Cell patterning in the Drosophila segment: spatial regulation of the segment polarity gene patched

Intrasegmental patterning in the Drosophila embryo requires the activity of the segment polarity genes. The acquisition of positional information by cells during embryogenesis is reflected in the dynamic patterns of expression of several of these genes. In the case of patched, early ubiquitous expression is followed by its repression in the anterior portion of each parasegment; subsequently each broad band of expression splits into two narrow stripes. In this study we analyse the contribution of other segment polarity gene functions to the evolution of this pattern; we find that the first step in patched regulation is under the control of engrailed whereas the second requires the activity of both cubitus interruptusD and patched itself. Furthermore, the products of engrailed, wingless and hedgehog are essential for maintaining the normal pattern of expression of patched.     

Mutations in the human homologue of the Drosophila segment polarity gene patched in oral squamous cell carcinoma cell lines

In the present study, we have analyzed tumor deoxyribonucleic acid from oral squamous cell carcinoma (OSCC) cells for patched mutations using an exon-by-exon single strand conformation polymorphism assay and direct sequencing. We found two missense mutations which affected the conserved residue in the transmembrane domains of the gene product and in the intracellular loop at the C-terminal residue implicated in regulating the smoothened molecule. In addition, we demonstrated that the N-terminal fragment of sonic hedgehog (Shh-N) stimulates the growth of normal epithelial cells, the OSCC cell line, NA, and the salivary gland adenocarcinoma cell lines, HSG and HSY, which have no detectable mutation in patched. On the other hand, Shh has no effect on human SCC cells (UE, KA, KO, NI, A431 cells) that have mutations in patched. These results strongly suggest that an Shh-patched signaling is involved in the cell growth of oral epithelial cells and in the tumorigenesis of OSCCs.     

The segment polarity gene costal-2 in Drosophila. II. The origin of imaginal pattern duplications

Imaginal pattern duplications caused by hypomorphic expression of the segment polarity gene costal-2 are described. These affect the anteroposterior coordinate of the imaginal disc. A very small part of the pattern is deleted and a large number of additional pattern elements arise in a progressive order, anterior-most first followed by more and more posterior structures. Mosaic analyses show that the duplications arise nonautonomously in the larval stages but that the costal-2 gene is not required after early embryogenesis. Arguments that the duplications are the result of cell interactions and intercalary growth that themselves arise from an abnormal polarity of the embryonic segment are presented.     

frizzled Gene expression and development of tissue polarity in the Drosophila wing

Almost every cell in the Drosophila pupal wing forms a single, distally pointing cuticular hair. The function of the frizzled (fz) gene is essential for the elaboration of the normal wing hair pattern. In the absence of fz function hairs develop, but they display an abnormal polarity. We have examined the developmental expression of the fi gene at the RNA level via in situ hybridization and at the protein level via Western blotting. We have found that fz is expressed in all regions of the epidermis before, during, and after the fz cold sensitive period. We have also found that fz function is not required for normal fi expression. We have further found that mutations in several other tissue polarity genes do not noticeably alter the expression or the modification state of the Fz protein. © 1994 Wiley-Liss, Inc.     

The core planar cell polarity gene prickle interacts with flamingo to promote sensory axon advance in the Drosophila embryo

The atypical cadherin Drosophila protein Flamingo and its vertebrate homologues play widespread roles in the regulation of both dendrite and axon growth. However, little is understood about the molecular mechanisms that underpin these functions. Whereas flamingo interacts with a well-defined group of genes in regulating planar cell polarity, previous studies have uncovered little evidence that the other core planar cell polarity genes are involved in regulation of neurite growth. We present data in this study showing that the planar cell polarity gene prickle interacts with flamingo in regulating sensory axon advance at a key choice point — the transition between the peripheral nervous system and the central nervous system. The cytoplasmic tail of the Flamingo protein is not required for this interaction. Overexpression of another core planar cell polarity gene dishevelled produces a similar phenotype to prickle mutants, suggesting that this gene may also play a role in regulation of sensory axon advance.     

Dynamics of decapentaplegic expression during regeneration of the Drosophila melanogaster wing imaginal disc

Regeneration of an imaginal disc involves highly ordered proliferation and pattern regulation of the newly formed tissue. Although the general principles of imaginal disc regeneration have been extensively studied, knowledge of the underlying molecular mechanisms is far from complete. Results from other model organisms suggest that regeneration is the result of local recapitulation of the normal patterning genes. To analyze the dynamics of one major Drosophila patterning gene, decapentaplegic (dpp), in wing imaginal disc regeneration, a vital GFP reporter together with iontophoretic cell labeling were used. Our observations reveal that the restoration of compartment-border-specific dpp expression is a common event in imaginal disc regeneration. However, we did not find evidence of an upregulation of dpp expression during the regeneration process.     

