Olfactory receptor accessory proteins play crucial roles in receptor function and gene choice
- Duke University Medical Center, United States
- Graduate School of Agricultural and Life Sciences, The University of Tokyo, Japan
- Boston University, United States
- Tokyo University of Agriculture and Technology, Japan
- Biogen Inc, United States
- Duke Institute for Brain Sciences, United States
Figures
Figure 1
Deletion of RTP1 and RTP2 causes defects in the OE.
(A) Strategy for knocking out RTP1 and RTP2 in series. (B) RNA in situ hybridization with probes specific to RTP1 and RTP2 in both wild-type and RTP1,2DKO mice showing that the knock out mice do not …
Figure 2 with 1 supplement
RTP1,2DKO mice have fewer mature sensory neurons.
(A) Paired comparison of the thickness of the OE measured at five matched positions (see methods) between RTP1,2DKO and their wild-type littermate. (p=0.02, paired student t test). (B) RNA in situ …
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Figure 2—source data 1
OE thickness and percent area occupied by the OMP layer, ACIII layer and GAP43 layer in the wild-type and RTP1,2DKO.
- https://doi.org/10.7554/eLife.21895.005
Figure 2—figure supplement 1
Low magnification view of OMP in situ hybridization signals in OE sections.
(A) Matching OE sections from Wild-type (left) and RTP1,2DKO (right).
Figure 3
Diminished activity in response to odorants in RTP1,2DKO.
(A) Electroolfactograms show the response to seven odorants wild-type. The grey line denotes the air only blank averaged over multiple interleaved trials interspersed within the series. (B) …
Figure 4 with 1 supplement
Representation of ORs in RTP1,2DKO.
(A) Comparison of all transcripts between the wild-type and RTP1,2DKO, the green dots represent ORs, higher read counts for ORs are observed in the wild-type compared to RTP1,2DKO. (B) A comparison …
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Figure 4—source data 1
Percent positive cell counts for the uORs and oORs in Figure 4E and 4 hr.
- https://doi.org/10.7554/eLife.21895.009
Figure 4—figure supplement 1
Representation of ORs in RTP1,2DKO using all genes.
(A) Volcano plot showing the fold change of the expression levels (x-axis) of the ORs between wild-types and RTP1,2DKO using read counts normalized by all genes from our sequencing data. Y axis …
Figure 5
The proportion of OSNs expressing oORs increases in older RTP1,2DKO.
(A) Representative images from 1 day, 21 day and 6 month OE stained with a probe mix against 11 of the uORs expressed in the dorsal OE. (B) Quantification of the percent dorsal uOR positive cells at …
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Figure 5—source data 1
Percent positive cell counts for the uOR and oOR probe mix at 1 day, 21 day and 6 month old OE.
- https://doi.org/10.7554/eLife.21895.012
Figure 6
OR protein sequences determine representation in RTP1,2DKO.
(A) Phylogenetic tree showing uORs in black and oORs in red. (B) Schematic depiction of β2AR-IRES-tau LacZ. (C) The percent Olfr151 positive cells is smaller in RTP1,2DKO mouse (left panels). β2Adren…
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Figure 6—source data 1
Normalized geometric mean for PE intensity obtained from our FACS experiment.
- https://doi.org/10.7554/eLife.21895.014
Figure 7
OSNs expressing oORs from RTP1,2DKO can mature and function.
(A) Representative images showing the colocalization of Olfr923 (uOR) and Olfr78 (oOR) (green) with OMP (red) for het (top) and RTP1,2DKO (bottom). OMP negative OSNs are indicated with arrows. (B) …
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Figure 7—source data 1
Numbers of uOR and oOR neurons found within the OMP layer and outside it.
- https://doi.org/10.7554/eLife.21895.016
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Figure 7—source data 2
Normalized pS6 staining intensity for Olfr923 and Olfr1395 positive cells from het and RTP1,2DKO OE in response to 1%acetophenone and 1%TMT respectively.
- https://doi.org/10.7554/eLife.21895.017
Figure 8
nATF5 expression persists in OSNs expressing uORs but not oORs in RTP1,2DKO.
(A) Expanded expression pattern of nATF5 is observed for RTP1,2DKO Scale bars = 25 μm. S.C. = Sustentacular cells. (B) Analysis of individual sections of wild-type and RTP1,2DKO OE for ATF5 …
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Figure 8—source data 1
Percent uOR/oOR positive cells that co-localize with nATF5 in wild-types and RTP1,2DKO.
- https://doi.org/10.7554/eLife.21895.019
Figure 9 with 1 supplement
Unstable expressing of M71 in RTP1,2DKO.
(A) Schematic depiction of OSN lineage tracing. We crossed a mouse carrying M71–IRES- Cre with Rosa26-lox-stop-lox-tdTomato. In the progeny, the expression of Olfr151 (M71) in an OSN will drive the …
Figure 9—figure supplement 1
OSNs expressing Olfr151 do not switch to Olfr143.
(A) Representative images from the wild-type and RTP1,2DKO OE stained against Olfr143 (green) and tdTomato (red). (B) The percent Olfr143 positive cells is greater in RTP1,2DKO mouse and tdTomato …
Figure 10
RTP1,2DKO mice are able to form glomeruli.
(A) OMP staining shown in red and nuclear staining in cyan. Both wild-type and RTP1,2DKO mice have OMP in the glomerular layer. Scale bar = 25 μm. (B) A whole mount GFP fluorescence from axons …
Figure 11
Model for the role of RTP1 and RTP2 in OR gene choice.
(A) A model showing that in the wild-type (left), ORs are transported to the surface of the cell in conjunction with the RTPs. In the absence of the RTPs (right), oORs reach the surface of the cell …
Additional files
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Supplementary file 1
The number of sequence reads that map each annotated gene in RNA-Seq from three wild-type and 3 RTP1,2DKO mice. FDR is calculated against the entire data set and the fold change (logFC) is displayed as the log (average wild-type reads/ average RTP1,2DKO reads).
- https://doi.org/10.7554/eLife.21895.024
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Supplementary file 2
List of oORs and uORs with their chromosomal locations and their expression zones (dorsal versus ventral) in the OE.
- https://doi.org/10.7554/eLife.21895.025
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Supplementary file 3
Results of one-way ANOVA and Tukey’s post hoc tests
- https://doi.org/10.7554/eLife.21895.026
