Differential activities of the core planar polarity proteins during Drosophila wing patterning

During planar polarity patterning of the Drosophila wing, a "core" group of planar polarity genes has been identified which acts downstream of global polarity cues to locally coordinate cell polarity and specify trichome production at distal cell edges. These genes encode protein products that assemble into asymmetric apicolateral complexes that straddle the proximodistal junctional region between adjacent cells. We have carried out detailed genetic analysis experiments, analysing the requirements of each complex component for planar polarity patterning. We find that the three transmembrane proteins at the core of the complex, Frizzled, Strabismus and Flamingo, are required earliest in development and are the only components needed for intercellular polarity signalling. Notably, cells that lack both Frizzled and Strabismus are unable to signal, revealing an absolute requirement for both proteins in cell-cell communication. In contrast the cytoplasmic components Dishevelled, Prickle and Diego are not needed for intercellular communication. These factors contribute to the cell-cell propagation of polarity, most likely by promotion of intracellular asymmetry. Interestingly, both local polarity propagation and trichome placement occur normally in mutant backgrounds where asymmetry of polarity protein distribution is undetectable, suggesting such asymmetry is not an absolute requirement for any of the functions of the core complex.     

Temporal regulation of apterous activity during development of the Drosophila wing

Dorsoventral axis formation in the Drosophila wing depends on the activity of the selector gene apterous. Although selector genes are usually thought of as binary developmental switches, we find that Apterous activity is negatively regulated during wing development by its target gene dLMO. Apterous-dependent expression of Serrate and fringe in dorsal cells leads to the restricted activation of Notch along the dorsoventral compartment boundary. We present evidence that the ability of cells to participate in this Apterous-dependent cell-interaction is under spatial and temporal control. Apterous-dependent expression of dLMO causes downregulation of Serrate and fringe and allows expression of delta in dorsal cells. This limits the time window during which dorsoventral cell interactions can lead to localized activation of Notch and induction of the dorsoventral organizer. Overactivation of Apterous in the absence of dLMO leads to overexpression of Serrate, reduced expression of delta and concomitant defects in differentiation and cell survival in the wing primordium. Thus, downregulation of Apterous activity is needed to allow normal wing development.     

The role of Suppressor of Hairless in Notch mediated signalling during zebrafish somitogenesis

Suppressor of Hairless (Su(H)) codes for a protein that interacts with the intracellular domain of Notch to activate the target genes of the Delta-Notch signalling pathway. We have cloned the zebrafish homologue of Su(H) and have analysed its function by morpholino mediated knockdown. While there are at least four notch and four delta homologues in zebrafish, there appears to be only one complete Su(H) homologue. We have analysed the function of Su(H) in the somitogenesis process and its influence on the expression of notch pathway genes, in particular her1, her7, deltaC and deltaD. The cyclic expression of her1, her7 and deltaC in the presomitic mesoderm is disrupted by the Su(H) knockdown mimicking the expression of these genes in the notch1a mutant deadly seven. deltaD expression is similarly affected by Su(H) knockdown like deltaC but shows in addition an ectopic expression in the developing neural tube. The inactivation of Su(H) in a fss/tbx24 mutant background leads furthermore to a clear breakdown of cyclic her1 and her7 expression, indicating that the Delta-Notch pathway is required for the creation of oscillation and not only for the synchronisation between neighbouring cells. The strongest phenotypes in the Su(H) knockdown embryos show a loss of all somites posterior to the first five to seven ones. This phenotype is stronger than the known amorphic phenotypes for notch1 (des) or deltaD (aei) in zebrafish, but mimicks the knockout phenotype of RBP-Jkappa gene in the mouse, which is the homologue of Su(H). This suggests that there is some functional redundancy among the Notch and Delta genes. This fact that the first five to seven somites are only weakly affected by Su(H) knockdown indicates that additional genetic pathways may be active in the specification of the most anterior somites.     

