A natural variant and engineered mutation in a GPCR promote DEET resistance in C. elegans

Abstract

DEET (N,N-diethyl-meta-toluamide) is a synthetic chemical identified by the US Department of Agriculture in 1946 in a screen for repellents to protect soldiers from mosquito-borne diseases1,2. Since its discovery, DEET has become the world’s most widely used arthropod repellent and is effective against invertebrates separated by millions of years of evolution—including biting flies3, honeybees4, ticks5, and land leeches3. In insects, DEET acts on the olfactory system5,6,7,8,9,10,11,12 and requires the olfactory receptor co-receptor Orco7,9,10,11,12, but exactly how it works remains controversial13. Here we show that the nematode Caenorhabditis elegans is sensitive to DEET and use this genetically tractable animal to study the mechanism of action of this chemical. We found that DEET is not a volatile repellent, but instead interferes selectively with chemotaxis to a variety of attractant and repellent molecules. In a forward genetic screen for DEET-resistant worms, we identified a gene that encodes a single G protein-coupled receptor, str-217, which is expressed in a single pair of chemosensory neurons that are responsive to DEET, called ADL neurons. Mis-expression of str-217 in another chemosensory neuron conferred responses to DEET. Engineered str-217 mutants, and a wild isolate of C. elegans that carries a str-217 deletion, are resistant to DEET. We found that DEET can interfere with behaviour by inducing an increase in average pause length during locomotion, and show that this increase in pausing requires both str-217 and ADL neurons. Finally, we demonstrated that ADL neurons are activated by DEET and that optogenetic activation of ADL neurons increased average pause length. This is consistent with the ‘confusant’ hypothesis, which proposes that DEET is not a simple repellent but that it instead modulates multiple olfactory pathways to scramble behavioural responses10,11. Our results suggest a consistent motif in the effectiveness of DEET across widely divergent taxa: an effect on multiple chemosensory neurons that disrupts the pairing between odorant stimulus and behavioural response.

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Fig. 1: DEET interferes with chemotaxis in wild-type C. elegans.
Fig. 2: str-217 mutants are resistant to DEET.
Fig. 3: Several C. elegans neurons respond to DEET but ADL neurons are required for sensitivity to DEET.
Fig. 4: str-217 is necessary for ADL responses to DEET and can confer DEET sensitivity to AWB neurons.
Fig. 5: DEET increases average pause length by activating str-217 and ADL neurons.

Data availability

All scripts and graphed data with the exception of raw video files are available in the Supplementary Data. Raw video files are available on request from the corresponding author.

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Acknowledgements

We thank M. Crickmore, K. J. Lee, A. Singhvi, N. Yapici and members of the Vosshall Laboratory for comments on the manuscript, and for experimental assistance and advice: S. Shaham and W. Wang for assistance with chemical mutagenesis; H. Jang for assistance with chemotaxis behaviour and imaging; A. Lopez-Cruz and E. Scheer for assistance with tracking; S. Levy and E. Scheer for plasmids and strains; and A. Nguyen for early analysis of mutants (with P.S.H.). This work was conducted with support from NIH (to E.J.D., F31 DC014222) and the CGC (P40 OD010440), which provided selected strains. L.B.V. is an investigator of the Howard Hughes Medical Institute.

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Nature thanks E. Poivet and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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E.J.D. and L.B.V. conceived the project, wrote the manuscript and produced the figures: E.J.D. performed the experiments and analyses shown in all figures, except for Figs. 3a–e, 4m–o (performed by M.D.) and Fig. 3g (performed by X.J.). L.B.D. performed HEK293T expression. C.I.B. provided guidance, experimental design advice and data interpretation. P.S.H. made the original observation that DEET interferes with chemotaxis, and performed initial genetic screens.

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Correspondence to Leslie B. Vosshall.

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Dennis, E.J., Dobosiewicz, M., Jin, X. et al. A natural variant and engineered mutation in a GPCR promote DEET resistance in C. elegans. Nature 562, 119–123 (2018). https://doi.org/10.1038/s41586-018-0546-8

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Keywords

  • Average Pause Length
  • Chemosensory Neurons
  • Buffer Solvent
  • Wild-type Worms
  • Read Buffer

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