Fig. 3.

Mutation of the Orco CBS disrupts Orco and OR47a trafficking to the ciliated OSN dendrites. a UAS-mCherry::Orco ^WT /UAS-myR::GFP (II); Orco-GAL4, Orco ¹ (III) flies show normal Orco protein localization in OSN soma and ciliated dendrites (magenta) and normal dendritic morphology (green). We confirmed this by visualization of mCherry and GFP, respectively. b Staining of UAS-mCherry::Orco ^WT /UAS-EGFP::OR47a (II); Orco-GAL4, Orco ¹ (III) antennae with an anti-GFP antibody shows normal trafficking of OR47a protein to the ciliated outer dendrites. c–h We then examined the localization of Orco and OR47a proteins in flies expressing Orco^CBSΔ (c, d), Orco^W341Δ (e, f), and Orco^RH344EE (g, h). For unknown reasons, perhaps increased protein degradation, the Orco CBS mutants all show lower mCherry signal than Orco^WT when imaged at the same confocal settings (c, e, g left inset panels). After boosting the confocal gain, it was clear none of the three Orco CBS mutants rescue the localization of Orco to the outer dendrites (c, e, g magenta) despite normal dendritic morphology (c, e, g green). Besides failing to rescue the localization of OR47a protein to the ciliated outer dendrites, expression of the Orco CBS mutants also seems to reduce OR47a staining in the OSN soma (d, f, h). For b, d, f, and h, the color of each image is inverted to improve dendrite visibility. All scale bars, 10 μm. CBS calmodulin binding site, GFP green fluorescent protein, OSN olfactory sensory neuron