonic tissues. Our results demonstrate that all of the neurogenic mutants show abnormalities in many different organs derived from all three germ layers. Defects caused by the neurogenic mutations in ectodermally derived tissues fell into two categories. First, all cell types that delaminate from the ectoderm (neuroblasts, sensory neurons, peripheral glia cells and oenocytes) are increased in number. Secondly, ectodermal tissues that in the wild type form epithelial structures lose their epithelial phenotype and dissociate (optic lobe, stomatogastric nervous system) or show significant differentiative abnormalities (trachea, Malpighian tubules and salivary gland). Abnormalities in tissues derived from the mesoderm were observed in all six neurogenic mutations. Most importantly, somatic myoblasts do not fuse and/or form an aberrant muscle pattern. Cardioblasts (which form the embryonic heart) are increased in number and show differentiative abnormalities; other mesodermal cell types (fat body, pericardial cells) are significantly decreased. The development of the endoderm (midgut rudiments) is disrupted in most of the neurogenic mutations (Notch, Delta, Enhancer of split and neuralized) during at least two stages. Defects occur as early as during gastrulation when the invaginating midgut rudiments prematurely lose their epithelial characteristics. Later, the transition of the midgut rudiments to form the midgut epithelium does not occur. In addition, the number of adult midgut precursor cells that segregate from the midgut rudiments is strongly increased. We propose that, at least in the ectodermally and endodermally derived tissues, neurogenic gene function is primarily involved in interactions among cells that need to acquire or to maintain an epithelial phenotype.     

Suppressor of Hairless, the Drosophila homolog of the mouse recombination signal-binding protein gene, controls sensory organ cell fates.

Suppressor of Hairless (Su(H)) is required at two stages of adult sensory organ development in Drosophila. Complete loss of Su(H) function results in a "neurogenic" phenotype in imaginal discs, in which too many cells adopt the sensory organ precursor cell fate. Su(H) is also involved in controlling the fates of sensillum accessory cells and is specifically expressed in two of these cells. Su(H) is the Drosophila homolog of the mouse J kappa RBP gene, whose product binds specifically to the recombination signal sequence of immunoglobulin J kappa segments. The Su(H) and J kappa RBP proteins are 82% identical over most of their length, and share with bacteriophage integrates and yeast recombinases a motif that includes residues directly involved in catalyzing recombination.     

Estimation of neuroblast numbers in insect neurogenesis using the lateral inhibition hypothesis of cell differentiation

Cells in the neurogenic region of an insect ectoderm have two alternative fates, making neurons or epidermis. The fates seem to be determined through a laterally inhibitory interaction among cells. That is, initially homogeneous cells are all competent to differentiate into neuroblasts. Once a cell has differentiated as a neuroblast, it inhibits its immediate neighbors from following this pathway. The differentiation process is simulated by a digital computer in a planar array of polygonal domains similar to a cell pattern. We find that the number of cells differentiating as neuronal precursors in insect neurogenesis is that expected under the hypothesis of lateral inhibition of cell differentiation between immediate neighbors.     

Formation of competent Bacillus subtilis cells.

The process of competent cell formation for transformation has been studied with early-stationary-phase (T1) cells of Bacillus subtilis which had been grown in an enriched Spizizen minimal medium and transferred to a second synthetic medium. Rifampin, chloramphenicol, and tunicamycin were strong inhibitors of competent cell formation, as well as vegetative growth. After formation, competent cells were no longer sensitive to the above agents. Methicillin and an inhibitor of chromosomal replication, hydroxyphenylazouracil, did not inhibit the development of competence. A D-alanine-requiring mutant strain developed competence even in the absence of D-alanine in the second medium. A T1-stage culture showed the activity of extracellular serine protease which is necessary for sporulation. Competent cell formation was completely blocked by 0.7 M ethanol, which is a specific inhibitor of early events during sporulation, including forespore septum formation. Competent cells were formed even in media which supported sporulation. The development of competence was also studied with spo0 mutants at 10 different loci. Most spo0 mutations repressed the development of competence except for spo0C, spo0G, and spo0J. These results suggest that competent cells are formed from early sporulating cells with the synthesis of cell wall materials and by factors whose genes are activated by the supply of nutrients. It is suggested that common steps are involved both in forespore septation and in competent cell formation.     

