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At least 577 records · Page 32Linked to original sources
PubMed2005

The Drosophila gene CG9918 codes for a pyrokinin-1 receptor.

The database from the Drosophila Genome Project contains a gene, CG9918, annotated to code for a G protein-coupled receptor. We cloned the cDNA of this gene and functionally expressed it in Chinese hamster ovary cells. We tested a library of about 25 Drosophila and other insect neuropeptides, and seven insect biogenic amines on the expressed receptor and found that it was activated by low concentrations of the Drosophila neuropeptide, pyrokinin-1 (TGPSASSGLWFGPRLamide; EC50, 5 x 10(-8) M). The receptor was also activated by other Drosophila neuropeptides, terminating with the sequence PRLamide (Hug-gamma, ecdysis-triggering-hormone-1, pyrokinin-2), but in these cases about six to eight times higher concentrations were needed. The receptor was not activated by Drosophila neuropeptides, containing a C-terminal PRIamide sequence (such as ecdysis-triggering-hormone-2), or PRVamide (such as capa-1 and -2), or other neuropeptides and biogenic amines not related to the pyrokinins. This paper is the first conclusive report that CG9918 is a Drosophila pyrokinin-1 receptor gene.

Amino Acid Sequence↗
PubMed2003

Drosophila proteasome regulator REGgamma: transcriptional activation by DNA replication-related factor DREF and evidence for a role in cell cycle progression.

The proteasome regulator REG (PA28gamma) is a conserved complex present in metazoan nuclei and is able to stimulate the trypsin-like activity of the proteasome in a non-ATP dependent manner. However, the in vivo function for REGgamma in metazoan cells is currently unknown. To understand the role of Drosophila REGgamma we have attempted to identify the type of promoter elements regulating its transcription. Mapping the site of the transcription initiation revealed a TATA-less promoter, and a sequence search identified elements found typically in Drosophila genes involved in cell cycle progression and DNA replication. In order to test the relevance of the motifs, REGgamma transcriptional assays were carried out with mutations in the proposed promoter. Our results indicate that a single Drosophila replication-related element sequence, DRE, is essential for REGgamma transcription. To confirm that REGgamma has a role in cell cycle progression, the effect of removing REGgamma from S2 cells was tested using RNA interference. Drosophila cells depleted of REGgamma showed partial arrests in G1/S cell cycle transition. Immuno-staining of Drosophila embryos revealed that REGgamma is typically localized to the nucleus during embryogenesis with increased levels present in invaginating cells during gastrulation. The REGgamma was found dispersed throughout the cell volume within mitotic domains undergoing cell division. Finally, database searches suggest that the DRE system may regulate key members of the proteasome system in Drosophila.

Animals↗
PubMed2002

Structure and expression patterns of Drosophila TULP and TUSP, members of the tubby-like gene family.

Tubby is a mouse gene that may provide a model for adult-onset obesity in humans. It is a member of a four gene family in mammals that collectively encode the Tubby-like proteins (TULPs), putative transcription factors which share similar 260 amino acid 'tubby domains' at their C-termini. The mammalian genome also encodes distant relatives of TULPs, which have been called TUSPs (tubby domain superfamily proteins). We have characterized the transcription unit of the single Drosophila TULP homolog, analyzed the expression pattern of the Drosophila TULP and TUSP genes, and determined the evolutionary relationships between the Drosophila proteins and members of the tubby domain superfamily in other organisms. Interestingly, like its mammalian homologs, Drosophila TULP is principally expressed in the embryonic central and peripheral nervous systems. This suggests that mammalian and Drosophila TULPs may possess some conserved functional properties in the nervous system. The Drosophila TUSP gene is also expressed in the central nervous system and olfactory organ but in few other peripheral sensory organs.

Adaptor Proteins, Signal Transducing↗
PubMed1999

The LIM homeodomain protein dLim1 defines a subclass of neurons within the embryonic ventral nerve cord of Drosophila.

