Semiochemical responsive olfactory sensory neurons are sexually dimorphic and plastic
- Department of Neurobiology, Neurobiology Graduate Program, Duke University Medical Center, United States
- Department of Molecular Genetics and Microbiology, Duke University Medical Center, United States
- Department of Chemistry, University at Albany, State University of New York, United States
- School of Medicine, Medical Sciences, Indiana University, United States
- Duke Institute for Brain Sciences, Duke University, United States
Figures
Figure 1
Identification of ORs exhibiting sexually dimorphic expression in sex-separated mice.
(A) Schematic of the housing setup. For sex-separation, male mice were co-housed exclusively with male mice. Female mice were co-housed exclusively with female mice. (B) Volcano plot comparing …
Figure 2 with 1 supplement
Sexually dimorphic expression of ORs is consistent with a change in the number of OSNs expressing those ORs.
(A) Left: representative in situ mRNA hybridization pictures probing for the expression of Olfr910 in 43-week-old sex-separated male (top) and female (bottom) mice. Scale bars indicate 50 μm. Right: …
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Figure 2—source data 1
Summary statistics for Figure 2.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig2-data1-v2.xlsx
Figure 2—figure supplement 1
Architecture of reads uniquely mapping to female-enriched ORs.
(A) Architecture of reads uniquely mapped to Olfr910 in male and female mice. Shown are the defined transcript variants, coding sequence (CDS), uniquely designed riboprobe, and 5′ to 3′ …
Figure 3 with 1 supplement
Sexually dimorphic ORs are activated by mature male mouse odor.
(A) Schematic of exposure experiment. A juvenile mouse (black) was exposed to (in descending order) a clean environment, mature male mice, mature female mice, or 1% (v/v) acetophenone for 1 hr in a …
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Figure 3—source data 1
Summary statistics for Figure 3.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig3-data1-v2.xlsx
Figure 3—figure supplement 1
Example in situ stainings showing sexually dimorphic ORs are activated by mature male mouse odor.
(A) Representative images showing in situ mRNA hybridizations probing for Olfr910 expression and pS6 immunostainings. Co-localization events are only seen following mature male mouse exposure as …
Figure 4 with 3 supplements
Sexually dimorphic ORs are activated by mature male mouse semiochemicals SBT and MTMT.
(A) Schematic of the pS6-IP-Seq experiment. Litter matched, ~3 week-old (juvenile) mice are used. Mice are habituated to an odor-free environment for 1 hr. One mouse then receives exposure to an …
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Figure 4—source data 1
Summary statistics for Figure 4.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig4-data1-v2.xlsx
Figure 4—figure supplement 1
Sexually dimorphic ORs are not activated by sex-specific or sex-enriched odorants that are not SBT or MTMT.
(A) Volcano plot showing the results of pS6-IP-Seq using 1% (v/v) β-caryophyllene dissolved in water as stimulus. Olfr910, Olfr912, and Olfr1295 are highlighted in red and not enriched. The red …
Figure 4—figure supplement 2
Cognate ORs for other sex-specific and sex-enriched volatiles are not sexually dimorphic.
(A) Left: the top five candidate ORs activated by 1% (v/v) 2-heptanone exposure, based on lowest FDR values, are highlighted in red. Right: none of the top five candidate ORs activated by 1% (v/v) …
Figure 4—figure supplement 3
Example in situ stainings showing sexually dimorphic ORs are activated by SBT and MTMT.
(A) Representative images showing in situ mRNA hybridizations probing for Olfr910 expression and pS6 immunostainings. Co-localization events are only seen following SBT exposure as indicated by …
Figure 5 with 1 supplement
OSN responses to semiochemicals are not sexually dimorphic between mature male and female mice.
(A) Comparison of responses of OSNs from 26-week-old male and female mice to various stimuli. One-way ANOVA with Tukey’s multiple comparisons test correction reveals only exposure to 0.1% (v/v) SBT …
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Figure 5—source data 1
Summary statistics for Figure 5.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig5-data1-v2.xlsx
Figure 5—figure supplement 1
Example in situ stainings showing responses to semiochemicals are not sexually dimorphic between mature male and female mice.
