Project description:Odorants are thought to activate sets of odorant receptors in vivo, but capturing sets of responsive receptors in vivo has never been accomplished. GeneChip microarrays were used to identify the odorant receptor mRNAs enriched in samples from activated olfactory neurons
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:Genetic variation governs protein expression through both transcriptional and post-transcriptional processes. To investigate this relationship, we combined a multiplexed, mass spectrometry-based method for protein quantification with an emerging mouse model harboring extensive genetic variation from 8 founder strains. We collected genome-wide mRNA and protein profiling measurements to link genetic variation to protein expression differences in livers from 192 diversity outcross mice. We observed nearly 3,700 protein-level quantitative trait loci (pQTL) with an equal proportion of proteins regulated directly by their cognate mRNA as uncoupled from their transcript. Our analysis reveals an extensive array of at least five models for genetic variant control of protein abundance including direct protein-to-protein associations that act to achieve stoichiometric balance of functionally related enzymes and subunits of multimeric complexes.
Project description:Genetic variation governs protein expression through both transcriptional and post-transcriptional processes. To investigate this relationship, we combined a multiplexed, mass spectrometry-based method for protein quantification with an emerging mouse model harboring extensive genetic variation from 8 founder strains. We collected genome-wide mRNA and protein profiling measurements to link genetic variation to protein expression differences in livers from 192 diversity outcross mice. We observed nearly 3,700 protein-level quantitative trait loci (pQTL) with an equal proportion of proteins regulated directly by their cognate mRNA as uncoupled from their transcript. Our analysis reveals an extensive array of at least five models for genetic variant control of protein abundance including direct protein-to-protein associations that act to achieve stoichiometric balance of functionally related enzymes and subunits of multimeric complexes.
Project description:Noncoding genetic variation is a major driver of phenotypic diversity but determining the underlying mechanisms and the cell types in which it acts remain challenging problems. Here, we investigate the impact of natural genetic variation provided by phenotypically diverse inbred strains of mice on gene expression and epigenetic landscapes of Kupffer cells. Analysis of gene expression in Kupffer cells and other liver cell types derived from C57BL/6J, BALB/cJ and A/J mice provided evidence for strain-specific differences in environmental factors influencing Kupffer cell phenotypes, including preferential Leptin signaling in BALB/cJ Kupffer cells. Systematic analysis of transcriptomic and epigenetic data from F1 hybrids of these mice, and transcriptomic data from strain-specific Kupffer cells engrafted into a common host enabled quantitative assessment of cis versus trans effects of genetic variation on gene expression and an estimate of cell autonomous versus non cell autonomous effects. Under homeostatic conditions, trans effects of genetic variation were dominant, with the majority of trans regulation being non cell autonomous. In contrast, strain specific responses to acutely administered LPS were primarily associated with genetic variation acting in cis to modify response elements for lineage determining and signal dependent transcription factors. Collectively, these findings reveal cell intrinsic and environmental effects of natural genetic variation on gene expression, demonstrate the use of enhancers as detectors of trans effects of genetic variation, and provide a new resource for understanding the impact of genetic variation on gene expression in Kupffer cells.