Members of the LIM homeodomain family of transcription factors have been implicated in specifying cell identity in a range of species. In Drosophila three LIM homeobox genes, apterous, lim3 and isl, have been shown to control axon pathfinding of subsets of neurons within the embryo. Here we describe the isolation and characterization of another LIM homeobox gene in Drosophila termed dlim1, a homolog of the vertebrate Lim-1 gene. The sequence and expression of dLim1 is highly related to its vertebrate homologs. Within the Drosophila embryo, dLim1 is expressed in the head primordia, the brain lobes, and in distinct sets of motorneurons and interneurons within the ventral nerve cord. Comparatively in vertebrates, Lim-1 (Lhx1) along with Lim-3 (Lhx3), Gsh-4 (Lhx4), Isl-1 and Isl-2 are expressed in developing motorneurons along the spinal column, where their overlapping expression suggests a role for these genes in the establishment of specific motorneuron subtypes. dLim1 is absent from all cells expressing Isl, Lim3, and Apterous, indicating that these proteins function independently within the Drosophila embryo. To investigate the function of dlim1, we generated loss-of-function mutations within the locus. Our findings show that dlim1 is an essential gene that when mutated results in lethality near the larval-pupal boundary. In contrast to vertebrate Lim-1, dlim1 has no apparent role in anterior patterning of the Drosophila embryo. Our analysis shows that dlim1 has been evolutionarily conserved, however the Drosophila lim1 gene exhibits unique properties that distinguishes it from its vertebrate homologs.

Amino Acid Motifs↗
PubMed2000

Drosophila models of human neurodegenerative disease.

Drosophila has provided a powerful genetic system in which to elucidate fundamental cellular pathways in the context of a developing and functioning nervous system. Recently, Drosophila has been applied toward elucidating mechanisms of human neurodegenerative disease, including Alzheimer's, Parkinson's and Huntington's diseases. Drosophila allows study of the normal function of disease proteins, as well as study of effects of familial mutations upon targeted expression of human mutant forms in the fly. These studies have revealed new insight into the normal functions of such disease proteins, as well as provided models in Drosophila that will allow genetic approaches to be applied toward elucidating ways to prevent or delay toxic effects of such disease proteins. These, and studies to come that follow from the recently completed sequence of the Drosophila genome, underscore the contributions that Drosophila as a model genetic system stands to contribute toward the understanding of human neurodegenerative disease.

Amyloid beta-Protein Precursor↗
PubMed2004

Expression of leukemic MLL fusion proteins in Drosophila affects cell cycle control and chromosome morphology.

The Mixed Lineage Leukemia (MLL) gene is involved in lymphoblastic and myeloid leukemia through chromosome translocations leading to fusion of MLL to partner genes, or through internal MLL rearrangements. MLL is the mammalian counterpart of the Drosophila trithorax (trx) gene, involved in maintaining active gene expression states. We have used transgenic Drosophila to assess the molecular targets and cellular processes affected by MLL and two of its leukemic fusion proteins. We find that whereas expression of normal human MLL in flies does not result in phenotypic alterations, overexpressing the human MLL-AF9 and MLL-AF4 proteins causes larval to pupal lethality, which interestingly resembles the phenotypes displayed by certain Drosophila trx mutant alleles. MLL-AF9 and MLL-AF4 transgenic flies exhibit antagonistic alterations in cell cycle progression. Additionally, flies expressing MLL-AF9 display impairment in higher order chromatin integrity, evidenced in decondensation of mitotic figures. The effects of MLL fusion proteins in Drosophila suggest that alteration of chromatin structure by MLL fusion proteins may contribute to the lethal phenotype. Our results indicate that the mode(s) of action of MLL-AF9 in Drosophila varies from that of MLL-AF4. Taken together, the expression of MLL fusion proteins in Drosophila provides a new and powerful system to reveal and characterize biological activities associated with MLL fusion proteins.

Animals↗
PubMed2002

Identification of Drosophila melanogaster yellow-f and yellow-f2 proteins as dopachrome-conversion enzymes.

