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Deep sequencing of the murine olfactory receptor neuron transcriptome.

Kanageswaran N, Demond M, Nagel M, Schreiner BS, Baumgart S, Scholz P, Altmüller J, Becker C, Doerner JF, Conrad H, Oberland S, Wetzel CH, Neuhaus EM, Hatt H, Gisselmann G - PLoS ONE (2015)

Bottom Line: In OE samples, nearly all OR and trace amine-associated receptor (TAAR) genes involved in the perception of volatile amines were detectably expressed.To identify OE-specific genes, we compared olfactory neuron expression profiles with RNA-Seq transcriptome data from different murine tissues.We also identified other previously undescribed membrane proteins as potential new players in olfaction.

View Article: PubMed Central - PubMed

Affiliation: Ruhr-University Bochum, Department of Cell Physiology, Bochum, Germany.

ABSTRACT
The ability of animals to sense and differentiate among thousands of odorants relies on a large set of olfactory receptors (OR) and a multitude of accessory proteins within the olfactory epithelium (OE). ORs and related signaling mechanisms have been the subject of intensive studies over the past years, but our knowledge regarding olfactory processing remains limited. The recent development of next generation sequencing (NGS) techniques encouraged us to assess the transcriptome of the murine OE. We analyzed RNA from OEs of female and male adult mice and from fluorescence-activated cell sorting (FACS)-sorted olfactory receptor neurons (ORNs) obtained from transgenic OMP-GFP mice. The Illumina RNA-Seq protocol was utilized to generate up to 86 million reads per transcriptome. In OE samples, nearly all OR and trace amine-associated receptor (TAAR) genes involved in the perception of volatile amines were detectably expressed. Other genes known to participate in olfactory signaling pathways were among the 200 genes with the highest expression levels in the OE. To identify OE-specific genes, we compared olfactory neuron expression profiles with RNA-Seq transcriptome data from different murine tissues. By analyzing different transcript classes, we detected the expression of non-olfactory GPCRs in ORNs and established an expression ranking for GPCRs detected in the OE. We also identified other previously undescribed membrane proteins as potential new players in olfaction. The quantitative and comprehensive transcriptome data provide a virtually complete catalogue of genes expressed in the OE and present a useful tool to uncover candidate genes involved in, for example, olfactory signaling, OR trafficking and recycling, and proliferation.

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Expression ranking of non-olfactory GPCRs (nGPCR).A. Heatmap showing the ranking of the 30 most highly expressed nGPCRs in the FACS-sorted ORNs. Only six nGPCRs were among the 1,000 most highly expressed genes (Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3). Of the 30 most highly expressed genes, the expression of 17 in the OE was no previously known. B. Heat map showing the ranking of nGPCRs that are specifically enriched in ORNs according the criteria that the nGPCRs’ FPKMs were greater than 1 and 5 times greater in the ORNs than in non-olfactory tissue (brain, liver, muscle and testes). A total of 18 specifically enriched nGPCRs were found in ORNs. Excluding the specifically enriched candidates that were already presented in the list of the 30 most highly expressed genes, an additional 10 genes were found to be specifically enriched and are new in terms of olfaction. Regarding the genes that were among the 30 most highly expressed and were specifically enriched in ORNs, 60% had neither been shown to be expressed in the OE or been ascribed any function in the OE in any previous study with the exception of for several candidates in a tabular form in a recent transcriptome-wide study of the total OE [52].
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pone.0113170.g006: Expression ranking of non-olfactory GPCRs (nGPCR).A. Heatmap showing the ranking of the 30 most highly expressed nGPCRs in the FACS-sorted ORNs. Only six nGPCRs were among the 1,000 most highly expressed genes (Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3). Of the 30 most highly expressed genes, the expression of 17 in the OE was no previously known. B. Heat map showing the ranking of nGPCRs that are specifically enriched in ORNs according the criteria that the nGPCRs’ FPKMs were greater than 1 and 5 times greater in the ORNs than in non-olfactory tissue (brain, liver, muscle and testes). A total of 18 specifically enriched nGPCRs were found in ORNs. Excluding the specifically enriched candidates that were already presented in the list of the 30 most highly expressed genes, an additional 10 genes were found to be specifically enriched and are new in terms of olfaction. Regarding the genes that were among the 30 most highly expressed and were specifically enriched in ORNs, 60% had neither been shown to be expressed in the OE or been ascribed any function in the OE in any previous study with the exception of for several candidates in a tabular form in a recent transcriptome-wide study of the total OE [52].