Wingless promotes cell survival but constrains growth during Drosophila wing development

During animal development, organs grow to a fixed size and shape. Organ development typically begins with a rapid growth phase followed by a gradual decline in growth rate as the organ matures, but the regulation of either stage of growth remains unclear. The Wnt/Wingless (Wg) proteins are critical for patterning most animal organs, have diverse effects on development and have been proposed to promote organ growth. Here we report that contrary to this view, Wg activity actually constrains wing growth during Drosophila melanogaster wing development. In addition, we demonstrate that Wg is required for wing cell survival, particularly during the rapid growth phase of wing development. We propose that the cell-survival- and growth-constraining activities of Wg function to sculpt and delimit final wing size as part of its overall patterning programme.     

Multiple roles of the Dcdc42 GTPase during wing development in Drosophila melanogaster

The Rho sub-family of GTPases, comprising Rho, Rac and Cdc42. regulates many biological processes, including morphogenesis, cell polarity, migration, the cell cycle and gene expression. It is important to develop genetic approaches to allow the dissection, in vivo, of the mechanisms of GTPase regulation and signal transmission, and their biological consequences. In this regard, wing development in Drosophila melanogaster is an excellent model system. To investigate the functions of the Drosophila Cdc42 GTPase (Dcdc42), we generated phenotypes during wing development, by expression of the dominant-negative N17 and L89 mutants of Dcdc42. We have identified roles for Dcdc42 in wing growth, and in cell fate choice during the development of the wing veins and the peripheral nervous system. Reduction of Dcdc42 signalling following over-expression of Dcdc42N17 resulted in a broader but more diffuse domain characterised by wing-margin sensory bristles. This was correlated with a broadened stripe of wingless expression along the dorsal-ventral boundary of third-instar wing imaginal discs. Together with genetic interactions with loss- and gain-of-function Notch alleles, these data support a role for wild-type Dcdc42 as a negative regulator of Notch signalling.     

Drawing lines in the Drosophila wing: initiation of wing vein development

It has been proposed that wing veins in Drosophila form at boundaries between discrete sectors of cells that subdivide the anterior-posterior axis of the developing wing primordium. Recently, analysis of events underlying initiation of vein formation suggests that there is a general developmental mechanism for drawing lines between adjacent domains of cells, which is referred to as 'for-export-only-signaling'. In this model, cells in one domain produce a short range signal to which they cannot respond. As a consequence of this constraint, cells lying in a narrow line immediately outside the signal-producing domain are the only cells that can respond to the signal by activating expression of vein-promoting genes.     

Nemo is an inducible antagonist of Wingless signaling during Drosophila wing development

The cellular events that govern patterning during animal development must be precisely regulated. This is achieved by extrinsic factors and through the action of both positive and negative feedback loops. Wnt/Wg signals are crucial across species in many developmental patterning events. We report that Drosophila nemo (nmo) acts as an intracellular feedback inhibitor of Wingless (Wg) and that it is a novel Wg target gene. Nemo antagonizes the activity of the Wg signal, as evidenced by the finding that reduction of nmo rescues the phenotypic defects induced by misexpression of various Wg pathway components. In addition, the activation of Wg-dependent gene expression is suppressed in wing discs ectopically expressing nmo and enhanced cell autonomously in nmo mutant clones. We find that nmo itself is a target of Wg signaling in the imaginal wing disc. nmo expression is induced upon high levels of Wg signaling and can be inhibited by interfering with Wg signaling. Finally, we observe alterations in Arm stabilization upon modulation of Nemo. These observations suggest that the patterning mechanism governed by Wg involves a negative feedback circuit in which Wg induces expression of its own antagonist Nemo.     

Expression of the blistered/DSRF gene is controlled by different morphogens during Drosophila trachea and wing development

The Drosophila serum response factor (DSRF) is expressed in the precursors of the terminal tracheal cells and in the future intervein territories of the third instar wing imaginal disc. Dissection of the DSRF regulatory region reveals that a single enhancer element, which is under the control of the fibroblast growth factor (FGF)-receptor signalling pathway, is sufficient to induce DSRF expression in the terminal tracheal cells. In contrast, two separate enhancers direct expression in distinct intervein sectors of the wing imaginal disc. One element is active in the central intervein sector and is induced by the Hedgehog signalling pathway. The other element is under the control of Decapentaplegic and is active in two separate territories, which roughly correspond to the intervein sectors flanking the central sector. Hence, each of the three characterized enhancers constitutes a molecular link between a specific territory induced by a morphogen signal and the localized expression of a gene required for the final differentiation of this territory.     