This study describes the identification of Drosophila yellow-f and yellow-f2 as dopachrome-conversion enzymes responsible for catalysing the conversion of dopachrome into 5,6-dihydroxyindole in the melanization pathway. Drosophila yellow -y gene and yellow -b, -c, -f and -f2 genes were expressed in an insect cell/baculovirus expression system and their corresponding recombinant proteins were screened for dopachrome-conversion enzyme activity. Among the yellow and yellow -related genes, the yellow -f and yellow -f2 genes were identified as the genes coding for Drosophila dopachrome-conversion enzyme based on the high activity of their recombinant proteins in catalysing the production of 5,6-dihydroxyindole from dopachrome. Both yellow-f and yellow-f2 are capable of mediating a decarboxylative structural rearrangement of dopachrome, as well as an isomerization/tautomerization of dopamine chrome and dopa methyl ester chrome. Northern hybridization revealed the transcription of yellow -f in larvae and pupae, but a high abundance of mRNA was observed in later larval and early pupal stages. In contrast, yellow-f2 transcripts were present at all stages, but high abundance of its mRNA was observed in later-stage pupae and adults. These data indicate that yellow-f and yellow-f2 complement each other during Drosophila development and that the yellow-f is involved in larval and pupal melanization, and yellow-f2 plays a major role in melanization reactions in Drosophila during later pupal and adult development. Results from this study provide the groundwork towards a better understanding of the physiological roles of the Drosophila yellow gene family.

Amino Acid Sequence↗
PubMed2006

Analysis of the hypoxia-sensing pathway in Drosophila melanogaster.

The mechanism by which hypoxia induces gene transcription involves the inhibition of HIF-1alpha (hypoxia-inducible factor-1 alpha subunit) PHD (prolyl hydroxylase) activity, which prevents the VHL (von Hippel-Lindau)-dependent targeting of HIF-1alpha to the ubiquitin/proteasome pathway. HIF-1alpha thus accumulates and promotes gene transcription. In the present study, first we provide direct biochemical evidence for the presence of a conserved hypoxic signalling pathway in Drosophila melanogaster. An assay for 2-oxoglutarate-dependent dioxygenases was developed using Drosophila embryonic and larval homogenates as a source of enzyme. Drosophila PHD has a low substrate specificity and hydroxylates key proline residues in the ODD (oxygen-dependent degradation) domains of human HIF-1alpha and Similar, the Drosophila homologue of HIF-1alpha. The enzyme promotes human and Drosophila [(35)S]VHL binding to GST (glutathione S-transferase)-ODD-domain fusion protein. Hydroxylation is enhanced by proteasomal inhibitors and was ascertained using an anti-hydroxyproline antibody. Secondly, by using transgenic flies expressing a fusion protein that combined an ODD domain and the green fluorescent protein (ODD-GFP), we analysed the hypoxic cascade in different embryonic and larval tissues. Hypoxic accumulation of the reporter protein was observed in the whole tracheal tree, but not in the ectoderm. Hypoxic stabilization of ODD-GFP in the ectoderm was restored by inducing VHL expression in these cells. These results show that Drosophila tissues exhibit different sensitivities to hypoxia.

Amino Acid Sequence↗
PubMed2006

Participation of a galactose-specific C-type lectin in Drosophila immunity.

A galactose-specific C-type lectin has been purified from a pupal extract of Drosophila melanogaster. This lectin gene, named DL1 (Drosophila lectin 1), is part of a gene cluster with the other two galactose-specific C-type lectin genes, named DL2 (Drosophila lectin 2) and DL3 (Drosophila lectin 3). These three genes are expressed differentially in fruit fly, but show similar haemagglutinating activities. The present study characterized the biochemical and biological properties of the DL1 protein. The recombinant DL1 protein bound to Escherichia coli and Erwinia chrysanthemi, but not to other Gram-negative or any other kinds of microbial strains that have been investigated. In addition, DL1 agglutinated E. coli and markedly intensified the association of a Drosophila haemocytes-derived cell line with E. coli. For in vivo genetic analysis of the lectin genes, we also established a null-mutant Drosophila. The induction of inducible antibacterial peptide genes was not impaired in the DL1 mutant, suggesting that the galactose-specific C-type lectin does not participate in the induction of antibacterial peptides, but possibly participates in the immune response via the haemocyte-mediated mechanism.

