Kunio Isono

Trehalose sensitivity in Drosophila correlates with mutations in and expression of the gustatory receptor gene Gr5a

Drosophila taste gene Tre is located on the distal X chromosome and controls gustatory sensitivity to a subset of sugars [1, 2]. Two adjacent, seven-transmembrane domain genes near the Tre locus are candidate genes for Tre. One (CG3171) encodes a rhodopsin family G protein receptor [3, 4], and the other (Gr5a) is a member of a chemosensory gene family encoding a putative gustatory receptor [5-7]. We carried out molecular analyses of mutations in Tre to elucidate their involvement in the gustatory phenotype. Here, we show that Tre mutations induced by P element-mediated genomic deletions disrupt Gr5a gene organization and the expression of Gr5a mRNA, while disruption of the CG3171 gene or its expression was not always associated with mutations in Tre. In flies with the spontaneous mutation Tre(01), both CG3171 and Gr5a mRNAs are transcribed. Coding sequences of these two candidate genes were compared among various strains. A total of three polymorphic sites leading to amino acid changes in CG3171 were not correlated with the gustatory phenotype. Among four nonsynonymous sites in Gr5a, a single nucleotide polymorphism leading to an Ala218Thr substitution in the predicted second intracellular loop cosegregated with Tre(01). Taken together, the mutation analyses support that Gr5a is allelic to Tre.

Genetic dimorphism in the taste sensitivity to trehalose inDrosophila melanogaster

SummaryA quantitative behavioral assay was developed for the measurement of taste responses to sugars inDrosophila. The amount of the intake of a sugar solution was measured colorimetrically after homogenization of flies which had consumed sugar solutions mixed with a food-dye. A two-choice method was utilized to determine the taste sensitivity to sugars. Two kinds of sugar solutions were marked with either blue or red food-dye and placed alternately in the wells of a micro test plate. Flies were allowed to choose between the two sugar solutions. By classifying and counting the coloured flies, the relative taste sensitivity could be determined. Employing these methods, a genetic dimorphism in the taste sensitivity to trehalose was found among some laboratory strains ofDrosophila melanogaster. No difference in the taste sensitivity to glucose, fructose and sucrose was found between the trehalose high-sensitivity (T-1) and the low-sensitivity (Oregon-R) strains. Trehalose concentration equivalent to 2 mmol/1 sucrose, in terms of stimulating activity, was 57 mmol/1 inOregon-R and was 10 mmol/1 inT-1. Genetic analysis showed that theTre gene, whose locus is closely linked tocx (13.6) on theX chromosome, is responsible for the difference in the taste sensitivity to trehalose.

Depth Perception from Image Defocus in a Jumping Spider

A Good Judge of Distance Jumping spiders actively pursue their prey, often jumping relatively long distances in order to catch them. Such feats require accurate depth perception. Nagata et al. (p. 469; see the Perspective by Herberstein and Kemp) show that jumping spiders use a process called image defocus, which allows depth perception to be obtained through the comparison of a nonfocused image to a focused image within the same eye. A single layer within the spiders eye that could not focus green light nevertheless contained a green sensitive pigment. Thus, this layer always receives an unfocused image, while other layers receive images in focus. Confirming this eye arrangements role in depth perception, spiders unlucky enough to be bathed in green light nearly always jumped short of their target. To jump exact distances to capture prey, spiders, like computers, use defocus as a major cue for depth perception. The principal eyes of jumping spiders have a unique retina with four tiered photoreceptor layers, on each of which light of different wavelengths is focused by a lens with appreciable chromatic aberration. We found that all photoreceptors in both the deepest and second-deepest layers contain a green-sensitive visual pigment, although green light is only focused on the deepest layer. This mismatch indicates that the second-deepest layer always receives defocused images, which contain depth information of the scene in optical theory. Behavioral experiments revealed that depth perception in the spider was affected by the wavelength of the illuminating light, which affects the amount of defocus in the images resulting from chromatic aberration. Therefore, we propose a depth perception mechanism based on how much the retinal image is defocused.

Molecular and cellular designs of insect taste receptor system

The insect gustatory receptors (GRs) are members of a large G-protein coupled receptor family distantly related to the insect olfactory receptors. They are phylogenetically different from taste receptors of most other animals. GRs are often coexpressed with other GRs in single receptor neurons. Taste receptors other than GRs are also expressed in some neurons. Recent molecular studies in the fruitfly Drosophila revealed that the insect taste receptor system not only covers a wide ligand spectrum of sugars, bitter substances or salts that are common to mammals but also includes reception of pheromone and somatosensory stimulants. However, the central mechanism to perceive and discriminate taste information is not yet elucidated. Analysis of the primary projection of taste neurons to the brain shows that the projection profiles depend basically on the peripheral locations of the neurons as well as the GRs that they express. These results suggest that both peripheral and central design principles of insect taste perception are different from those of olfactory perception.

