Wingless signaling and the control of cell shape in Drosophila wing imaginal discs

The control of cell morphology is important for shaping animals during development. Here we address the role of the Wnt/Wingless signal transduction pathway and two of its target genes, vestigial and shotgun (encoding E-cadherin), in controlling the columnar shape of Drosophila wing disc cells. We show that clones of cells mutant for arrow (encoding an essential component of the Wingless signal transduction pathway), vestigial or shotgun undergo profound cell shape changes and are extruded towards the basal side of the epithelium. Compartment-wide expression of a dominant-negative form of the Wingless transducer T-cell factor (TCF/Pangolin), or double-stranded RNA targeting vestigial or shotgun, leads to abnormally short cells throughout this region, indicating that these genes act cell autonomously to maintain normal columnar cell shape. Conversely, overexpression of Wingless, a constitutively-active form of the Wingless transducer β-catenin/Armadillo, or Vestigial, results in precocious cell elongation. Co-expression of Vestigial partially suppresses the abnormal cell shape induced by dominant-negative TCF. We conclude that Wingless signal transduction plays a cell-autonomous role in promoting and maintaining the columnar shape of wing disc cells. Furthermore, our data suggest that Wingless controls cell shape, in part, through maintaining vestigial expression.     

Ectopic Repo suppresses expression of castor gene in Drosophila central nervous system

The ventral nerve cord (VNC) of the Drosophila embryo is derived from neuroblasts (NBs). NBs divide in a stem cell lineage to generate a series of ganglion mother cells (GMCs), each of which divides once to produce a pair of neurons or glial cells. One of the NB genes, castor (cas), is expressed in a subset of NBs and has never been identified in neurons and the peripheral nervous system; cas plays a role in axonogenesis. But its limited expression along the dorsal-ventral axis within the central nervous system has not been investigated yet. In the present study, we examined the expression patterns of both genes using confocal microscopy to determine the effects of repo mutation on cas expression. Cas was mainly expressed in layers different from repo-expressed layers during early embryogenesis: repo was expressed mostly from deep to mid layers, while cas, from mid to superficial layers. Loss-of-function of repo did not result in an ectopic expression of cas, but rather, a scattering of cas-expressing cells. However, repo gain-of-function mutation caused repression of cas. In addition, repo-expressing cells seemed to block the migration of cas-expressing cells.     

Spalt major controls the development of the notum and of wing hinge primordia of the Drosophila melanogaster wing imaginal disc

The Drosophila wing and the dorsal thorax develop from primordia within the wing imaginal disc. Here we show that spalt major (salm) is expressed within the presumptive dorsal body wall primordium early in wing disc development to specify notum and wing hinge tissue. Upon ectopic salm expression, dorsally located second leg disc cells develop notum and wing hinge tissue instead of sternopleural tissue. Similarly, by salm over-expression within the wing disc, wing blade formation is suppressed and a mirror-image duplication of the notum and wing hinge is formed. In large dorsal clones, which lack salm and its neighboring paralogue spalt related (salr), the cells of the notum primordium do not grow; these dorsal cells are not specified as notum, hence no notum outgrowth develops. These results suggest that the zinc finger factors encoded by the salm/salr complex play important roles in defining cells of the early wing disc as dorsal body wall cells, which develop into a large dorsal body wall territory and form mesonotum and some wing hinge tissue, and in delimiting the wing primordium. We also find that salm activity is down-regulated by its own product and by that of the Pax gene eyegone.     

