Drosophila Eye Imaginal Disc Research Articles

Ectopic expression of human p53 inhibits entry into S phase and induces apoptosis in the Drosophila eye imaginal disc.

Transgenic flies in which ectopic expression of human p53 was targeted to the Drosophila eye imaginal disc were established. On sectioning of adult fly eyes which displayed a severe rough eye phenotype, most ommatidia were found to be fused and irregular shapes of rabdomeres were observed. In addition, many pigment cells were lost. In the developing eye imaginal disc, photoreceptor cell differentiation was initiated normally despite the ectopic expression of p53. However, expression of p53 inhibited cell cycle progression in eye imaginal disc cells and the S phase zone (the second mitotic wave) behind the morphogenetic furrow was almost completely abolished. Furthermore, expression of p53 induced extensive apoptosis of eye imaginal disc cells, and co-expression of baculovirus P35 in the eye imaginal disc suppressed the p53-induced rough eye phenotype. These results are consistent with the known functions of human p53 and indicate the existence of signaling systems with elements corresponding to human p53 in Drosophila eye imaginal disc cells. Genetic crosses of transgenic flies expressing p53 to a collection of Drosophila deficiency stocks allowed us to identify several genomic regions, deletions of which caused enhancement or suppression of the p53-induced rough eye phenotype. The transgenic flies established in this study should be useful to identify novel targets of p53 and its positive or negative regulators in Drosophila.

Eya1-deficient mice lack ears and kidneys and show abnormal apoptosis of organ primordia.

Haploinsufficiency for human EYA1, a homologue of the Drosophila melanogaster gene eyes absent (eya), results in the dominantly inherited disorders branchio-oto-renal (BOR) syndrome and branchio-oto (BO) syndrome, which are characterized by craniofacial abnormalities and hearing loss with (BOR) or without (BO) kidney defects. To understand the developmental pathogenesis of organs affected in these syndromes, we inactivated the gene Eya1 in mice. Eya1 heterozygotes show renal abnormalities and a conductive hearing loss similar to BOR syndrome, whereas Eya1 homozygotes lack ears and kidneys due to defective inductive tissue interactions and apoptotic regression of the organ primordia. Inner ear development in Eya1 homozygotes arrests at the otic vesicle stage and all components of the inner ear and specific cranial sensory ganglia fail to form. In the kidney, Eya1 homozygosity results in an absence of ureteric bud outgrowth and a subsequent failure of metanephric induction. Gdnf expression, which is required to direct ureteric bud outgrowth via activation of the c-ret Rtk (refs 5, 6, 7, 8), is not detected in Eya1-/- metanephric mesenchyme. In Eya1-/- ear and kidney development, Six but not Pax expression is Eya1 dependent, similar to a genetic pathway elucidated in the Drosophila eye imaginal disc. Our results indicate that Eya1 controls critical early inductive signalling events involved in ear and kidney formation and integrate Eya1 into the genetic regulatory cascade controlling kidney formation upstream of Gdnf. In addition, our results suggest that an evolutionarily conserved Pax-Eya-Six regulatory hierarchy is used in mammalian ear and kidney development.

Decapentaplegic and wingless are regulated by eyes absent and eyegone and interact to direct the pattern of retinal differentiation in the eye disc.

Signaling by the secreted hedgehog, decapentaplegic and wingless proteins organizes the pattern of photoreceptor differentiation within the Drosophila eye imaginal disc; hedgehog and decapentaplegic are required for differentiation to initiate at the posterior margin and progress across the disc, while wingless prevents it from initiating at the lateral margins. Our analysis of these interactions has shown that initiation requires both the presence of decapentaplegic and the absence of wingless, which inhibits photoreceptor differentiation downstream of the reception of the decapentaplegic signal. However, wingless is unable to inhibit differentiation driven by activation of the epidermal growth factor receptor pathway. The effect of wingless is subject to regional variations in control, as the anterior margin of the disc is insensitive to wingless inhibition. The eyes absent and eyegone genes encode members of a group of nuclear proteins required to specify the fate of the eye imaginal disc. We show that both eyes absent and eyegone are required for normal activation of decapentaplegic expression at the posterior and lateral margins of the disc, and repression of wingless expression in presumptive retinal tissue. The requirement for eyegone can be alleviated by inhibition of the wingless signaling pathway, suggesting that eyegone promotes eye development primarily by repressing wingless. These results provide a link between the early specification and later differentiation of the eye disc.

