GENETIC DISSECTION OF AVERSION TO ALLYL ISOTHIOCYANATE IN Drosophila melanogaster

Many plant species have evolved to produce a variety of compounds that activate the Transient Receptor Potential (TRP) Channels, leading to many animal's behavioral aversion to the chemicals the plants produce. The TRPA subfamily, which in mammals has one member (TRPA1) responds to many electrophiles, including allyl isothiocyanate (AITC). Drosophila have four homologs to the mammalian TRPA1: dTRPA1, painless, pyrexia, and water witch. dTRPA1 and painless are thought to play a role in the behavioral aversion to AITC, however the role of painless is controversial in the published literature. We have analyzed the behavioral phenotypes of painless, dTRPA1, and the double mutant using four behavioral assays: the proboscis extension reflex (PER), two-choice capillary feeding assay (CAFE), oviposition assay, and the locomotion assay. Results from studies using these assays all demonstrated the necessity of both receptors in the behavioral aversion to AITC. Expression patterns of dTRPA1 and painless were evaluated in the periphery to determine if there was any coexpression between these two receptors. We observed an absence of colocalization in sensory cells in the legs. We also expressed UAS-ORK, which electrically silences cells, in sensory neurons, to evaluate the necessity of dTRPA1 and painless cell populations. Using all four behavioral assays, we determined the outputs from both dTRPA1 and painless cell populations were necessary for aversion to AITC. To assess if either dTRPA1 or painless cell populations were sufficient to elicit behavioral aversion, we expressed mammalian TRPV1 in dTRPA1 and painless cell populations to determine if we could confer aversion to capsaicin. Expression of TRPV1 in either cell population was not sufficient to elicit aversion to capsaicin, demonstrating neither dTRPA1 nor painless cell populations are sufficient to elicit aversion. We used a dTRPA1-RNAi to determine if there was a subpopulation of cells with coexpression of these two receptors that we had not yet identified. With all four behavioral assays, we determined there is a subset of cells with coexpresion that are necessary for aversion to AITC. To assess if painless can act as a direct sensor to AITC, we expressed painless in Gr5A, a sugar sensor, in a painless mutant background and evaluated the behavioral phenotype using the CAFE assay. We saw a dose dependent appetitive response to AITC, suggesting painless acted as a direct sensor to AITC. Collectively, these results imply that each channel is necessary for behavioral aversion and there is a complex circuit involved in the chemical detection of AITC.

Grantor dc:publisher

Wake Forest University

Year dc:date.issued

2014

Author dc:creator

• Mandel, Samantha Jill

• Drosophila

Language dc:language.iso

en

Handle dc:identifier.uri

http://hdl.handle.net/10339/39397

OAI identifier oai:identifier

oai:wakespace.lib.wfu.edu:10339/39397