Pattern formation in the lateral line of zebrafish.

The lateral line of fish and amphibians is a sensory system that comprises a number of individual sense organs, the neuromasts, arranged in a defined pattern on the surface of the body. A conspicuous part of the system is a line of organs that extends along each flank (and which gave the system its name). At the end of zebrafish embryogenesis, this line comprises 7-8 neuromasts regularly spaced between the ear and the tip of the tail. The neuromasts are deposited by a migrating primordium that originates from the otic region. Here, we follow the development of this pattern and show that heterogeneities within the migrating primordium prefigure neuromast formation.

Early efferent innervation of the zebrafish lateral line.

We examined the efferent innervation of the lateral line in zebrafish larvae. Three efferent nuclei were previously reported for the posterior line, two in the hindbrain and one in the ventral hypothalamus. Here we show that the same three nuclei innervate the anterior line as well. The rhombencephalic neurons innervate either the anterior or the posterior line. The diencephalic neurons seem to innervate both lines as well as the ear. The diencephalic efferents are labeled by anti-tyrosine hydroxylase antibodies and probably use dopamine as a transmitter. They are among the very first catecholaminergic neurons to differentiate in the brain and extend branches into the lateral line system almost as soon as the latter forms. We discuss possible functions of the rhombencephalic and diencephalic efferents.

Neuronal differences prefigure somatotopy in the zebrafish lateral line.

The central projection of the fish lateral line displays somatotopic ordering. In order to know when and how this ordering is established, we have labelled single sensory neurones and followed the growth of their neurites. We show that the neuromast cells and the corresponding neurones are not related by a fixed lineage, and also that somatotopic differences between anterior and posterior line neurones, and among neurones of the posterior line, are present before innervation of the sense organs. We propose that the position of the central projection defines the peripheral position that the neurone will innervate.

A genetic programme for neuronal connectivity.

What is the nature of the genetic programme that allows neurons to extend their axons and connect to other neurons with a high degree of specificity? Work on the sensory neurons of the fly has shown how the control of neuronal identity is embedded in the general developmental programme of the organism. The ongoing analysis of pathfinding mutants suggests plausible mechanisms for the translation of neuronal identity into axonal behaviour.

Deconstructing cell determination: proneural genes and neuronal identity.

Vertebrates express scores of bHLH proteins during neural development. Earlier studies inspired by the established role of "proneural" genes in fly neurogenesis, as well as by the vertebrate bHLH myogenic program, focused on the reconstruction of bHLH gene cascades, which are thought to control successive steps leading to neuronal differentiation. Little attention has been paid thus far to the relationship between the diversity of neural bHLH genes and the diversity of neuronal phenotypes. This article reviews recent evidence that, akin to their fly counterparts, vertebrate neural bHLH genes probably confer not only "generic" neuronal properties, but also neuronal type-specific properties, inextricably linking neural determination and the specification of neuronal identity. We also speculate on the relations between positional information and gene activity, and on the evolutionary significance of the diversity of bHLH genes.

Somatotopy of the lateral line projection in larval zebrafish.

We examined the topography of the lateral line primary projection in zebrafish larvae by double labeling. The projections of two identified neuromasts of the posterior lateral line are seen as two separate sets of fibers that show reproducible spatial relationships: the projection of the anterior neuromast is always ventrolateral to that of a more posteriorly located neuromast. The same rule applies to the projection of anterior lateral line neuromasts. The position of the neuromasts along the antero posterior axis of the fish therefore is represented in the central projection of the sensory neurons. This somatotopy is similar to, and may be at the origin of, the tonotopic projection of the cochlear hair cells in mammals.

Isolation and characterization of single-chain Fv genes encoding antibodies specific for Drosophila Poxn protein.

The usefulness of intrabodies as specific inhibitors of gene function has been extensively demonstrated in cell culture assays. However, very few experiments have been conducted with intrabodies expressed in whole organisms. To evaluate the intrabody technology in Drosophila, we focused on poxn protein, since its effects can be easily studied. We purified the recombinant poxn protein. We next isolated three single-chain variable fragments (scFv) which specifically recognize poxn protein. Two scFvs, designated alpha-Poxn2 and alpha-Poxn4, react with both denatured and native Poxn with half maximal inhibition values of 100 nM and 40 nM, respectively. The alpha-Poxn5 scFv also recognizes denatured Poxn but either does not recognize native Poxn or its half maximal inhibition value for native Poxn is high.

The regulated expression of an intrabody produces a mutant phenotype in Drosophila.