(A) Representative images from mature male and female mice showing in situ mRNA hybridizations probing for Olfr910 expression and pS6 immunostainings. Co-localization events are only seen following …
Figure 6
Sex-combined housing leads to the attenuation of the dimorphic OR representations.
(A) Schematic of the housing setup. For sex-combined housing, one male mouse was co-housed with one female mouse. (B) Volcano plot comparing expression of Olfrs between 43-week-old sex-combined male …
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Figure 6—source data 1
Summary statistics for Figure 6.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig6-data1-v2.xlsx
Figure 7 with 1 supplement
Olfr1437 and Olfr235 are male-enriched and respond to the macrocyclic musk molecule CPD.
(A) Volcano plot comparing expression of Olfrs between 43-week-old sex-separated male and female mice. Olfr1437 and Olfr235 and are highlighted in red. The red dashed line indicates an FDR = 0.05. …
Figure 7—figure supplement 1
Male over-expression is also consistent with a difference in the number of cells expressing Olfr1437 between male and female mice.
(A) Left: representative in situ mRNA hybridization pictures probing for the expression of Olfr1437 in 43-week-old sex-separated male (top) and female (bottom) mice. Scale bars indicate 50 μm. …
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Figure 7—figure supplement 1—source data 1
Summary statistics for Figure 7—figure supplement 1.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig7-figsupp1-data1-v2.xlsx
Figure 8
Single-cell RNA-Seq analysis reveals an enrichment of activity-associated gene expression in male OSNs expressing Olfr910, Olfr912, and Olfr1295.
(A) Chemosensory receptor-independent UMAP embedding of 17,666 singly sequenced mature OSNs. OSNs expressing Olfr910, Olfr912, Olfr1295, Olfr1437, and Olfr235 are highlighted. (B) Gene expression …
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Figure 8—source data 1
Summary statistics for Figure 8.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig8-data1-v2.xlsx
Figure 9 with 2 supplements
Sex-separated Bax-/- mice fail to generate sexually dimorphic representations of Olfr910, Olfr912, and Olfr1295.
(A) Volcano plot comparing expression of Olfrs between 26-week-old Bax-/- sex separated male and female mice. Olfr910, Olfr912, and Olfr1295 are highlighted in red. The red dashed line indicates an …
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Figure 9—source data 1
Summary statistics for Figure 9.
- https://cdn.elifesciences.org/articles/54501/elife-54501-fig9-data1-v2.xlsx
Figure 9—figure supplement 1
Sex-separated Bax-/- mice fail to generate sexually dimorphic representations of Olfr1437 and Olfr235. .
(A) Volcano plot comparing expression of Olfrs between 26-week-old Bax-/- sex-separated male and female mice. Olfr1437 and Olfr235 are highlighted in red. The red dashed line indicates an FDR = 0.05.…
Figure 9—figure supplement 2
Bax-/- mice exhibit significant changes in their OR repertoire.
