loco encodes an RGS protein required for Drosophila glial differentiation.

In Drosophila, glial cell development depends on the gene glial cells missing (gcm). gcm activates the expression of other transcription factors such as pointed and repo, which control subsequent glial differentiation. In order to better understand glial cell differentiation, we have screened for genes whose expression in glial cells depends on the activity of pointed.

Identification and characterization of kraken, a gene encoding a putative hydrolytic enzyme in Drosophila melanogaster.

Kraken, a novel Drosophila gene isolated from a 4-8-h-old Drosophila embryo cDNA library, shows homology to a family of serine hydrolases whose common feature is that they all catalyse breakage of substrates with a carbonyl-containing group. It is a single-copy gene with at least two introns and maps to position 21D on polytene chromosomes. kraken is a member of a conserved gene family.

Identification and characterization of the gene for Drosophila L3 ribosomal protein.

A cDNA clone that encodes a Drosophila homologue of ribosomal protein L3 was isolated from a Drosophila ovary gridded cDNA library. The Drosophila ribosomal protein L3 gene (RpL3) is highly conserved with ribosomal protein L3 genes in other organisms. It is a single copy gene and maps to position 86D5-10 on polytene chromosomes.

Identification and characterization of the gene for Drosophila S20 ribosomal protein.

A cDNA clone that encodes a Drosophila homologue of ribosomal protein S20 was isolated from a Drosophila ovary cDNA library. The Drosophila S20 gene (RpS20) is highly conserved with S20 genes in other organisms. It is a single copy gene and maps to position 92F-93A on polytene chromosomes.

Genetic feminization of pheromones and its behavioral consequences in Drosophila males.

Pheromones are intraspecific chemical signals important for mate attraction and discrimination. In the fruit fly Drosophila melanogaster, hydrocarbons on the cuticular surface of the animal are sexually dimorphic in both their occurrence and their effects: Female-specific molecules stimulate male sexual excitation, whereas the predominant male-specific molecule tends to inhibit male excitation. Complete feminization of the pheromone mixture produced by males was induced by targeted expression of the transformer gene in adult oenocytes (subcuticular abdominal cells) or by ubiquitous expression during early imaginal life.

Associative learning disrupted by impaired Gs signaling in Drosophila mushroom bodies.

Disruptions in mushroom body (MB) or central complex (CC) brain structures impair Drosophila associative olfactory learning. Perturbations in adenosine 3',5' monophosphate signaling also disrupt learning. To integrate these observations, expression of a constitutively activated stimulatory heterotrimeric guanosine triphosphate-binding protein alpha subunit (Galphas*) was targeted to these brain structures.

Activation of protein kinase A-independent pathways by Gs alpha in Drosophila.

One of the best-described transmembrane signal transduction mechanisms is based on receptor activation of the alpha subunit of the heterotrimeric G protein Gs, leading to stimulation of adenylyl cyclase and the production of cAMP. Intracellular cAMP is then thought to mediate its effects largely, if not entirely, by activation of protein kinase A and the subsequent phosphorylation of substrates which in turn control diverse cellular phenomena. In this report we demonstrate, by two different methods, that reduction or elimination of protein kinase A activity had no effect on phenotypes generated by activation of Gs alpha pathways in Drosophila wing epithelial cells.

Inducible ternary control of transgene expression and cell ablation in Drosophila.

In Drosophila, P-GAL4 enhancer trap lines can target expression of a cloned gene, under control of a UASGAL element, to any cells of interest. However, additional expression of GAL4 in other cells can produce unwanted lethality or side-effects, particularly when it drives expression of a toxic gene product. To target the toxic gene product ricin A chain specifically to adult neurons, we have superimposed a second layer of regulation on the GAL4 control.

Mutations in shaking-B prevent electrical synapse formation in the Drosophila giant fiber system.

The giant fiber system (GFS) is a simple network of neurons that mediates visually elicited escape behavior in Drosophila. The giant fiber (GF), the major component of the system, is a large, descending interneuron that relays visual stimuli to the motoneurons that innervate the tergotrochanteral jump muscle (TTM) and dorsal longitudinal flight muscles (DLMs). Mutations in the neural transcript from the shaking-B locus abolish the behavioral response by disrupting transmission at some electrical synapses in the GFS.

Syntaxin and synaptobrevin function downstream of vesicle docking in Drosophila.

In synaptic transmission, vesicles are proposed to dock at presynaptic active zones by the association of synaptobrevin (v-SNARE) with syntaxin (t-SNARE). We test this hypothesis in Drosophila strains lacking neural synaptobrevin (n-synaptobrevin) or syntaxin. We showed previously that loss of either protein completely blocks synaptic transmission.

Targeted expression of tetanus toxin light chain in Drosophila specifically eliminates synaptic transmission and causes behavioral defects.

Tetanus toxin cleaves the synaptic vesicle protein synaptobrevin, and the ensuing loss of neurotransmitter exocytosis has implicated synaptobrevin in this process. To further the study of synaptic function in a genetically tractable organism and to generate a tool to disable neuronal communication for behavioural studies, we have expressed a gene encoding tetanus toxin light chain in Drosophila. Toxin expression in embryonic neurons removes detectable synaptobrevin and eliminates evoked, but not spontaneous, synaptic vesicle release.

