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Fig. 6. Rh1 and Rh4 may function upstream of TRPA1 to amplify chemical signals. The norpA locus encodes phospholipase Cβ. The flies were defective in avoiding aristolochic acid after knocking down rh1 (UAS-rh1-RNAi) in trpA1-expressing cells (trpA1-GAL4), using the GAL4/UAS system.
The goal of my study is to test the following model: can rhodopsins function as chemosensors? This is an iconoclastic question since rhodopsins were originally discovered more than 100 years ago, and until recently, were thought to function exclusively in light sensation. To explore a non-canonical role for rhodosins in taste, I am taking advantage of the fruit fly, Drosophila melanogaster, as a model organism. There are six characterized rhodopsins in flies (Rh1 through Rh6), all of which are light sensors1. However, rhodopsins across the animal kingdom are found in extra-retinal tissues. However, the functions of these rhodopsins are enigmatic2,3,4. Our lab discovered recently that the major Drosophila rhodopsin, Rh1, is essential for thermal discrimination5, suggesting that rhodopsins have additional sensory roles. In this study, I provide evidence that two rhodopsins (Rh1 and Rh4) are necessary to facilitate gustatory sensing in flies. I am using a combination of genetic, molecular, behavioral and electrophysiological approaches to provide the first demonstration that rhodopsins function in chemosensation.
Fig. 1. Flies lacking Rh1 (NINAE) or Rh4 failed to avoid aristolochic acid. Two-way choice assay (shown in upper left) using 1 mm sucrose alone (mixed with red food dye) versus 5 mM sucrose plus 10 mM aristolochic acid (mixed with blue food dye). Using a blind strategy, we scored the flies with red, blue and purple abdomens, and calculated the preference index. PI = 1 indicates a complete preference for 1 mM sucrose, while PI = 0 shows complete preference for 5 mM sucrose. PI=0.5 indicates no preference. Error bars indicate S.E.M.s. **p<0.01 using the Mann Whitney U test. n=10.
Fig. 3. Neuronal firing frequency of S6 sensilla in response to 1 mM aristolochic acid. (A) Average number of action potentials over the first 500 ms following contact with aristolochic acid. (B) Representative traces. Error bars indicate S.E.M.s. **p<0.01 using the Mann Whitney U test. n≥12
Fig. 2. Flies lacking Rh1 or Rh4 showed normal avoidance to caffeine. Two-way choice assay using 1 mM sucrose alone versus 5 mM sucrose plus 10 mM caffeine. Error bars indicate S.E.M.s. **p<0.01 using the Mann Whitney U test, n=10.
Fig. 4. RT-PCR amplification of rh1 and rh4 RNA from labella.
Fig. 5. Rh1 was not required for sensing higher concentration of aristolochic acid. Average number of action potentials over the first 500 ms following contact with 5 mM aristolochic acid.
1. Rh1 and Rh4 are required in labellar Gustatory Receptor Neurons (GRNs) to sense aristolochic acid.
2. Rh1 is required to mediate responses to low concentration of aristolochic acid by coupling to Gαq and PLCβ.
Functional Characterization of Rhodopsins in Gustatory Signalingof Drosophila Melanogaster
Chao Liu, Adishthi S. Gurav, Letitia A. Mueller, Craig Montell
Craig Montell, Ph.D. Duggan Professor of Neuroscience Department of Molecular, Cellular, & Developmental Biology2109 Life Sciences BuildingEmail: [email protected]: https://labs.mcdb.ucsb.edu/montell/craig/Phone: 805-893-3634
Contact1. Craig Montell. Drosophila visual transduction. Cell Review, 2012. 35(6).2. Mano, H., D. Kojima, and Y. Fukada, Exo-rhodopsin: a novel rhodopsin expressed in the zebrafish pineal gland. Brain Res Mol Brain Res, 1999. 73(1-2).3. Baggiolini, M., et al., Activation of neutrophil leukocytes: chemoattractant receptors and respiratory burst. FASEB J, 1993. 7(11).4. Blackshaw, S. and S.H. Snyder, Encephalopsin: a novel mammalian extraretinal opsin discretely localized in the brain. J Neurosci, 1999. 19(10).5. Wei L. Shen., et al., Function of Rhodopsin in Temperature Discrimination in Drosophila. Science, 2011. 331(1333).
References
Abstract
Results
Chao Liu, Ph.DDepartment of Molecular, Cellular, & Developmental BiologyEmail: [email protected]: 615-818-8947
Pref
eren
ce In
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5 mM sucrose + 10 mM aristolochic acid
1 mM sucrose
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Pref
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5 mM sucrose + 10 mM aristolochic acid
1 mM sucrose
1 mM sucrose
5 mM sucrose + 10 mM caffeine
Summary
PI
(PLCβ)