G2 arrest of cell cycle ensures a determination process of sensory mother cell formation in Drosophila

 In Drosophila, the sensory mother cells of macrochaetes are chosen from among the mitotically quiescent clusters of cells in wing imaginal discs, where other cells are proliferating. The pattern of cyclin A, one of the G2 cyclins, reveals that mitotically quiescent clusters of cells are arrested in G2. When precocious mitoses are induced during sensory mother cell determination by the ectopic expression of string, a known G2/M transition regulator, the formation of sensory mother cells is disturbed, resulting in the loss of macrochaetes in the adult notum. This suggests that G2 arrest of the cell cycle ensures the proper determination of sensory mother cells. Received: 16 December 1996 / Accepted: 14 March 1997     

Expression of proneural and neurogenic genes in the zebrafish lateral line primordium correlates with selection of hair cell fate in neuromasts

Expression of a mouse atonal homologue, math1, defines cells with the potential to become sensory hair cells in the mouse inner ear (Science 284 (1999) 1837) and Notch signaling limits the number of cells that are permitted to adopt this fate (Nat. Genet. 21 (1999) 289; J. Neurocytol. 28 (1999) 809). Failure of lateral inhibition mediated by Notch signaling is associated with an overproduction of ear hair cells in the zebrafish mind bomb (mib) and deltaA mutants (Development 125 (1998a) 4637; Development 126 (1999) 5669), suggesting a similar role for these genes in limiting the number of hair cells in the zebrafish ear. This study extends the analysis of proneural and neurogenic gene expression to the lateral line system, which detects movement via clusters of related sensory hair cells in specialized structures called neuromasts. We have compared the expression of a zebrafish atonal homologue, zath1, and neurogenic genes, deltaA, deltaB and notch3, in neuromasts and the posterior lateral line primordium (PLLP) of wild-type and mib mutant embryos. We describe progressive restriction of proneural and neurogenic gene expression in the migrating PLLP that appears to correlate with selection of hair cell fate in maturing neuromasts. In mib mutants there is a failure to restrict expression of zath1 and Delta homologues in the neuromasts revealing similarities with the phenotype previously described in the ear.     

Cross-regulation of Ngn1 and Math1 coordinates the production of neurons and sensory hair cells during inner ear development

Temporal and spatial coordination of multiple cell fate decisions is essential for proper organogenesis. Here, we define gene interactions that transform the neurogenic epithelium of the developing inner ear into specialized mechanosensory receptors. By Cre-loxP fate mapping, we show that vestibular sensory hair cells derive from a previously neurogenic region of the inner ear. The related bHLH genes Ngn1 (Neurog1) and Math1 (Atoh1) are required, respectively, for neural and sensory epithelial development in this system. Our analysis of mouse mutants indicates that a mutual antagonism between Ngn1 and Math1 regulates the transition from neurogenesis to sensory cell production during ear development. Furthermore, we provide evidence that the transition to sensory cell production involves distinct autoregulatory behaviors of Ngn1 (negative) and Math1 (positive). We propose that Ngn1, as well as promoting neurogenesis, maintains an uncommitted progenitor cell population through Notch-mediated lateral inhibition, and Math1 irreversibly commits these progenitors to a hair-cell fate.     