The sidekick gene, a member of the immunoglobulin superfamily, is required for pattern formation in the Drosophila eye.

In the Drosophila eye imaginal disc the photoreceptor cells (R cells) differentiate according to a precise spatial and temporal order. The sidekick (sdk) gene is necessary to prevent extra R cells from differentiating during eye disc development. The extra cell appears between R3 and R4 early in R cell clusters and is most likely the result of the mystery cell inappropriately differentiating as an R cell. Mosaic analysis shows that sdk is required neither in the R cells nor in the extra cell, suggesting that sdk is necessary in the surrounding undifferentiated cells. The sdk gene codes for a protein that is a member of the immunoglobulin superfamily, having six immunoglobulin domains, thirteen fibronectin repeats and a transmembrane domain. The protein structure is consistent with its participation in cell-cell interaction during eye development.

The regulation of hedgehog and decapentaplegic during Drosophila eye imaginal disc development

The hedgehog signalling pathway is a conserved mechanism which acts in inductive processes in both vertebrate and invertebrate development to direct growth and patterning. in Drosophila, the secreted Hedgehog protein acts as a signal to induce non-autonomous activation in adjacent cells of either the decapentaplegic or wingless genes (both of which encode growth factor-like molecules), via inactivation of patched activity. In the eye disc, this pathway drives progression of the morphogenetic furrow, while in the wing (and leg and antennal) discs it is required to set up an organising centre along the anteroposterior compartment boundary. We have compared the regulation and function of hedgehog pathway activity in the eye and wing discs, and find that there are significant differences. Whereas in the wing disc, engrailed function is required for hedgehog expression, in the eye disc activation and maintenance of hedgehog expression is achieved independently of engrailed. Regulation of decapentaplegic expression also differs: in the wing disc it is repressed in the anterior compartment by patched and in the posterior compartment by engrailed. In the eye disc, however, it is repressed posterior to the morphogenetic furrow in the absence of either patched or engrailed activity. We conclude that in the eye disc there are novel aspectsto hedgehog pathway function. Moreover, engrailed does not play an essential conserved role.

Mutations in the Drosophila hook gene inhibit endocytosis of the boss transmembrane ligand into multivesicular bodies.

Transmembrane ligands can be internalized across cell boundaries into receptor-expressing cells. In the developing Drosophila eye imaginal disc, the bride of sevenless transmembrane protein (boss) is expressed on the surface of R8 cells. After internalization into neighboring R7 cells, the boss protein accumulates in multivesicular bodies. In a search for genes that affect this cell-type-specific pattern of boss endocytosis, we found that mutations in the hook gene inhibit the accumulation of boss in multivesicular bodies of R7 cells. In addition, hook flies exhibit pleiotropic phenotypes including abnormal bristle morphology and eye degeneration. The wild-type-pattern of boss endocytosis was restored in hook mutants by a genomic rescue fragment containing the hook gene or by a hook cDNA expressed in R7 cells under control of a sevenless (sev) enhancer. The hook gene encodes a novel cytoplasmic protein of 679 amino acids with a central coiled-coil domain of some 200 amino acids. Truncated, epitope-tagged hook proteins coimmunoprecipitated the full-length protein, indicating dimerization mediated by the coiled-coil domain. The hook protein localizes to vesicular structures that are part of the endocytic compartment. The requirement of the hook protein in R7 cells for the accumulation of boss protein in multivesicular bodies, and the localization of the hook protein to endocytic vesicles indicate that the hook gene encodes a novel component of the endocytic compartment that plays an important role in the endocytosis of transmembrane ligands or their transport to multivesicular bodies.