Intrabodies show great promise for controlling gene expression. As an initial attempt to evaluate the intrabody technology in Drosophila, the gene poxn was used as target. Transgenic flies harboring different anti-Poxn scFv genes integrated into various chromosomes were obtained. In one transformant, a phenocopy resembling the hypomorphic poxn-phenotype was produced in embryos and larvae following induction of expression of alpha-Poxn2 intrabody. The antisense approach was used as control. Parameters that can affect the success of intrabody technology are described.

The iroquois complex controls the somatotopy of Drosophila notum mechanosensory projections.

Sensory neurons can establish topologically ordered projections in the central nervous system, thereby building an internal representation of the external world. We analyze how this ordering is genetically controlled in Drosophila, using as a model system the neurons that innervate the mechanosensory bristles on the back of the fly (the notum). Sensory neurons innervating the medially located bristles send an axonal branch that crosses the central nervous system midline, defining a 'medial' identity, while the ones that innervate the lateral bristles send no such branch, defining a 'lateral' identity. We analyze the role of the proneural genes achaete and scute, which are involved in the formation of the medial and lateral bristles, and we show that they have no effect on the 'medial' and 'lateral' identities of the neurons. We also analyze the role of the prepattern genes araucan and caupolican, two members of the iroquois gene complex which are required for the expression of achaete and scute in the lateral region of the notum, and we show that their expression is responsible for the 'lateral' identity of the projection.

Expression and function of tap in the gustatory and olfactory organs of Drosophila.

We have recently described the identification of a gene, tap, which encodes a bHLH protein expressed in one neuron of each larval chemosensory organ. Here we show that tap is expressed at a late stage in the development of one type of adult chemosensory organ, the gustatory bristles of the leg, wing and proboscis. We also show that tap is expressed very early in the development of a second type of chemosensory receptors, the olfactory organs of the antenna. The results of behavioral experiments suggest that the ectopic expression of tap affects the response to sugar and salt.

Genetic basis of the formation and identity of type I and type II neurons in Drosophila embryos.

The embryonic peripheral nervous system of Drosophila contains two main types of sensory neurons: type I neurons, which innervate external sense organs and chordotonal organs, and type II multidendritic neurons. Here, we analyse the origin of the difference between type I and type II in the case of the neurons that depend on the proneural genes of the achaete-scute complex (ASC). We show that, in Notch- embryos, the type I neurons are missing while type II neurons are produced in excess, indicating that the type I/type II choice relies on Notch-mediated cell communication. In contrast, both type I and type II neurons are absent in numb- embryos and after ubiquitous expression of tramtrack, indicating that the activity of numb and the absence of tramtrack are required to produce both external sense organ and multidendritic neural fates. The analysis of string- embryos reveals that when the precursors are unable to divide they differentiate mostly into type II neurons, indicating that the type II is the default neuronal fate. We also report a new mutant phenotype where the ASC-dependent neurons are converted into type II neurons, providing evidence for the existence of one or more genes required for maintaining the alternative (type I) fate. Our results suggest that the same mechanism of type I/type II specification may operate at a late step of the ASC-dependent lineages, when multidendritic neurons arise as siblings of the external sense organ neurons and, at an early step, when other multidendritic neurons precursors arise as siblings of external sense organ precursors.

tap, a Drosophila bHLH gene expressed in chemosensory organs.

We have isolated a Drosophila bHLH gene, tap, that is expressed in a small subset of neurons when they undergo differentiation. In the peripheral nervous system, tap is expressed exclusively in one of the neurons that innervate each larval chemosensory organ, possibly controlling the specific properties of that neuron. Sequence comparisons suggest that tap is most closely related to two bHLH genes identified in several vertebrate species, neurogenin and neuroD, which are involved respectively in neural determination and in neuronal differentiation.

Cell fate determination in Drosophila.

A major issue in development is to understand how local heterogeneities are interpreted to determine specific cell fates. The sense organs of Drosophila provide an accessible system for addressing this issue. Most sense organs comprise four types of cells, and their differentiation is the outcome of a complex developmental programme comprising several steps. Recent results illuminate, for several of these steps, the nature of the local heterogeneities and the mechanism used to interpret them in terms of cell fate decisions.

Lineage and fate in Drosophila: role of the gene tramtrack in sense organ development.

The tactile bristles of the fly comprise four cells that originate from a single precursor cell through a fixed lineage. The gene tramtrack (ttk) plays a crucial role in defining the fates of these cells. Here we analyse the normal pattern of expression of ttk, as well as the effect of ttk overexpression at different steps of the lineage. We show that ttk is never expressed in cells having a neural potential, and that in cells where ttk is expressed, there is a delay between division and the onset of expression. The ectopic expression of ttk before some stage of the cell cycle can block further cell division. Furthermore, this expression transforms neural into non-neural cells, suggesting that ttk acts as a repressor of neural fate at each step of the lineage. Our results suggest that ttk is probably not involved in setting up the mechanism that creates an asymmetry between sister cells, but rather in the implementation of that choice.