Mutations of early neurogenesis inDrosophila

Summary Embryonic lethal mutations at the Notch locus are known to produce a conspicuous central nervous system hypertrophy accompanied by a hypotrophy of the epidermal sheath. We have studied several zygotic mutants belonging to four different autosomal complementation groups which produce the same phenotype. The embryonic development of the new mutants, as well as that of Notch, consists of an initial enlargement of the neurogenic region at the expenses of epidermal cell precursors. The possibility is discussed that these five loci are involved in the determination of neural and epidermal cell precursors.     

Podial sensory receptors and the induction of metamorphosis in echinoids

Larvae of Strongylocentrotus droebachiensis (Müller), Lytechinus pictus (Verrill), and Lytechinus variegatus (Leske) which are competent to metamorphose display what appears to be substratum-testing behavior prior to metamorphosis. Larvae cease swimming, partially evert the adult rudiment, and walk about examining the substratum with their five primary podia. Larvae eithe r metamorphose or withdraw their podia and resume swimming to settle again elsewhere. Scanning and transmission electron microscopic examinations of the primary podia revealed sensory receptor cells on the rim and on a conical projection at the center of the podial sucker. Each sensory cell has a single short cilium on its apical surface a axonal process at its base which contributes to the basiepithelial nerve plexus. Mature adults of the same species also have comparable sensory structures on their tube feet suckers. It is suggested that the sensory receptors on the primary podia of setting larvae, although they are not specialized larval structures, may be involved in the perception of tactil e stimuli which have been previously demonstrated to be involved in the induction of metamorphosis.     

Role of neurogenic genes in establishment of follicle cell fate and oocyte polarity during oogenesis in Drosophila

Oogenesis in Drosophila involves specification of both germ cells and the surrounding somatic follicle cells, as well as the determination of oocyte polarity. We found that two neurogenic genes, Notch and Delta, are required in oogenesis. These genes encode membrane proteins with epidermal growth factor repeats and are essential in the decision of an embryonic ectodermal cell to take on the fate of neuroblast or epidermoblast. In oogenesis, mutation in either gene leads to an excess of posterior follicle cells, a cell fate change reminiscent of the hyperplasia of neuroblasts seen in neurogenic mutant embryos. Furthermore, the Notch mutation in somatic cells causes mislocalization of bicoid in the oocyte. These results suggest that the neurogenic genes Notch and Delta are involved in both follicle cell development and the establishment of anterior-posterior polarity in the oocyte.     

Identification and isolation of multipotential neural progenitor cells from the subcortical white matter of the adult human brain

The subcortical white matter of the adult human brain harbors a pool of glial progenitor cells. These cells can be isolated by fluorescence-activated cell sorting (FACS) after either transfection with green fluorescent protein (GFP) under the control of the CNP2 promoter, or A2B5-targeted immunotagging. Although these cells give rise largely to oligodendrocytes, in low-density culture we observed that some also generated neurons. We thus asked whether these nominally glial progenitors might include multipotential progenitor cells capable of neurogenesis. We found that adult human white-matter progenitor cells (WMPCs) could be passaged as neurospheres in vitro and that these cells generated functionally competent neurons and glia both in vitro and after xenograft to the fetal rat brain. WMPCs were able to produce neurons after their initial isolation and did not require in vitro expansion or reprogramming to do so. These experiments indicate that an abundant pool of mitotically competent neurogenic progenitor cells resides in the adult human white matter.     

Molecular interactions between the protein products of the neurogenic loci Notch and Delta, two EGF-homologous genes in Drosophila

Genetic analyses have raised the possibility of interactions between the gene products of the neurogenic loci Notch and Delta, each of which encodes a transmembrane protein with EGF homology. To examine the possibility of intermolecular association between the products of these two genes, we studied the effects of their expression on aggregation in Drosophila S2 cells. We find that Notch-expressing cells form mixed aggregates specifically with cells that express Delta and that this process is calcium dependent. In addition, we show that Notch and Delta can associate within the membrane of a single cell, and further, that they form detergent-soluble intermolecular complexes. Our analyses suggest that Notch and Delta proteins interact at the cell surface via their extracellular domains.