Temporal and spatial windows delimit activation of the outer ring of wingless in the Drosophila wing

Extracellular signalling molecules play many roles in the development of higher organisms. They are used reiteratively in different tissues and stages, but the response of the receiving cells is controlled in a context dependent manner. The pattern of expression of the signalling molecule Wingless/WNT in Drosophila is extraordinarily complex. We have studied the mechanism that controls its expression and function in the outer ring of the Drosophila wing hinge. Our findings indicate that wingless expression is controlled by a dual mechanism: its initial activation requires the product of zinc finger homeodomain 2 and is subsequently repressed by the product of the gene complex elbow/no ocelli. This tight regulation restricts the activation of wingless temporally and spatially. Later in development, wingless expression is maintained by an autoregulatory loop that involves the product of homothorax. We have analyzed the phenotype of a wingless allelic combination that specifically removes the outer ring, and our results show that Wingless is required to promote local proliferation of the wing base cells. Thus, cell proliferation in the proximal-distal axis is controlled by the sequential activation of wingless in the inner ring and the outer ring at different stages of development.     

Regulation of EGFR and Notch signaling by distinct isoforms of D-cbl during Drosophila development

Cells receive and interpret extracellular signals to regulate cellular responses such as proliferation, cell survival and differentiation. However, proper inactivation of these signals is critical for appropriate homeostasis. Cbl proteins are E3-ubiquitin ligases that restrict receptor tyrosine kinase (RTK) signaling, most notably EGFR (Epidermal Growth Factor Receptor), via the endocytic pathway. Consistently, many mutant phenotypes of Drosophila cbl (D-cbl) are due to inappropriate activation of EGFR signaling. However, not all D-cbl phenotypes can be explained by increased EGFR activity. Here, we report that D-Cbl also negatively regulates Notch activity during eye and wing development. D-cbl produces two isoforms by alternative splicing. The long isoform, D-CblL, regulates the EGFR. We found that the short isoform, D-CblS, preferentially restricts Notch signaling. Specifically, our data imply that D-CblS controls the activity of the Notch ligand Delta. Taken together, these data suggest that D-Cbl controls the EGFR and Notch/Delta signaling pathways through production of two alternatively spliced isoforms during development in Drosophila.     

Myoblast fusion in Drosophila

The body wall musculature of a Drosophila larva is composed of an intricate pattern of 30 segmentally repeated muscle fibers in each abdominal hemisegment. Each muscle fiber has unique spatial and behavioral characteristics that include its location, orientation, epidermal attachment, size and pattern of innervation. Many, if not all, of these properties are dictated by founder cells, which determine the muscle pattern and seed the fusion process. Myofibers are then derived from fusion between a specific founder cell and several fusion competent myoblasts (FCMs) fusing with as few as 3-5 FCMs in the small muscles on the most ventral side of the embryo and as many as 30 FCMs in the larger muscles on the dorsal side of the embryo. The focus of the present review is the formation of the larval muscles in the developing embryo, summarizing the major issues and players in this process. We have attempted to emphasize experimentally-validated details of the mechanism of myoblast fusion and distinguish these from the theoretically possible details that have not yet been confirmed experimentally. We also direct the interested reader to other recent reviews that discuss myoblast fusion in Drosophila, each with their own perspective on the process [1], [2], [3] and [4]. With apologies, we use gene nomenclature as specified by Flybase (http://flybase.org) but provide Table 1 with alternative names and references.     

The deubiquitinase emperor's thumb is a regulator of apoptosis in Drosophila

We have characterized the gene emperor's thumb (et) and showed that it is required for the regulation of apoptosis in Drosophila. Loss-of-function mutations in et result in apoptosis associated with a decrease in the concentration of DIAP1. Overexpression of one form of et inhibits apoptosis, consistent with et having an anti-apoptotic function; however, overexpression of a second form of et induces apoptosis, indicating that the two forms of et may have competing functions. et encodes a protein deubiquitinase, suggesting it regulates apoptosis by controlling the stability of apoptotic regulatory proteins.     