Genetic determinants of sense organ identity in Drosophila: regulatory interactions between cut and poxn.

Two genes involved in defining the type of sense organ have been identified in Drosophila. The gene cut differentiates the external sense organs (where it is expressed) from the chordotonal organs (where it is not); among the external sense organs poxn differentiates the poly-innervated organs (where it is expressed) from the mono-innervated organs (where it is not). Here we show that the expression of poxn in normal embryos does not depend on cut, and that poxn is capable of inducing the expression of cut. We have identified a small domain of the very large cut regulatory region as a likely target for activation by poxn.

[Development of the nervous system in Drosophila]. / Le développement du système nerveux chez la drosophile.

The formation of sense organs in Drosophila involves a series of choices, each of which depends on the co-ordinated activity of a small battery of genes. Two essential steps of this process have been extensively studied over the past few years: the determination of neural precursor cells, and their diversification. In both cases, the choices are dichotomous, and each choice reflects the fact that a specific control gene is or is not expressed. This principle is illustrated in the case of the genes "cut" and "poxn", the expression of which controls the type of sense organ that a given precursor will form.

Peripheral axonal pathway and cleaning behavior are correlated in Drosophila microchaetes.

The notum of Drosophila is covered with evenly spaced small mechanosensory bristles (microchaetes). Four groups of microchaetes can be distinguished on the basis of the pathways followed by the axons of their neurons. In three of the four groups, the axons extend along fibrous processes, the prospective tendons of the adult indirect flight muscles, formed by the epithelial cells. We show that the subdivision into four fasciculation groups is correlated to differences in the cleaning behavior elicited by stimulating the microchaetes.

Position-reading and the emergence of sense organ precursors in Drosophila.

Genetic analysis of development in Drosophila melanogaster has advanced our understanding of "position reading", where the expression of particular genes informs a cell of its position in the developing animal. The first step in localization of fly sense organs is the local expression of a gene conferring neural competence on epidermal cells. The four genes of the achaete-scute (AS-C) complex play crucial roles in the localization of sense organs. The resolution of local expression of AS-C genes along one dimension is about 10%; accuracy is improved by the balancing local expression of AS-C antagonist genes such as extramacrochaete. Position reading seems to depend primarily on such patterns of gene expression, and not upon the compartmental identity of the cells. No evidence has been found for differing roles of the four AS-C genes in the generation of sense organ progenitor cells or in the specification of neuronal properties of innervating neurons. The formation of each sense organ may be a unique case where the different proneural and neurogenic gene products have varying importance, and fortuitous local effects acting on this complex combination of factors have come to be important. The fly may be evolving from a flexible regular pattern to an inflexible irregular pattern strongly dependent on local factors, turning the fly into a crystallized system. (Written by R. Wayne Davies.).

The gene poxn controls different steps of the formation of chemosensory organs in Drosophila.

The gene poxn codes for a transcriptional regulator that specifies poly-innervated (chemosensory), as opposed to mono-innervated (mechanosensory), organs in Drosophila. The ectopic expression of poxn during metamorphosis results in a transformation of the morphology and central projection of adult mechanosensory organs toward those of chemosensory organs. Here we show, by electron microscopy analysis of normal and transformed bristles and by Dil labeling of the innervating neurons, that poxn also controls the number of neurons. To determine whether poxn can transform not only the sense organ precursor cells but also their daughter cells, we examine the effects of the ectopic expression of poxn at different stages of the lineage, and we conclude that poxn can act at a late stage to affect the fate of the undifferentiated neuron.

The leg of Drosophila as a model system for the analysis of neuronal diversity.

The neurons innervating insect sense organs vary in number, shape, dendritic morphology, axonal projections and connectivity, providing abundant material for the genetic analysis of neuronal diversity. Here we describe the leg of Drosophila as a potential model system for this analysis. The leg of Drosophila comprises a variety of sense organs arranged in a precise and reproducible pattern. The cell bodies of the sensory neurons are located near the organ they innervate, which greatly facilitates their identification and accessibility. The development of the leg from its progenitor structure, the imaginal disc, is known in good detail. In particular, the time of appearance and of divisions of the sense organ precursors is known. The origin and mode of formation of the leg nerve (through which all sensory axons project into the central nervous system) has been described. The central projections of some of the sensory neurons have been examined by horseradish peroxidase backfill or DiI labelling. Finally, the expression of several genes that control the differentiation of various types of sensory neurons can be manipulated at will. We illustrate these different aspects, and discuss the potentials and shortcomings of this system.