Wingless inhibits morphogenetic furrow movement in the Drosophila eye disc.

Differentiation of the Drosophila eye imaginal disc is an asynchronous, repetitive process which proceeds across the disc from posterior to anterior. Its propagation correlates with the expression of decapentaplegic at the front of differentiation, in the morphogenetic furrow. Both differentiation and decapentaplegic expression are maintained by Hedgehog protein secreted by the differentiated cells posterior to the furrow. However, their initiation at the posterior margin occurs prior to hedgehog expression by an unknown mechanism. We show here that the wingless gene contributes to the correct spatial localization of initiation. Initiation of the morphogenetic furrow is restricted to the posterior margin by the presence of wingless at the lateral margins; removal of wingless allows lateral initiation. Ectopic expression of wingless at the posterior margin can also inhibit normal initiation. In addition, the presence of wingless in the center of the disc can prevent furrow progression. These effects of wingless are achieved without altering the expression of decapentaplegic.

Shortsighted acts in the decapentaplegic pathway in Drosophila eye development and has homology to a mouse TGF-β-responsive gene

Differentiation in the Drosophila eye imaginal disc traverses the disc as a wave moving from posterior to anterior. The propagation of this wave is driven by hedgehog protein secreted by the differentiated cells in the posterior region of the disc. Hedgehog induces decapentaplegic expression at the front of differentiation, in the morphogenetic furrow. We have identified a gene, shortsighted, which is expressed in a hedgehog-dependent stripe in the undifferentiated cells just anterior to the furrow and which appears to be involved in the transmission of the differentiation-inducing signal; a reduction in shortsighted function leads to a delay in differentiation and to a loss of photoreceptors in the adult. shortsighted is also required for a morphogenetic movement in the brain that reorients the second optic lobe relative to the first. shortsighted encodes a cytoplasmic leucine zipper protein with homology to a mouse gene, TSC-22, which is transcriptionally induced in response to TGF-beta.

Star Is Required for Neuronal Differentiation in the Drosophila Retina and Displays Dosage-Sensitive Interactions with Ras1

The photoreceptor cells R8, R2, and R5 are the first cells to initiate neuronal differentiation in the Drosophila eye imaginal disc. We have shown previously that these three cells require Star gene function for proper ommatidial assembly. Due to the embryonic lethality associated with mutations in Star, we have analyzed the consequence of loss of Star function in mosaic eye imaginal discs. Presumptive R8, R2, and R5 cells that lack Star function fail to differentiate neuronally and die a few hours later. Enhancer trap insertions reveal that Star expression in the eye disc is restricted to the developing R8, R2, and R5 cells. Taken together, these data suggest that Star is required for the reception of a signal and/or the execution of a developmental program that leads to the neuronal differentiation of R8, R2, and R5. Star is also required for the formation of wing veins and is expressed in developing veins, suggesting that at least partially overlapping pathways may operate during photoreceptor cell differentiation and wing vein formation. The role of Star in cell-cell signaling is supported by the observation of genetic interactions between Star and mutations that reduce signaling through both sevenless and the Drosophila EGF-receptor homologue, including Ras1 and Son of sevenless.

The structure of protein phosphatase 2A is as highly conserved as that of protein phosphatase I

cDNA coding for protein phosphatase 2A (PP2A) has been isolated from Drosophila head and eye imaginal disc libraries. Drosophila PP2A mRNA is expressed throughout development, but is most abundant in the early embryo. The cDNA hybridises to a single site on the left area of the second chromosomes at position 28D2-4. The deduced amino acid sequence (309 residues) of Drosophila PP2A shows 94% identity with either rabbit PP2Aα or PP2Aβ, indicating that PP2A maybe the most conserved of all known enzymes.