Gustatory avoidance of fatty acids by <em>Aedes aegypti</em> depends on an arthropod-specific TRP channel

Mosquito-disease vectors, such as Aedes aegypti, use their sense of taste before deciding whether to consume a blood meal, or fly away. However, the molecular mechanisms controlling gustatory decisions in mosquitoes are largely unknown. The TRP channel, Painless1 (Pain1), is an intriguing candidate for participating in Ae. aegypti taste since pain1 transcripts are detected in gustatory receptor neurons (GRNs). The Drosophila homolog, painless (pain) is also expressed in GRNs, where it is required for sensing allyl-isothiocyanate. Here, to identify additional gustatory roles for pain homologs, we first focused on Drosophila pain, which is widely expressed in multiple GRNs classes. We demonstrated that pain mutations eliminated gustatory attraction to low fatty acids levels, repulsion to high levels, and fatty acid-induced action potentials. The attraction and repulsion depended on pain expression in different GRN classes. In contrast to Drosophila, when Aedes contacts fatty acids, they induce gustatory rejection only. Aedes pain1 is expressed in taste organs, and is required for gustatory avoidance of fatty acids, and for fatty acid-induced action potentials. Given that Pain homologs are found in insects but not vertebrates, Pain1 represents an intriguing target for developing repellents to diminish biting, and transmission of infectious agents by mosquito disease vectors. Methods Data were generated using established behavioral, electrophysiological, molecular, and imaging approaches in Drosophila melanogaster and Aedes aegypti, as described in detail in the associated article. Proboscis extension response (PER) assays were performed by stimulating the labella or tarsi with sucrose alone or sucrose mixed with fatty acids or other tastants, and scoring feeding responses. Electrophysiological recordings were obtained using extracellular tip-recording techniques from individual labellar taste sensilla following chemical stimulation, and action potential frequencies were quantified over defined analysis windows. Gene expression was assessed using RT-PCR and quantitative real-time PCR (RT-qPCR), with ribosomal protein genes used as internal controls. Confocal microscopy was used to image gustatory receptor neuron populations following immunohistochemistry or reporter expression, and scanning electron microscopy (SEM) was used to characterize labellar sensilla morphology. Full experimental protocols, sample sizes, and statistical methods are provided in the main article Methods and figure legends.