(A) Volcano plot comparing expression of Olfrs between 26-week-old wild-type and Bax-/- sex-separated male mice. Olfr910, Olfr912, Olfr1295, Olfr1437, and Olfr235 are highlighted in red. The red …
Author response image 1
Author response image 2
Author response image 3
Tables
Key resources table
| Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
|---|---|---|---|---|
| Strain, strain background (Mus musculus C57BL/6J) | C57BL/6J | Jackson Labs | 000664 | |
| Strain, strain background (Mus musculus C57BL/6J) | Bax-/- | Jackson Labs | 002994 | |
| Commercial assay, kit | TRIzol | Life Technologies | 15596026 | |
| Commercial assay, kit | QUBIT HS RNA Assay Kit | ThermoFisher | Q32855 | |
| Commercial assay, kit | RNase-free DNaseI | Roche | 04 716 728 001 | |
| Commercial assay, kit | RNeasy Mini Kit | Qiagen | 74104 | |
| Commercial assay, kit | SMART-Seq v4 Ultra Low Input RNA Kit | Takara | 634898 | |
| Commercial assay, kit | Nexterra XT DNA Library Preparation Kit | Illumina | 15032354 | |
| Sequence-based reagent | Olfr910 UTR F | This paper | 5'-AAACGCGTGTGAAAATTGTGACAGATCCA-3' | |
| Sequence-based reagent | Olfr910 UTR R | This paper | 5'-AAGCGGCCGCCATTTACAAGAAGGGAATCAG-3' | |
| Sequence-based reagent | Olfr912 UTR F | This paper | 5'-AAACGCGTACTTTGTTCTGATTCAGTTGTT-3' | |
| Sequence-based reagent | Olfr912 UTR R | This paper | 5′-AAGCGGCCGCGTCCACAGAGCAATACAACA-3′ | |
| Sequence-based reagent | Olfr1295 UTR F | This paper | 5'-AAACGCGTACTCCTCTCCTAAATCCAAC-3' | |
| Sequence-based reagent | Olfr1295 UTR R | This paper | 5'-AAGCGGCCGCGGCAGCACCACTGATCAA-3' | |
| Commercial assay, kit | Phusion DNA Polymerase | NEB | F530S | |
| Commercial assay, kit | pCI vector | Promega | E1731 | |
| Commercial assay, kit | DNA polymerase | Qiagen | 203203 | |
| Commercial assay, kit | MinElute Kit | Qiagen | 28004 | |
| Commercial assay, kit | T3 RNA polymerase | Promega | P2083 | |
| Commercial assay, kit | DIG RNA labeling mix | Roche | 11277073910 | |
| Commercial assay, kit | Micro Bio-Spin P-30 Gel Columns | Bio-Rad | 732–6223 | |
| Commercial assay, kit | Tissue-Tek O.C.T. Compound | Sakura Finetek | 4583 | |
| Commercial assay, kit | Superfrost Plus Slides | Fisherbrand | 1255015 | |
| Commercial assay, kit | Parafilm | Sigma | P7793 | |
| Commercial assay, kit | Blocking reagent | Roche | 11096176001 | |
| Commercial assay, kit | Maleic acid | Sigma | M0375 | |
| Antibody | Anti-DIG antibody (sheep polyclonal) | Roche | 11207733910 | 1:1000 |
| Commercial assay, kit | TSA-Fluorescein | PerkinElmer | NEL741B001KT | 1:400 |
| Antibody | pS6 Antibody for IF (rabbit polyclonal) | ThermoFisher | 44–923G | 1:300 |
| Antibody | Anti-rabbit IgG Cy3 Antibody (donkey polyclonal) | Jackson Immuno | 711-165-152 | 1:200 |
| Other | Bisbenzimide | Sigma | H 33258 | 1:100,000 |
| Other | Paper bucket | International Paper | DFM85 | |
| Other | Blotting pad | VWR | 28298–014 | |
| Other | Odor cassette | Sakura Finetek | 0006772–01 | |
| Commercial assay, kit | LoBind Tube | Eppendorf | 22431021 | |
| Commercial assay, kit | NP40 Substitute | Sigma | 11332473001 | |
| Commercial assay, kit | DHPC | Avanti Polar Lipids | 850306P | |
| Antibody | pS6 Antibody for IP (rabbit monoclonal) | Cell Signaling | D68F8 | 6 μL |
| Commercial assay, kit | Dynabeads Protein A | Invitrogen | 10002D | |
| Commercial assay, kit | RNeasy Micro Kit | Qiagen | 74004 | |
| Other | β-Caryophyllene | Sigma | W225207 | |
| Other | 2,5-DMP | Sigma | 175420 | |
| Other | 2-Heptanone | Sigma | W254401 | |
| Other | (E)-β-Farnesene | Bedoukian | P3500-90 | |
| Other | CPD | Sigma | C111201 | |
| Other | MTMT | Lin et al., 2005 | Synthesized | |
| Other | SBT | Meijer et al., 1973; Abrunhosa et al., 2001; Tashiro and Mori, 1999 | Synthesized | |
| Other | DHB | Wiesler et al., 1984 | Synthesized |