The Him gene inhibits the development of Drosophila flight muscles during metamorphosis

During Drosophila metamorphosis some larval tissues escape the general histolysis and are remodelled to form adult tissues. One example is the dorso-longitudinal muscles (DLMs) of the indirect flight musculature. They are formed by an intriguing process in which residual larval oblique muscles (LOMs) split and fuse with imaginal myoblasts associated with the wing disc. These myoblasts arise in the embryo, but remain undifferentiated throughout embryogenesis and larval life, and thus share characteristics with mammalian satellite cells. However, the mechanisms that maintain the Drosophila myoblasts in an undifferentiated state until needed for LOM remodelling are not understood. Here we show that the Him gene is expressed in these myoblasts, but is undetectable in developing DLM fibres. Consistent with this, we found that Him could inhibit DLM development: it inhibited LOM splitting and resulted in fibre degeneration. We then uncovered a balance between mef2, a positive factor required for proper DLM development, and the inhibitory action of Him. Mef2 suppressed the inhibitory effect of Him on DLM development, while Him could suppress the premature myosin expression induced by mef2 in myoblasts. Furthermore, either decreased Him function or increased mef2 function disrupted DLM development. These findings, together with the co-expression of Him and Mef2 in myoblasts, indicate that Him may antagonise mef2 function during normal DLM development and that Him participates in a balance of signals that controls adult myoblast differentiation and remodelling of these muscle fibres. Lastly, we provide evidence for a link between Notch function and Him and mef2 in this balance.     

On the roles of Notch, Delta, kuzbanian, and inscuteable during the development of Drosophila embryonic neuroblast lineages

The generation of cellular diversity in the nervous system involves the mechanism of asymmetric cell division. Besides an array of molecules, including the Par protein cassette, a heterotrimeric G protein signalling complex, Inscuteable plays a major role in controlling asymmetric cell division, which ultimately leads to differential activation of the Notch signalling pathway and correct specification of the two daughter cells. In this context, Notch is required to be active in one sibling and inactive in the other. Here, we investigated the requirement of genes previously known to play key roles in sibling cell fate specification such as members of the Notch signalling pathway, e.g., Notch (N), Delta (Dl), and kuzbanian (kuz) and a crucial regulator of asymmetric cell division, inscuteable (insc) throughout lineage progression of 4 neuroblasts (NB1-1, MP2, NB4-2, and NB7-1). Notch-mediated cell fate specification defects were cell-autonomous and were observed in all neuroblast lineages even in cells born from late ganglion mother cells (GMC) within the lineages. We also show that Dl functions non-autonomously during NB lineage progression and clonal cells do not require Dl from within the clone. This suggests that within a NB lineage Dl is dispensable for sibling cell fate specification. Furthermore, we provide evidence that kuz is involved in sibling cell fate specification in the central nervous system. It is cell-autonomously required in the same postmitotic cells which also depend on Notch function. This indicates that KUZ is required to facilitate a functional Notch signal in the Notch-dependent cell for correct cell fate specification. Finally, we show that three neuroblast lineages (NB1-1, NB4-2, and NB7-1) require insc function for sibling cell fate specification in cells born from early GMCs whereas insc is not required in cells born from later GMCs of the same lineages. Thus, there is differential requirement for insc for cell fate specification depending on the stage of lineage progression of NBs.     

JNK signaling pathway required for wound healing in regenerating Drosophila wing imaginal discs

We have examined wound healing during regeneration of Drosophila wing imaginal discs fragments by confocal microscopy and assessed the role of components of the JNK pathway in this process. After cutting, columnar and peripodial epithelia cells at the wound edge start to close the wound through formation and contraction of an actin cable. This is followed by a zipping process through filopodial protrusions from both epithelia knitting the wound edges from proximal to distal areas of the disc. Activation of the JNK pathway is involved in such process. puckered (puc) expression is induced in several rows of cells at the edge of the wound, whereas absence of JNK pathway activity brought about by hemipterous, basket, and Dfos mutants impair wound healing. These defects are accompanied by lowered or loss of expression of puc. In support of a role of puc in wound healing, hep mutant phenotypes are rescued by reducing puc function, whereas overexpression of puc inhibits wound healing. Altogether, these results demonstrate a role for the JNK pathway in imaginal disc wound healing, similar to that reported for other healing processes such as embryonic dorsal closure, thoracic closure, and adult epithelial wound healing in Drosophila. Differences with such processes are also highlighted.