Mentions: We classified the nGPCRs into five main GPCR families: glutamate, rhodopsin, adhesion, frizzled and secretin. This classification reveals that the nGPCRs (including genes with FPKMs below 1) that were expressed in ORNs mainly belong to the rhodopsin family (64%). Ten percent belong to the adhesion family, approximately 4% to the frizzled family, 6% to the glutamate family, and 0.2% to the secretin family. An additional set of nGPCRs with unknown functions, no known ligands and atypical structure could not be classified (approximately 12%) nGPCRs that are highly expressed may hypothetically function as co-receptors in ORNs [86, 87]. Therefore, we created a ranking of the expression of all nGPCRs to identify the highly expressed candidates (Fig. 6). Interestingly, only six nGPCRs were ranked among the 1,000 most highly expressed genes in ORNs: Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3 (Fig. 6). An additional 30 nGPCRs exhibited medium expression profiles (FPKMs between 52–10), and 65 nGPCRs exhibited low expression profiles (FPKMs between 1–10) (S11 Fig.). Some of these GPCRs have previously been reported to be expressed in ORNs [48, 49]. Regarding the most highly expressed 30 nGPCRs, our study describes the expression of an additional 17 genes: Gpr178, Lphn1, Gpr137, Gpr162, Gpr155, Gpr63, Paqr9, Gpr108, Lgr4, Gpr89, Wls, A030009H04Rik, Gpr107, Gpr87, Gpr137b, Ptger1 and P2ry14 (Fig. 6). A few of these genes have also been tabularly presented in a recent transcriptome wide study of the total OE [52].


Deep sequencing of the murine olfactory receptor neuron transcriptome.

Kanageswaran N, Demond M, Nagel M, Schreiner BS, Baumgart S, Scholz P, Altmüller J, Becker C, Doerner JF, Conrad H, Oberland S, Wetzel CH, Neuhaus EM, Hatt H, Gisselmann G - PLoS ONE (2015)

Expression ranking of non-olfactory GPCRs (nGPCR).A. Heatmap showing the ranking of the 30 most highly expressed nGPCRs in the FACS-sorted ORNs. Only six nGPCRs were among the 1,000 most highly expressed genes (Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3). Of the 30 most highly expressed genes, the expression of 17 in the OE was no previously known. B. Heat map showing the ranking of nGPCRs that are specifically enriched in ORNs according the criteria that the nGPCRs’ FPKMs were greater than 1 and 5 times greater in the ORNs than in non-olfactory tissue (brain, liver, muscle and testes). A total of 18 specifically enriched nGPCRs were found in ORNs. Excluding the specifically enriched candidates that were already presented in the list of the 30 most highly expressed genes, an additional 10 genes were found to be specifically enriched and are new in terms of olfaction. Regarding the genes that were among the 30 most highly expressed and were specifically enriched in ORNs, 60% had neither been shown to be expressed in the OE or been ascribed any function in the OE in any previous study with the exception of for several candidates in a tabular form in a recent transcriptome-wide study of the total OE [52].
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4295871&req=5

