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The vrille gene of Drosophila is a maternal enhancer of decapentaplegic and encodes a new member of the bZIP family of transcription factors.

We report here the genetical and molecular characterization of a new Drosophila zygotic lethal locus, vrille (vri). Vri alleles act not only as dominant maternal enhancers of embryonic dorsoventral patterning defects caused by easter and decapentaplegic (dpp) mutations, but also as dominant zygotic enhancers of dpp alleles for phenotypes in wing. The vri gene encodes a new member of the bZIP family of transcription factors closely related to gene 9 of Xenopus laevis, induced by thyroid hormone during the tadpole tail resorption program, and NF-IL3A, a human T cell transcription factor that transactivates the interleukin-3 promoter. NF-IL3A shares 93% similarity and 60% identity with Vri for a stretch of 68 amino acids that includes the bZIP domain. Although all the alleles tested behave like antimorphs, the dominant enhancement is also seen with a nonsense mutation allele that prevents translation of the bZIP domain. Because of the strong domainant enhancement of dpp phenotypes by vri alleles in both embryo and wing, and also the similarity between the wing vein phenotypes caused by the vri and shortvein dpp alleles, we postulate that vri interacts either directly or indirectly with certain components of the dpp (a TGF beta homologue) signal transduction pathway.

Alleles↗

The catecholamines up (Catsup) protein of Drosophila melanogaster functions as a negative regulator of tyrosine hydroxylase activity.

We report the genetic, phenotypic, and biochemical analyses of Catecholamines up (Catsup), a gene that encodes a negative regulator of tyrosine hydroxylase (TH) activity. Mutations within this locus are semidominant lethals of variable penetrance that result in three broad, overlapping effective lethal phases (ELPs), indicating that the Catsup gene product is essential throughout development. Mutants from each ELP exhibit either cuticle defects or catecholamine-related abnormalities, such as melanotic salivary glands or pseudotumors. Additionally, Catsup mutants have significantly elevated TH activity that may arise from a post-translational modification of the enzyme. The hyperactivation of TH in Catsup mutants results in abnormally high levels of catecholamines, which can account for the lethality, visible phenotypes, and female sterility observed in these mutants. We propose that Catsup is a component of a novel system that downregulates TH activity, making Catsup the fourth locus found within the Dopa decarboxylase (Ddc) gene cluster that functions in catecholamine metabolism.

Animals↗

Inhibition of patterned cell shape change and cell invasion by Discs large during Drosophila oogenesis.

Drosophila Discs large (Dlg) is a tumor suppressor gene whose loss in epithelial tissues causes disrupted cell polarity and increased cell proliferation. A human Dlg homolog, hDlg, has been implicated in tumorigenic processes via its association with the product of the Adenomatous Polyposis Coli (APC) gene. We show for the first time that Drosophila Dlg is required to block cell invasion. Loss of dlg activity during oogenesis causes follicle cells to change shape and invade in a pattern similar to border cells, a small population of cells that break from the post-mitotic follicular epithelium during wild-type oogenesis, yet dlg mutant cells have not adopted a border cell fate. Both functional and morphological evidence indicates that cooperation between germ cell and follicle cell Dlg, probably mediated by Dlg PDZ domains, is crucial for regulating cell mixing, suggesting a novel developmental mechanism and mode of action for the Dlg family of molecules. These findings suggest that Dlg does not simply inhibit individual cell behaviors during oogenesis, but rather acts in a developmental pathway essential for blocking cell proliferation and migration in a spatio-temporally defined manner. A model for Dlg action in blocking cell invasion is presented.

Animals↗

Dorsal-ventral patterning of the Drosophila embryo depends on a putative negative growth factor encoded by the short gastrulation gene.

Pattern formation in the dorsal region of the Drosophila embryo depends on the activity of a small group of zygotically acting genes. dpp, a key gene in this group, encodes a TGF-beta-like product (Dpp) that has been proposed to function as a morphogen with peak levels of Dpp-specifying amnioserosa, the dorsal-most cell type, and lower Dpp levels specifying dorsal ectoderm. The short gastrulation gene also contributes to patterning the dorsal region, but unlike the other genes involved in this process, sog activity is only required in ventral cells. Genetic evidence indicates that sog functions to antagonize dpp activity. In this report we present further phenotypic characterization of sog mutant embryos in dorsal and lateral regions and describe the cloning of the sog locus. sog is expressed in a broad lateral stripe of cells that abuts the dorsal territory of dpp-expressing cells. sog is predicted to encode a protein with an internal signal sequence and a large extracellular domain containing four repeats of a novel motif defined by the spacing of 10 cysteine residues that is distantly related to domains present in thrombospondin and procollagen. We propose that one or more of these cysteine repeats can be liberated by proteolytic cleavage of the primary Sog protein. These putative soluble Sog peptides may then diffuse into the dorsal region to antagonize the activity of Dpp, leading to the subdivision of the dorsal territory into amnioserosa and dorsal ectoderm.

