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Abstract
Insect odorant receptors (ORs) detect volatile chemical cues with high sensitivity. These ORs operate as ligand-gated ion channels and are formed by heptahelical OrX and Orco (co-receptor) proteins. A highly conserved calmodulin (CaM) binding site (CBS) 336SAIKYWVER344 within the second intracellular loop of Drosophila melanogaster Orco constitutes a target for regulating OR performance. Here we asked how a point mutation K339N in this CBS affects the olfactory performance of Drosophila melanogaster. We first asked how this mutation would affect the odor responses of olfactory sensory neurons (OSNs). Using Ca2+ imaging in an ex-vivo antenna preparation, we activated all OR (OrX/Orco) expressing neurons using the synthetic agonist VUAA1. In a next attempt, we restricted the OR spectrum to Or22a expressing neurons (Or22a/Orco) and stimulated these OSNs with the ligand ethyl hexanoate. In both approaches, we found that flies carrying the K339N point mutation in Orco display a reduced olfactory response. We also found that the mutation abolishes the capability of OSNs to sensitize by repeated weak odor stimuli. Next, we asked whether OrcoK339N might affect the odor localization performance. Using a wind tunnel bioassay, we found that odor localization in flies carrying the OrcoK339N mutation was severely diminished.
Highlights
Insect odorant receptors (ORs) detect volatile chemical cues with high sensitivity
We checked by immunohistochemistry whether the mutation affects OR trafficking to the olfactory sensory neurons (OSNs) dendrites, and compared the expression of Or22a in mutant flies with UAS-OrcoK339N, as well as in Orco wild type expressing OSNs (OrX/OrcoWT) and in Orco-/- mutant flies rescued with a wild type Orco (OrX/OrcoWT-R) control lines
OR stimulation with the synthetic Orco agonist VUAA1 initiated a C a2+ influx that lead to a rise in [ Ca2+]i visualized as an increase in GCaMP6f fluorescence intensity
Summary
Insect odorant receptors (ORs) detect volatile chemical cues with high sensitivity These ORs operate as ligand-gated ion channels and are formed by heptahelical OrX and Orco (co-receptor) proteins. We restricted the OR spectrum to Or22a expressing neurons (Or22a/Orco) and stimulated these OSNs with the ligand ethyl hexanoate In both approaches, we found that flies carrying the K339N point mutation in Orco display a reduced olfactory response. The sense of smell plays a significant role in detecting and distinguishing between different types of odors present in nature This function is vital for finding mating partners, food sources and oviposition sites, as well as to avoid harmful sites indicated by alarm signals[1,2]. In flies expressing Orco with mutated PKC phosphorylation sites, or in those with pharmacologically downregulated PKC or AC function, no sensitization could be o bserved[24]
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