pone.0113170.g006: Expression ranking of non-olfactory GPCRs (nGPCR).A. Heatmap showing the ranking of the 30 most highly expressed nGPCRs in the FACS-sorted ORNs. Only six nGPCRs were among the 1,000 most highly expressed genes (Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3). Of the 30 most highly expressed genes, the expression of 17 in the OE was no previously known. B. Heat map showing the ranking of nGPCRs that are specifically enriched in ORNs according the criteria that the nGPCRs’ FPKMs were greater than 1 and 5 times greater in the ORNs than in non-olfactory tissue (brain, liver, muscle and testes). A total of 18 specifically enriched nGPCRs were found in ORNs. Excluding the specifically enriched candidates that were already presented in the list of the 30 most highly expressed genes, an additional 10 genes were found to be specifically enriched and are new in terms of olfaction. Regarding the genes that were among the 30 most highly expressed and were specifically enriched in ORNs, 60% had neither been shown to be expressed in the OE or been ascribed any function in the OE in any previous study with the exception of for several candidates in a tabular form in a recent transcriptome-wide study of the total OE [52].
Mentions: We classified the nGPCRs into five main GPCR families: glutamate, rhodopsin, adhesion, frizzled and secretin. This classification reveals that the nGPCRs (including genes with FPKMs below 1) that were expressed in ORNs mainly belong to the rhodopsin family (64%). Ten percent belong to the adhesion family, approximately 4% to the frizzled family, 6% to the glutamate family, and 0.2% to the secretin family. An additional set of nGPCRs with unknown functions, no known ligands and atypical structure could not be classified (approximately 12%) nGPCRs that are highly expressed may hypothetically function as co-receptors in ORNs [86, 87]. Therefore, we created a ranking of the expression of all nGPCRs to identify the highly expressed candidates (Fig. 6). Interestingly, only six nGPCRs were ranked among the 1,000 most highly expressed genes in ORNs: Adipor1, Gpr178, Gabbr1, Gprc5c, Drd2, and Lphn3 (Fig. 6). An additional 30 nGPCRs exhibited medium expression profiles (FPKMs between 52–10), and 65 nGPCRs exhibited low expression profiles (FPKMs between 1–10) (S11 Fig.). Some of these GPCRs have previously been reported to be expressed in ORNs [48, 49]. Regarding the most highly expressed 30 nGPCRs, our study describes the expression of an additional 17 genes: Gpr178, Lphn1, Gpr137, Gpr162, Gpr155, Gpr63, Paqr9, Gpr108, Lgr4, Gpr89, Wls, A030009H04Rik, Gpr107, Gpr87, Gpr137b, Ptger1 and P2ry14 (Fig. 6). A few of these genes have also been tabularly presented in a recent transcriptome wide study of the total OE [52].

Bottom Line: In OE samples, nearly all OR and trace amine-associated receptor (TAAR) genes involved in the perception of volatile amines were detectably expressed.To identify OE-specific genes, we compared olfactory neuron expression profiles with RNA-Seq transcriptome data from different murine tissues.We also identified other previously undescribed membrane proteins as potential new players in olfaction.

View Article: PubMed Central - PubMed

Affiliation: Ruhr-University Bochum, Department of Cell Physiology, Bochum, Germany.

ABSTRACT
The ability of animals to sense and differentiate among thousands of odorants relies on a large set of olfactory receptors (OR) and a multitude of accessory proteins within the olfactory epithelium (OE). ORs and related signaling mechanisms have been the subject of intensive studies over the past years, but our knowledge regarding olfactory processing remains limited. The recent development of next generation sequencing (NGS) techniques encouraged us to assess the transcriptome of the murine OE. We analyzed RNA from OEs of female and male adult mice and from fluorescence-activated cell sorting (FACS)-sorted olfactory receptor neurons (ORNs) obtained from transgenic OMP-GFP mice. The Illumina RNA-Seq protocol was utilized to generate up to 86 million reads per transcriptome. In OE samples, nearly all OR and trace amine-associated receptor (TAAR) genes involved in the perception of volatile amines were detectably expressed. Other genes known to participate in olfactory signaling pathways were among the 200 genes with the highest expression levels in the OE. To identify OE-specific genes, we compared olfactory neuron expression profiles with RNA-Seq transcriptome data from different murine tissues. By analyzing different transcript classes, we detected the expression of non-olfactory GPCRs in ORNs and established an expression ranking for GPCRs detected in the OE. We also identified other previously undescribed membrane proteins as potential new players in olfaction. The quantitative and comprehensive transcriptome data provide a virtually complete catalogue of genes expressed in the OE and present a useful tool to uncover candidate genes involved in, for example, olfactory signaling, OR trafficking and recycling, and proliferation.

Show MeSH
Related in: MedlinePlus