Amino Acid Sequence↗
PubMed2002

Dbeta3, an atypical nicotinic acetylcholine receptor subunit from Drosophila : molecular cloning, heterologous expression and coassembly.

Insect nicotinic acetylcholine receptors (nAChRs) play a central role in mediating neuronal synaptic transmission and are the target sites for the increasingly important group of neonicotinoid insecticides. Six nicotinic acetylcholine receptor (nAChR) subunits (four alpha-type and two beta-type) have been cloned previously from the model insect species Drosophila melanogaster. Despite extensive efforts, it has not been possible to generate functional recombinant nAChRs by heterologous expression of any combination of these six subunits. It has, however, been possible to express functional hybrid receptors when Drosophila alpha subunits are co-expressed with vertebrate beta subunits. This has led to the assumption that successful heterologous expression might require an, as yet, uncloned beta-type insect subunit. Examination of the recently completed Drosophila genomic sequence data has identified a novel putative nAChR beta-type subunit. Here we report the molecular cloning, heterologous expression and characterization of this putative Drosophila nAChR subunit (Dbeta3). Phylogenetic comparisons with other ligand-gated ion channel subunit sequences support its classification as a nAChR subunit but show it to be a distantly related member of this neurotransmitter receptor subunit family. Evidence that the Dbeta3 subunit is able to coassemble with other Drosophila nAChR subunits and contribute to recombinant nAChRs has been obtained by both radioligand binding and coimmunoprecipitation studies in transfected Drosophila S2 cells.

Animals↗
PubMed2006

Anthrax lethal factor and edema factor act on conserved targets in Drosophila.

Many bacterial toxins act on conserved components of essential host-signaling pathways. One consequence of this conservation is that genetic model organisms such as Drosophila melanogaster can be used for analyzing the mechanism of toxin action. In this study, we characterize the activities of two anthrax virulence factors, lethal factor (LF) and edema factor, in transgenic Drosophila. LF is a zinc metalloprotease that cleaves and inactivates most human mitogen-activated protein kinase (MAPK) kinases (MAPKKs). We found that LF similarly cleaves the Drosophila MAPK kinases Hemipterous (Hep) and Licorne in vitro. Consistent with these observations, expression of LF in Drosophila inhibited the Hep/c-Jun N-terminal kinase pathway during embryonic dorsal closure and the related process of adult thoracic closure. Epistasis experiments confirmed that LF acts at the level of Hep. We also found that LF inhibits Ras/MAPK signaling during wing development and that LF acts upstream of MAPK and downstream of Raf, consistent with LF acting at the level of Dsor. In addition, we found that edema factor, a potent adenylate cyclase, inhibits the hh pathway during wing development, consistent with the known role of cAMP-dependent PKA in suppressing the Hedgehog response. These results demonstrate that anthrax toxins function in Drosophila as they do in mammalian cells and open the way to using Drosophila as a multicellular host system for studying the in vivo function of diverse toxins and virulence factors.

Amino Acid Sequence↗
PubMed1997

Activation of distinct caspase-like proteases by Fas and reaper in Drosophila cells.

The cytoplasmic region of Fas, a mammalian death factor receptor, shares a limited homology with reaper, an apoptosis-inducing protein in Drosophila. Expression of either the Fas cytoplasmic region (FasC) or of reaper in Drosophila cells caused cell death. The death process induced by FasC or reaper was inhibited by crmA or p35, suggesting that its death process is mediated by caspase-like proteases. Both Ac-YVAD aldehyde and Ac-DEVD aldehyde, specific inhibitors of caspase 1- and caspase 3-like proteases, respectively, inhibited the FasC-induced death of Drosophila cells. However, the cell death induced by reaper was inhibited by Ac-DEVD aldehyde, but not by Ac-YVAD aldehyde. A caspase 1-like protease activity that preferentially recognizes the YVAD sequence gradually increased in the cytosolic fraction of the FasC-activated cells, whereas the caspase 3-like protease activity recognizing the DEVD sequence was observed in the reaper-activated cells. Partial purification and biochemical characterization of the proteases indicated that there are at least three distinct caspase-like proteases in Drosophila cells, which are differentially activated by FasC and reaper. The conservation of the Fas-death signaling pathway in Drosophila cells, which is distinct from that for reaper, may indicate that cell death in Drosophila is controlled not only by the reaper suicide gene, but also by a Fas-like killer gene.