Amino Acid Sequence↗

A circadian output in Drosophila mediated by neurofibromatosis-1 and Ras/MAPK.

Output from the circadian clock controls rhythmic behavior through poorly understood mechanisms. In Drosophila, null mutations of the neurofibromatosis-1 (Nf1) gene produce abnormalities of circadian rhythms in locomotor activity. Mutant flies show normal oscillations of the clock genes period (per) and timeless (tim) and of their corresponding proteins, but altered oscillations and levels of a clock-controlled reporter. Mitogen-activated protein kinase (MAPK) activity is increased in Nf1 mutants, and the circadian phenotype is rescued by loss-of-function mutations in the Ras/MAPK pathway. Thus, Nf1 signals through Ras/MAPK in Drosophila. Immunohistochemical staining revealed a circadian oscillation of phospho-MAPK in the vicinity of nerve terminals containing pigment-dispersing factor (PDF), a secreted output from clock cells, suggesting a coupling of PDF to Ras/MAPK signaling.

Alleles↗

Interaction of short-range repressors with Drosophila CtBP in the embryo.

Human CtBP attenuates transcriptional activation and tumorigenesis mediated by the adenovirus E1A protein. The E1A sequence motif that interacts with CtBP, Pro-X-Asp-Leu-Ser-X-Lys (P-DLS-K), is present in the repression domains of two unrelated short-range repressors in Drosophila, Knirps and Snail, and is essential for the interaction of these proteins with Drosophila CtBP (dCtBP). A P-element-induced mutation in dCtBP exhibits gene-dosage interactions with a null mutation in knirps, which is consistent with the occurrence of Knirps-dCtBP interactions in vivo. These observations suggest that CtBP and dCtBP are engaged in an evolutionarily conserved mechanism of transcriptional repression, which is used in both Drosophila and mammals.

Alcohol Oxidoreductases↗

Cooperative regulation of cell polarity and growth by Drosophila tumor suppressors.

Loss of cell polarity and tissue architecture are characteristics of malignant cancers derived from epithelial tissues. We provide evidence from Drosophila that a group of membrane-associated proteins act in concert to regulate both epithelial structure and cell proliferation. Scribble (Scrib) is a cell junction-localized protein required for polarization of embryonic and, as demonstrated here, imaginal disc and follicular epithelia. We show that the tumor suppressors lethal giant larvae (lgl) and discs-large (dlg) have identical effects on all three epithelia, and that scrib also acts as a tumor suppressor. Scrib and Dlg colocalize and overlap with Lgl in epithelia; activity of all three genes is required for cortical localization of Lgl and junctional localization of Scrib and Dlg. scrib, dlg, and lgl show strong genetic interactions. Our data indicate that the three tumor suppressors act together in a common pathway to regulate cell polarity and growth control.

Animals↗

Genome-wide analysis of mRNAs regulated by Drosha and Argonaute proteins in Drosophila melanogaster.

RNA silencing pathways are conserved gene regulation mechanisms that elicit decay and/or translational repression of mRNAs complementary to short interfering RNAs and microRNAs (miRNAs). The fraction of the transcriptome regulated by these pathways is not known, but it is thought that each miRNA may have hundreds of targets. To identify transcripts regulated by silencing pathways at the genomic level, we examined mRNA expression profiles in Drosophila melanogaster cells depleted of four Argonaute paralogs (i.e., AGO1, AGO2, PIWI, or Aubergine) that play essential roles in RNA silencing. We also profiled cells depleted of the miRNA-processing enzyme Drosha. The results reveal that transcripts differentially expressed in Drosha-depleted cells have highly correlated expression in the AGO1 knockdown and are significantly enriched in predicted and validated miRNA targets. The levels of a subset of miRNA targets are also regulated by AGO2. Moreover, AGO1 and AGO2 silence the expression of a common set of mobile genetic elements. Together, these results indicate that the functional overlap between AGO1 and AGO2 in Drosophila is more important than previously thought.