Microbial growth on the surfaces of various orthodontic bonding cements.

An experimental technique was developed to examine the adhesion of a strain of Streptococcus mutans to the surface of discs of selected orthodontic bonding composites and a glass ionomer cement. Colonization by the bacterium was assessed by weight gain and scanning electron microscopy. Initial colonization occurred after about 3 days, and the specimens were usually completely covered by the fourth day.

Surface roughness and droplet contact angle measurement of various orthodontic bonding cements.

Surface characteristics that are considered important for bacterial attachment to thirteen orthodontic bonding composite cements and one glass ionomer cement were examined in vitro before and after toothbrush abrasion. The surface roughness and contact angle measurements were found to be statistically significantly different between the materials, both before and after brushing, and there were also statistically significant changes within materials after brushing. There were low correlation coefficients between surface roughness and contact angle for both pre-and post-brushed materials.

Selective cell ablation and genetic surgery.

Selective ablation is a useful tool to investigate the origin, fate or function of particular cells. It can be achieved either using physical methods or toxigenic methods. Recent successes with conditional ablation should make it easier to ablate a wider range of cells than has hitherto been possible.

Inducible cell ablation in Drosophila by cold-sensitive ricin A chain.

We have developed a system for temperature-inducible killing of specific cells in the fruitfly Drosophila melanogaster. The system overcomes many of the limitations of existing cell ablation methods and is in principle applicable to any non-homeothermic eukaryote. Temperature-sensitive and cold-sensitive mutations in the ricin toxin A chain (RTA) of castor bean were generated in yeast.

Use of a cytoplasmically localised P-lacZ fusion to identify cell shapes by enhancer trapping inDrosophila.

The N-terminal 125 amino acids of theDrosophila P element transposase are necessary and sufficient for the nuclear localisation of a hybridlacZ gene product in most cell types of theDrosophila embryo. A P-lacZ enhancer-trap element lacking these residues is of use in visualizing the shapes of P-lacZ-expressing cells.

P-element-mediated enhancer detection allows rapid identification of developmentally regulated genes and cell specific markers in Drosophila.

We have employed a new technique in Drosophila that allows in vivo detection of genomic regulatory elements using a beta-galactosidase reporter gene. A translational fusion of the reporter gene to the P-transposase gene, which is encoded by the P-transposon of Drosophila, places the expression of beta-galactosidase under the control of the weak P-transposase promoter. Flies carrying single insertions of this P-element construct at different locations in the Drosophila genome frequently stain for beta-galactosidase activity in a temporally and spatially restricted fashion in embryos, larvae and adult ovaries, reflecting the influence of nearby genomic regulatory elements on the P-transposase promoter.

P-element-mediated enhancer detection: an efficient method for isolating and characterizing developmentally regulated genes in Drosophila.

We describe a new approach for identifying and studying genes involved in Drosophila development. Single copies of an enhancer detector transposon, P[1ArB], have been introduced into flies at many different genomic locations. The beta-galactosidase reporter gene in this construct is influenced by a wide range of genomic transcriptional regulatory elements in its vicinity.

P-element-mediated enhancer detection: a versatile method to study development in Drosophila.

We generated and characterized greater than 500 Drosophila strains that carry single copies of a novel P-element enhancer detector. In the majority of the strains, the beta-galactosidase reporter gene in the P-transposon responds to nearby transcriptional regulatory sequences in the genome. A remarkable diversity of spatially and temporally regulated staining patterns is observed in embryos carrying different insertions.

Neural enhancer-like elements as specific cell markers in Drosophila.

We have analysed four strains of Drosophila melanogaster which each carry the transposon P[lac,ry+] at a unique genomic location. In one of the strains, P[lac,ry+]A37, all the peripheral neurones that we can identify express the P-lac fusion protein; in at least some cases, and the support cells associated to particular neurones are also labelled. Expression of the fusion protein can be detected in subepidermal cells of the body segments as early as 4-5 h of development, according to a precise and reproducible pattern.

Detection in situ of genomic regulatory elements in Drosophila.

We have developed an approach for the in situ detection of genomic elements that regulate transcription zin Drosophila melanogaster. The approach is analogous to a powerful method of bacterial genetics, the random generation of operon fusions, that enables the isolation and characterization of genes simply by knowing or postulating their pattern of expression; it is not necessary initially to screen for mutant phenotypes. To apply this approach to Drosophila, we have used the expression of the lacZ gene of Escherichia coli from the P-element promoter in germ-line transformant flies to screen for chromosomal elements that can act at a distance to stimulate expression from this apparently weak promoter.

Integrable alpha-amylase plasmid for generating random transcriptional fusions in Bacillus subtilis.

An integrable plasmid, pOK4, which replicated independently in Escherichia coli was constructed for generating transcriptional fusions in vivo in Bacillus DNA. It did not replicate independently in Bacillus subtilis, but it could be made to integrate into the chromosome of B. subtilis if sequences homologous to chromosomal sequences were inserted into it.