Membrane trafficking in Drosophila wing and eye development

It is clear that membrane transport is essential to the proper sorting and delivery of membrane bound receptors and ligands, and secreted signaling molecules. Molecular genetic studies in Drosophila are particularly well suited to studies of membrane transport in development. The conservation of cell signaling pathways and membrane transport molecules between Drosophila and other species makes the results obtained in these studies of general interest. In addition, the ability to generate gain- and loss-of-function genetic mutations of various strengths, and the ability to generate transgenic flies that direct protein expression to tissues during development are of particular advantage. Several recent papers suggest that interesting and novel roles for membrane transport processes will be uncovered by studying classically defined membrane transport proteins in developmental contexts. Together these studies suggest that regulation of membrane transport may represent an additional mechanism to regulate the strength of cell-cell signaling during development.     

Drosophila wing development in the absence of dorsal identity

The developing wing disc of Drosophila is divided into distinct lineage-restricted compartments along both the anterior/posterior (A/P) and dorsal/ventral (D/V) axes. At compartment boundaries, morphogenic signals pattern the disc epithelium and direct appropriate outgrowth and differentiation of adult wing structures. The mechanisms by which affinity boundaries are established and maintained, however, are not completely understood. Compartment-specific adhesive differences and inter-compartment signaling have both been implicated in this process. The selector gene apterous (ap) is expressed in dorsal cells of the wing disc and is essential for D/V compartmentalization, wing margin formation, wing outgrowth and dorsal-specific wing structures. To better understand the mechanisms of Ap function and compartment formation, we have rescued aspects of the ap mutant phenotype with genes known to be downstream of Ap. We show that Fringe (Fng), a secreted protein involved in modulation of Notch signaling, is sufficient to rescue D/V compartmentalization, margin formation and wing outgrowth when appropriately expressed in an ap mutant background. When Fng and alphaPS1, a dorsally expressed integrin subunit, are co-expressed, a nearly normal-looking wing is generated. However, these wings are entirely of ventral identity. Our results demonstrate that a number of wing development features, including D/V compartmentalization and wing vein formation, can occur independently of dorsal identity and that inter-compartmental signaling, refined by Fng, plays the crucial role in maintaining the D/V affinity boundary. In addition, it is clear that key functions of the ap selector gene are mediated by only a small number of downstream effectors.     

Regulation of Apterous activity in Drosophila wing development.

Apterous is a LIM-homeodomain protein that confers dorsal compartment identity in Drosophila wing development. Apterous activity requires formation of a complex with a co-factor, Chip/dLDB. Apterous activity is regulated during wing development by dLMO, which competes with Apterous for complex formation. Here, we present evidence that complex formation between Apterous, Chip and DNA stabilizes Apterous protein in vivo. We also report that a difference in the ability of Chip to bind the LIM domains of Apterous and dLMO contributes to regulation of activity levels in vivo.     

Morphological intergration between development compartments in the Drosophila wing

Abstract. Developmental integration is the covariation among morphological structures due to connections between the developmental processes that built them. Here we use the methods of geometric morphometrics to study integration in the wing of Drosophila melanogaster . In particular, we focus on the hypothesis that the anterior and posterior wing compartments are separate developmental units that vary independently. We measured both variation among genetically diverse individuals and random differences between body sides of single individuals (fluctuating asymmetry, FA). For both of these sources of variation, the patterns of variation identified by principal component analyses all involved landmarks in both the anterior and posterior compartments simultaneously. Analyses focusing exclusively on the covariation between the anterior and posterior compartments, by the partial least-squares method, revealed pervasive integration of the two compartments, for both individual variation and FA. These analyses clearly indicate that the anterior and posterior compartments are not separate units of variation, but that the covariation between compartments is sufficient to account for nearly all the variation throughout the entire wing. We conclude that variation among individuals as well as the developmental perturbations responsible for FA generate shape variation primarily through developmental processes that are integrated across both compartments. In contrast, much less of the shape variation in our sample can be attributed to the localized processes that establish the identity of particular wing veins.