Animals↗

The Drosophila cytokine receptor Domeless controls border cell migration and epithelial polarization during oogenesis.

In mammals, the JAK/STAT (Janus Kinase/Signal Transducer and Activator of Transcription) signaling pathway is activated in response to cytokines and growth factors to control blood cell development, proliferation and cell determination. In Drosophila, a conserved JAK/STAT signaling pathway controls segmentation in embryos, as well as blood cell development and other processes in larvae and adults. During embryogenesis, transduction of the Unpaired [Upd; also known as Outstretched (Os)] ligand through the JAK/STAT pathway requires Domeless, a putative membrane protein with distant homology to vertebrate type I cytokine receptors. We have isolated domeless (dome) in a screen to identify genes essential in epithelial morphogenesis during oogenesis. The level of dome activity is critical for proper border cell migration and is controlled in part through a negative feedback loop. In addition to its essential role in border cells, we show that dome is required in the germarium for the polarization of follicle cells during encapsulation of germline cells. In this process, dome controls the expression of the apical determinant Crumbs. In contrast to the ligand Upd, whose expression is limited to a pair of polar cells at both ends of the egg chamber, dome is expressed in all germline and follicle cells. However, the Dome protein is specifically localized at apicolateral membranes and undergoes ligand-dependent internalization in the follicle cells. dome mutations interact genetically with JAK/STAT pathway genes in border cell migration and abolish the nuclear translocation of Stat92E in vivo. We also show that dome functions downstream of upd and that both the extracellular and intracellular domains of Dome are required for JAK/STAT signaling. Altogether, our data indicate that Dome is an essential receptor molecule for Upd and JAK/STAT signaling during oogenesis.

Animals↗

E(z): a polycomb group gene or a trithorax group gene?

The products of the Polycomb group of genes are cooperatively involved in repressing expression of homeotic selector genes outside of their appropriate anterior/posterior boundaries. Loss of maternal and/or zygotic function of Polycomb group genes results in the ectopic expression of both Antennapedia Complex and Bithorax Complex genes. The products of the trithorax group of genes are cooperatively involved in maintaining active expression of homeotic selector genes within their appropriate anterior/posterior boundaries. Loss of maternal and/or zygotic function of trithorax group genes results in reduced expression of both Antennapedia Complex and Bithorax Complex genes. Although Enhancer of zeste has been classified as a member of the Polycomb group, in this paper we show that Enhancer of zeste can also be classified as a member of the trithorax group. The requirement for Enhancer of zeste activity as either a trithorax group or Polycomb group gene depends on the homeotic selector gene locus as well as on spatial and temporal cues.

Alleles↗

hermaphrodite and doublesex function both dependently and independently to control various aspects of sexual differentiation in Drosophila.

The hermaphrodite (her) gene is necessary for sexual differentiation in Drosophila. Our characterization of her's zygotic function suggests that one set of female-specific terminal differentiation genes, the yolk protein (yp) genes, is transcriptionally activated by two separate pathways. One is a female-specific pathway, which is positively regulated by the female-specific doublesex protein (DSXF). The other is a non-sex-specific pathway, that is positively regulated by HER. The HER pathway is prevented from functioning in males by the action of the male-specific doublesex protein (DSXM). The HER and DSX pathways also function independently to control downstream target genes in the precursor cells that give rise to the vaginal teeth and dorsal anal plate of females, and the lateral anal plates of males. However, a female-specific pathway that is dependent on both DSXF and HER controls the female-specific differentiation of the foreleg bristles and tergites 5 and 6, and the male-specific differentiation of these tissues does not require the suppression of HER's function by DSXM.

Animals↗

The CNS midline cells control the spitz class and Egfr signaling genes to establish the proper cell fate of the Drosophila ventral neuroectoderm.