Animals↗
PubMed1999

Proapoptotic activity of Caenorhabditis elegans CED-4 protein in Drosophila: implicated mechanisms for caspase activation.

CED-4 protein plays an important role in the induction of programmed cell death in Caenorhabditis elegans through the activation of caspases. However, the precise mechanisms by which it activates caspases remain unknown. To investigate the conservation of CED-4 function in evolution, transgenic Drosophila lines that express CED-4 in the compound eye were generated. Ectopic expression of CED-4 in the eyes induced massive apoptotic cell death through caspase activation. An ATP-binding site (P-loop) mutation in CED-4 (K165R) causes a loss of function in its ability to activate Drosophila caspase, and an ATPase inhibitor blocks the CED-4-dependent caspase activity in Drosophila S2 cells. Immunoprecipitation analysis showed that both CED-4 and CED-4 (K165R) bind directly to Drosophila caspase drICE, and the overexpression of CED-4 (K165R) inhibits CED-4-, ecdysone-, or cycloheximide-dependent caspase activation in S2 cells. Furthermore, CED-4 (K165R) partially prevented cell death induced by CED-4 in Drosophila compound eyes. Thus, CED-4 function is evolutionarily conserved in Drosophila, and the molecular mechanisms by which CED-4 activates caspases might require ATP binding and direct interaction with the caspases.

Adenosine Triphosphate↗
PubMed1997

A novel Drosophila receptor tyrosine kinase expressed specifically in the nervous system. Unique structural features and implication in developmental signaling.

We report the identification and characterization of Dnrk (Drosophila neurospecific receptor kinase), a Drosophila gene encoding a putative receptor tyrosine kinase (RTK) highly related to the Trk and Ror families of RTKs. During Drosophila embryogenesis, the Dnrk gene is expressed specifically in the developing nervous system. The Dnrk protein possesses two conserved cysteine-containing domains and a kringle domain within its extracellular domain, resembling those observed in Ror family RTKs (Ror1, Ror2, and a Drosophila Ror, Dror). This protein contains the catalytic tyrosine kinase (TK) domain with two putative ATP-binding motifs, resembling those observed in another Drosophila RTK (Dtrk) that mediates homophilic cell adhesion. The TK domain of Dnrk, expressed in bacteria or mammalian cells, exhibits apparent autophosphorylation activities in vitro. The TK domain lacking the distal ATP-binding motif also exhibits autophosphorylation activity, yet to a lesser extent. In addition to its TK activity, there are several putative tyrosine-containing motifs that upon phosphorylation may interact with Src homology 2 regions of other signaling molecules. Collectively, these results suggest that Dnrk may play an important role in neural development during Drosophila embryogenesis.

Amino Acid Sequence↗
PubMed1999

Preferential expression of biotransformation enzymes in the olfactory organs of Drosophila melanogaster, the antennae.

Biotransformation enzymes have been found in the olfactory epithelium of vertebrates. We now show that in Drosophila melanogaster, a UDP-glycosyltransferase (UGT), as well as a short chain dehydrogenase/reductase and a cytochrome P450 are expressed specifically or preferentially in the olfactory organs, the antennae. The evolutionarily conserved expression of biotransformation enzymes in olfactory organs suggests that they play an important role in olfaction. In addition, we describe five Drosophila UGTs belonging to two families. All five UGTs contain a putative transmembrane domain at their C terminus as is the case for vertebrate UGTs where it is required for enzymatic activity. The primary sequence of the C terminus, including part of the transmembrane domain, differs between the two families but is highly conserved not only within each Drosophila family, but also between the members of one of the Drosophila families and vertebrate UGTs. The partial overlap of the conserved primary sequence with the transmembrane domain suggests that this part of the protein is involved in specific interactions occurring at the membrane surface. The presence of different C termini in the two Drosophila families suggests that they interact with different targets, one of which is conserved between Drosophila and vertebrates.

