Project description:We present a high-throughput in vivo method to identify odorant receptors responding to odorants, using phosphorylated ribosome immunoprecipitation of mRNA from olfactory epithelium of odor-stimulated mice followed by RNA-Seq. pS6-IP RNA-Seq in odor stimulated vs control mice olfactory epithelium
Project description:We present a high-throughput in vivo method to identify odorant receptors responding to odorants, using phosphorylated ribosome immunoprecipitation of mRNA from olfactory epithelium of odor-stimulated mice followed by RNA-Seq.
Project description:A fundamental challenge in studying principles of organization used by the olfactory system to encode odor concentration information has been to identify comprehensive sets of activated odorant receptors (ORs) across a broad concentration range inside freely behaving animals. In mammals, this has recently become feasible with high-throughput sequencing-based methods that identify populations of activated ORs in vivo. In this study, we characterized the mouse OR repertoires activated by the two odorants, acetophenone and 2,5-dihydro-2,4,5-trimethylthiazoline, from 0.01% to 100% (v/v) as starting concentrations using phosphorylated ribosomal protein S6 capture followed by RNA-Seq. We found Olfr923 to be one of the most sensitive ORs that is enriched by acetophenone. Using a mouse line that genetically labels Olfr923-positive axons, we provided evidence that acetophenone activates the Olfr923 glomeruli in the olfactory bulb. Through molecular dynamics stimulations, we identified amino acid residues in the Olfr923 binding cavity that facilitates acetophenone binding. This study sheds light on the active process by which unique OR repertoires may collectively facilitate the discrimination of odorant concentrations.
Project description:Our study shows reduced expression of Trace-Amine Associated Receptors (TAARs) in the olfactory epithelium of mice in which Taar elements 1 and 2 (TE1+2) are deleted in cis.
Project description:Phosphorylation of ribosomal protein S6 (pS6) serves as a molecular marker of neuronal activation by external stimuli. In this study, olfactory epithelium tissues were collected from mice exposed to single odorants (acetophenone, decanal, octanal, and cis-3-hexenol), binary mixtures (acetophenone–decanal and octanal–cis-3-hexenol), or complex fragrances (floral, mint, and floral–mint combination). To selectively capture transcripts undergoing active translation in odor-activated cells, pS6-associated ribosome complexes were isolated from tissue lysates through immunoprecipitation. The enriched mRNAs were subsequently purified and subjected to RNA sequencing, enabling transcriptomic profiling specifically within odor-activated neuronal populations, particularly olfactory sensory neurons. This approach provides a targeted view of gene expression programs induced by diverse odor stimulation paradigms in the heterogeneous olfactory epithelium.