The spitz class genes, pointed (pnt), rhomboid frho), single-minded (sim), spitz (spi)and Star (S), as well as the Drosophila epidermal growth factor receptor (Egfr) signaling genes, argos (aos), Egfr, orthodenticle (otd) and vein (vn), are required for the proper establishment of ventral neuroectodermal cell fate. The roles of the CNS midline cells, spitz class and Egfr signaling genes in cell fate determination of the ventral neuroectoderm were determined by analyzing the spatial and temporal expression patterns of each individual gene in spitz class and Egfr signaling mutants. This analysis showed that the expression of all the spitz class and Egfrsignaling genes is affected by the sim gene, which indicates that sim acts upstream of all the spitz class and Egfr signaling genes. It was shown that overexpression of sim in midline cells fails to induce the ectodermal fate in the spi and Egfr mutants. On the other hand, overexpression of spi and Draf causes ectopic expression of the neuroectodermal markers in the sim mutant. Ectopic expression of sim in the en-positive cells induces the expression of downstream genes such as otd, pnt, rho, and vn, which clearly demonstrates that the sim gene activates the EGFR signaling pathway and that CNS midline cells, specified by sim, provide sufficient positional information for the establishment of ventral neuroectodermal fate. These results reveal that the CNS midline cells are one of the key regulators for the proper patterning of the ventral neuroectoderm by controlling EGFR activity through the regulation of the expression of spitz class genes and Egfr signaling genes.

Animals↗

Probing for gene specificity in epithelial development.

We surveyed a total of 228 random insertions of a P[GawB] element to determine the fraction of regulatory regions in the Drosophila genome that activate gene expression specifically in follicle cells versus producing more complex patterns of expression. We monitored the GAL4 expression encoded by this construct in the ovarian follicle cells by crossing the lines to a strain containing a lacZ reporter construct. Sixty four per cent of the insertions showed ovarian expression. To assess the specificity of this expression, 124 of the 228 lines were crossed to strains containing either an activated form of Armadillo, the Drosophila homolog of beta-catenin, or an activated form of Torpedo/Egfr, the Drosophila homolog of the Epidermal Growth Factor receptor, under the control of GAL4 target sites. The lethality and imaginal disc phenotypes observed in these crosses suggest that most random insertions cause GAL4 expression in a variety of tissues. Very few insertions appear to drive expression only in follicle cells. Although the activated form of Armadillo produced higher frequencies of lethality and disk phenotypes, expression in the follicle cell epithelium at later stages of oogenesis did not lead to a visible phenotype. This contrasts with the dorsalized phenotypes observed in the combination of the same GAL4 lines with the activated Torpedo construct.

Animals↗

High sequence turnover in the regulatory regions of the developmental gene hunchback in insects.

Extensive sequence analysis of the developmental gene hunchback and its 5' and 3' regulatory regions in Drosophila melanogaster, Drosophila virilis, Musca domestica, and Tribolium castaneum, using a variety of computer algorithms, reveals regions of high sequence simplicity probably generated by slippage-like mechanisms of turnover. No regions are entirely refractory to the action of slippage, although the density and composition of simple sequence motifs varies from region to region. Interestingly, the 5' and 3' flanking regions share short repetitive motifs despite their separation by the gene itself, and the motifs are different in composition from those in the exons and introns. Furthermore, there are high levels of conservation of motifs in equivalent orthologous regions. Detailed sequence analysis of the P2 promoter and DNA footprinting assays reveal that the number, orientation, sequence, spacing, and protein-binding affinities of the BICOID-binding sites varies between species and that the 'P2' promoter, the nanos response element in the 3' untranslated region, and several conserved boxes of sequence in the gene (e.g., the two zinc-finger regions) are surrounded by cryptically-simple-sequence DNA. We argue that high sequence turnover and genetic redundancy permit both the general maintenance of promoter functions through the establishment of coevolutionary (compensatory) changes in cis- and trans-acting genetic elements and, at the same time, the possibility of subtle changes in the regulation of hunchback in the different species.

Amino Acid Sequence↗

Nuclear receptors of the honey bee: annotation and expression in the adult brain.