Amino Acid Sequence↗
PubMed1999

A novel sulfonylurea receptor family member expressed in the embryonic Drosophila dorsal vessel and tracheal system.

Sulfonylurea receptors (SURx) are required subunits of the ATP-sensitive potassium channel. SURx alone is electrophysiologically inert. However, when SURx is combined with an inward rectifier Kir6.2 subunit, ATP-sensitive potassium channel activity is generated. We report the identification, characterization, and localization of Dsur, a novel Drosophila gene that is highly related to the vertebrate SUR family. The Dsur coding sequence contains structural features characteristic of the ABC transporter family and, in addition, harbors 1.7 kilobases of a distinctive sequence that does not share homology with any known gene. When Dsur alone is expressed in Xenopus oocytes glibenclamide-sensitive potassium channel activity occurs. During Drosophila embryogenesis, the Dsur gene is specifically expressed in the developing tracheal system and dorsal vessel. Studies of the Drosophila genome support that only a single Dsur gene is present. Our data reveal conservation of glibenclamide-sensitive potassium channels in Drosophila and suggest that Dsur may play an important role during Drosophila embryogenesis. The lack of gene duplication in the Drosophila system provides a unique opportunity for functional studies of SUR using a genetic approach.

ATP-Binding Cassette Transporters↗
PubMed2002

Down-regulation of DIAP1 triggers a novel Drosophila cell death pathway mediated by Dark and DRONC.

Members of the inhibitor of apoptosis protein (IAP) family can inhibit caspases and cell death in a variety of insect and vertebrate systems. Drosophila IAP1 (DIAP1) inhibits cell death to facilitate normal embryonic development. Here, using RNA interference, we showed that down-regulation of DIAP1 is sufficient to induce cell death in Drosophila S2 cells. Although this cell death process was accompanied by elevated caspase activity, this activation was not essential for cell death. We found that DIAP1 depletion-induced cell death was strongly suppressed by a reduction in the Drosophila caspase DRONC or the Drosophila apoptotic protease-activating factor-1 (Apaf-1) homolog, Dark. RNA interference studies in Drosophila embryos also demonstrated that the action of Dark is epistatic to that of DIAP1 in this cell death pathway. The cell death caused by down-regulation of DIAP1 was accelerated by overexpression of DRONC and Dark, and a caspase-inactive mutant form of DRONC could functionally substitute the wild-type DRONC in accelerating cell death. These results suggest the existence of a novel mechanism for cell death signaling in Drosophila that is mediated by DRONC and Dark.

Animals↗
PubMed2000

Expression and functional characterization of a Drosophila neuropeptide precursor with homology to mammalian preprotachykinin A.

Peptides structurally related to mammalian tachykinins have recently been isolated from the brain and intestine of several insect species, where they are believed to function as both neuromodulators and hormones. Further evidence for the signaling role of insect tachykinin-related peptides was provided by the cloning and characterization of cDNAs for two tachykinin receptors from Drosophila melanogaster. However, no endogenous ligand has been isolated for the Drosophila tachykinin receptors to date. Analysis of the Drosophila genome allowed us to identify a putative tachykinin-related peptide prohormone (prepro-DTK) gene. A 1.5-kilobase pair cDNA amplified from a Drosophila head cDNA library contained an 870-base pair open reading frame, which encodes five novel Drosophila tachykinin-related peptides (called DTK peptides) with conserved C-terminal FXGXR-amide motifs common to other insect tachykinin-related peptides. The tachykinin-related peptide prohormone gene (Dtk) is both expressed and post-translationally processed in larval and adult midgut endocrine cells and in the central nervous system, with midgut expression starting at stage 17 of embryogenesis. The predicted Drosophila tachykinin peptides have potent stimulatory effects on the contractions of insect gut. These data provide additional evidence for the conservation of both the structure and function of the tachykinin peptides in the brain and gut during the course of evolution.

Animals↗