A Putative Blue-Light Receptor From Drosophila melanogaster

Abstract— A gene encoding a 62.5 kDa homolog of Drosophila melanogaster photolyase was isolated. Purified recombinant protein contained a flavin adenine dinucleotide chromophore. The recombinant protein did not show photolyase activity for either cyclobutane pyrimidine dimers or 6–4 photoproducts in vitro as well as in vivo in Escherichia coli host cells, suggesting that the protein is not a DNA repair enzyme but a blue‐light photoreceptor. Reverse transcription polymerase chain reaction analysis showed that the gene is more expressed in head than in body and that it is more expressed in antennae than in legs, wings and mouth appendages. In a phylogenetic tree of the photolyase family, the Drosophila photolyase homolog is located in a cluster containing 6–4 photolyases and mammalian photolyase homologs, which is only distantly related to the clade of higher plant blue‐light photoreceptors. The mammalian photolyase homologs are more closely related to Drosophila 6–4 photolyase than to the Drosophila photolyase homolog, suggesting different roles of the photolyase homologs.

The specific receptor site for aliphatic carboxylate anion in the labellar sugar receptor of the fleshfly

Abstract A two minute treatment of a single sugar receptor cell with 10 mg/ml pronase did not affect its response to d -fructose, but depressed markedly its response to l -valine. This is the first direct evidence for a specific site for certain aliphatic amino acids. All six amino acids that can stimulate the sugar receptor were examined and classified into two groups according to the presence or absence of the inhibitory effects of pronase treatment. Responses to certain aliphatic amino acids and a corresponding fatty acid were depressed whereas responses to phenylalanine and trytophan were not. Further evidence for the existence of two classes of amino acids comes from the fact that the α amino group of valine is not essential whereas that of phenylalanine is. It was concluded that the first class of amino acids react with a specific receptive site for carboxylate anions whereas the second react with the furanose site.

3‐H‐YDROXYRETINAL AS A CHROMOPHORE OF Drosophila melanogaster VISUAL PIGMENT ANALYZED BY HIGH‐PRESSURE LIQUID CHROMATOGRAPHY

Abstract— The chromophore of the visual pigment of the fruit fly Drosophila melanogaster was studied by high‐pressure liquid chromatography (HPLC). Oxime derivatives of the chromophore were successfully analyzed with a more polar solvent than has been used for retinaloximes. A preceding monochromatic irradiation resulted in reversible interchanges in the amount among geometric isomers of the oximes that reflected the photointerconversion of rhodopsin and metarhodopsin of the peripheral photoreceptors (Rl‐R6). Authentic all‐trans‐3‐h‐ydroxyretinal was used to identify the chromophore from metarhodopsin. Results demonstrated that the chromophore of the visual pigment in Drosophila is 3‐h‐ydroxyretinal as was proposed in larger flies.

TARGETED EXPRESSION OF IP3 SPONGE AND IP3 DSRNA IMPAIRES SUGAR TASTE SENSATION IN DROSOPHILA

We evaluated the role of IP3 in sugar taste reception in Drosophila melanogaster by inactivating the IP3 signaling using genetic tools. We used the “IP3 sponge,” composed of the modified ligand-binding domain from the mouse IP3 receptor, which was designed to absorb IP3 in competition with native IP3 receptors. Another tool was a transgene that generates double-stranded RNA against IP3 receptor mRNA. Both inhibitors diminished the sensitivity of flies to trehalose and sucrose, as estimated by behavioral assays and electrophysiological recordings from the sugar receptor cells. The result indicates that IP3 signaling is indispensable for sugar reception in Drosophila.

A putative binding protein for lipophilic substances related to butterfly oviposition

A unique protein of 23 kDa (Jf23) was found in the tarsus of the female swallowtail butterfly, Atrophaneura alcinous. Jf23 has 38% identity with a bilin‐binding protein, which was found in the cabbage butterfly, Pieris brassicae, and which has two consensus sequences in common with the members of the lipocalin family, suggesting that it is a binding protein for lipophilic ligands. Western blot analysis showed that Jf23 was expressed only in the female, and not in the male. Electrophysiological response of the female tarsi was stimulated by methanolic extract of their host plant, Dutchmans pipe (Aristolochia debilis). The stimulated response was depressed by the presence of Jf23 antiserum. These results suggest that Jf23 is one of the chemosensory signaling proteins, which plays one or more roles in female butterfly oviposition.

Cyclic AMP-dependent memory mutants are defective in the food choice behavior of Drosophila

Acute choice behavior in ingesting two different concentrations of sucrose in Drosophila is presumed to include learning and memory. Effects on this behavior were examined for four mutations that block associative learning (dunce, rutabaga, amnesiac, and radish). Three of these mutations cause cyclic AMP signaling defects and significantly reduced taste discrimination. The exception was radish, which affects neither. Electrophysiological recordings confirmed that the sensitivity of taste receptors is almost indistinguishable in all flies, whether wild type or mutant. These results suggest that food choice behavior in Drosophila involves central nervous learning and memory operating via cyclic AMP signaling pathways.