The Drosophila genome encodes 18 canonical nuclear receptors. All of the Drosophila nuclear receptors are here shown to be present in the genome of the honey bee (Apis mellifera). Given that the time since divergence of the Drosophila and Apis lineages is measured in hundreds of millions of years, the identification of matched orthologous nuclear receptors in the two genomes reveals the fundamental set of nuclear receptors required to 'make' an endopterygote insect. The single novelty is the presence in the A. mellifera genome of a third insect gene similar to vertebrate photoreceptor-specific nuclear receptor (PNR). Phylogenetic analysis indicates that this novel gene, which we have named AmPNR-like, is a new member of the NR2 subfamily not found in the Drosophila or human genomes. This gene is expressed in the developing compound eye of the honey bee. Like their vertebrate counterparts, arthropod nuclear receptors play key roles in embryonic and postembryonic development. Studies in Drosophila have focused primarily on the role of these transcription factors in embryogenesis and metamorphosis. Examination of an expressed sequence tag library developed from the adult bee brain and analysis of transcript expression in brain using in situ hybridization and quantitative RT-PCR revealed that several members of the nuclear receptor family (AmSVP, AmUSP, AmERR, AmHr46, AmFtz-F1, and AmHnf-4) are expressed in the brain of the adult bee. Further analysis of the expression of AmUSP and AmSVP in the mushroom bodies, the major insect brain centre for learning and memory, revealed changes in transcript abundance and, in the case of AmUSP, changes in transcript localization, during the development of foraging behaviour in the adult. Study of the honey bee therefore provides a model for understanding nuclear receptor function in the adult brain.

Amino Acid Sequence↗

Spliceosomal small nuclear RNA genes in 11 insect genomes.

The removal of introns from the primary transcripts of protein-coding genes is accomplished by the spliceosome, a large macromolecular complex of which small nuclear RNAs (snRNAs) are crucial components. Following the recent sequencing of the honeybee (Apis mellifera) genome, we used various computational methods, ranging from sequence similarity search to RNA secondary structure prediction, to search for putative snRNA genes (including their promoters) and to examine their pattern of conservation among 11 available insect genomes (A. mellifera, Tribolium castaneum, Bombyx mori, Anopheles gambiae, Aedes aegypti, and six Drosophila species). We identified candidates for all nine spliceosomal snRNA genes in all the analyzed genomes. All the species contain a similar number of snRNA genes, with the exception of A. aegypti, whose genome contains more U1, U2, and U5 genes, and A. mellifera, whose genome contains fewer U2 and U5 genes. We found that snRNA genes are generally more closely related to homologs within the same genus than to those in other genera. Promoter regions for all spliceosomal snRNA genes within each insect species share similar sequence motifs that are likely to correspond to the PSEA (proximal sequence element A), the binding site for snRNA activating protein complex, but these promoter elements vary in sequence among the five insect families surveyed here. In contrast to the other insect species investigated, Dipteran genomes are characterized by a rapid evolution (or loss) of components of the U12 spliceosome and a striking loss of U12-type introns.

Animals↗

Insect storage proteins: gene families and receptors.

The accumulation and utilization of storage proteins are prominent events linked to the metamorphosis of holometabolous insects. Storage proteins are synthesized in fat body, secreted into the larval hemolymph and taken up by fat body shortly before pupation. Within the pupal fat body, these proteins are initially stored in protein granules, and later proteolytically broken down to supply amino acid resources necessary for the completion of adult development. Most, but not all storage proteins belong to a superfamily of hexameric larval serum proteins that are evolutionarily related to hemocyanin. This article reviews the classification of these proteins, based on their amino acid sequences, and the current knowledge of the receptors that mediate their selective uptake into pupal fat body.

Animals↗

Ultrabithorax protein is necessary but not sufficient for full activation of decapentaplegic expression in the visceral mesoderm.

To elucidate the mechanisms by which homeotic selector (HOM) genes specify the unique features of Drosophila segments, we have analyzed the regulation of decapentaplegic (dpp), a transforming growth factor (TGF)-beta superfamily member, and have found that the Ultrabithorax (Ubx) HOM protein directly activates dpp expression in parasegment 7 (PS7) of the embryonic visceral mesoderm. Other factors are also required, including one that appears to act through homeodomain protein binding sites and may be encoded by extradenticle (exd). The exd protein binds in a highly co-operative manner to regulatory sequences mediating PS7-specific dpp expression, consistent with a genetic requirement for exd function in normal visceral mesoderm expression of dpp. A second mechanism contributing to PS7 expression of dpp appears not to require Ubx protein directly, and involves a general visceral mesoderm enhancer coupled to a spatially specific repression element. Thus, even in an apparently simple case where visceral mesoderm expression of the dpp target gene mirrors that of the Ubx HOM protein, full activation by Ubx protein requires at least one additional factor. In addition, a distinct regulatory mode not directly involving Ubx protein also appears to contribute to PS7-specific expression.

Animals↗