Cell biology of planar polarity transmission in the Drosophila wing

The coordination of epithelial planar polarization is a critical step in the formation of well-ordered tissues. The process has been extensively studied in Drosophila, where genetic analysis has identified a set of "tissue polarity" genes that serve to coordinate planar polarity of cells in the developing wings, bristles and eyes. In the last several years, it has emerged that six of these genes encode junctional proteins. In the wing epithelium, these proteins undergo a polarized redistribution, forming separate proximal and distal cortical domains within each cell. The mechanisms that mediate cortical polarization and cue its direction have been the subject of intense investigation. Cuing the orientation of cortical polarization appears to depend on the atypical Cadherins Fat and Dachsous, although these proteins do not become polarized themselves, nor do they colocalize with components of polarized cortical domains. Interestingly, these Cadherins also act at earlier developmental stages to polarize tissue growth along the proximal-distal axis and it will be interesting to see whether these processes are mechanistically related. Once the axis of polarization is determined, cortical polarity seems to be propagated, at least locally, by a cascade of direct cell-cell interactions mediated by the proximal and distal domains. The cell biological mechanisms leading to polarization are still unclear, but the process depends on the control of Protein Phosphatase 2A activity by its regulatory subunit, Widerborst. Interestingly, Widerborst is found on a planar web of microtubules with connections to apical junctions, suggesting that these microtubules may have an important function in polarizing the cortex.     

A role for the segment polarity gene shaggy/GSK-3 in the Drosophila circadian clock.

Tissue-specific overexpression of the glycogen synthase kinase-3 (GSK-3) ortholog shaggy (sgg) shortens the period of the Drosophila circadian locomotor activity cycle. The short period phenotype was attributed to premature nuclear translocation of the PERIOD/TIMELESS heterodimer. Reducing SGG/GSK-3 activity lengthens period, demonstrating an intrinsic role for the kinase in circadian rhythmicity. Lowered sgg activity decreased TIMELESS phosphorylation, and it was found that GSK-3 beta specifically phosphorylates TIMELESS in vitro. Overexpression of sgg in vivo converts hypophosphorylated TIMELESS to a hyperphosphorylated protein whose electrophoretic mobility, and light and phosphatase sensitivity, are indistinguishable from the rhythmically produced hyperphosphorylated TIMELESS of wild-type flies. Our results indicate a role for SGG/GSK-3 in TIMELESS phosphorylation and in the regulated nuclear translocation of the PERIOD/TIMELESS heterodimer.     

blistery encodes Drosophila tensin protein and interacts with integrin and the JNK signaling pathway during wing development

Tensin is an actin-binding protein that is localized in focal adhesions. At focal adhesion sites, tensin participates in the protein complex that establishes transmembrane linkage between the extracellular matrix and cytoskeletal actin filaments. Even though there have been many studies on tensin as an adaptor protein, the role of tensin during development has not yet been clearly elucidated. Thus, this study was designed to dissect the developmental role of tensin by isolating Drosophila tensin mutants and characterizing its role in wing development. The Drosophila tensin loss-of-function mutations resulted in the formation of blisters in the wings, which was due to a defective wing unfolding process. Interestingly, by(1)-the mutant allele of the gene blistery (by)-also showed a blistered wing phenotype, but failed to complement the wing blister phenotype of the Drosophila tensin mutants, and contains Y62N/T163R point mutations in Drosophila tensin coding sequences. These results demonstrate that by encodes Drosophila tensin protein and that the Drosophila tensin mutants are alleles of by. Using a genetic approach, we have demonstrated that tensin interacts with integrin and also with the components of the JNK signaling pathway during wing development; overexpression of by in wing imaginal discs significantly increased JNK activity and induced apoptotic cell death. Collectively, our data suggest that tensin relays signals from the extracellular matrix to the cytoskeleton through interaction with integrin, and through the modulation of the JNK signal transduction pathway during Drosophila wing development.     

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.     

The tissue polarity gene nemo carries out multiple roles in patterning during Drosophila development

Drosophila nemo was first identified as a gene required for tissue polarity during ommatidial development. We have extended the analysis of nemo and found that it participates in multiple developmental processes. It is required during wing development for wing shape and vein patterning. We observe genetic interactions between nemo and mutations in the Notch, Wingless, Frizzled and Decapentaplegic pathways. Our data support the findings from other organisms that Nemo proteins act as negative regulators of Wingless signaling. nemo mutations cause polarity defects in the adult wing and overexpression of nemo leads to abdominal polarity defects. The expression of nemo during embryogenesis is dynamic and dsRNA inhibition and ectopic expression studies indicate that nemo is essential during embryogenesis.     

The tricornered Ser/Thr protein kinase is regulated by phosphorylation and interacts with furry during Drosophila wing hair development

The Trc/Ndr/Sax1/Cbk1 family of ser/thr kinases plays a key role in the morphogenesis of polarized cell structures in flies, worms, and yeast. Tricornered (Trc), the Drosophila nuclear Dbf2-related (Ndr) serine/threonine protein kinase, is required for the normal morphogenesis of epidermal hairs, bristles, laterals, and dendrites. We obtained in vivo evidence that Trc function was regulated by phosphorylation and that mutations in key regulatory sites resulted in dominant negative alleles. We found that wild-type, but not mutant Trc, is found in growing hairs, and we failed to detect Trc in pupal wing nuclei, implying that in this developmental context Trc functions in the cytoplasm. The furry gene and its homologues in yeast and Caenorhabditis elegans have previously been implicated as being essential for the function of the Ndr kinase family. We found that Drosophila furry (Fry) also is found in growing hairs, that its subcellular localization is dependent on Trc function, and that it can be coimmunoprecipitated with Trc. Our data suggest a feedback mechanism involving Trc activity regulates the accumulation of Fry in developing hairs.