Fig. 5
VRs with sex-specific expression differences exhibit sex-specific expression frequencies and detect sex-specific odors. a Hierarchical clustering of VRs identified via RNA-seq as differentially expressed between SF and SM mice (green arrows). VRs labeled asterisk, FDR < 0.05; other VRs shown, unadjusted p < 0.01 (Cufflinks output). b RNA-seq read alignments to Vmn1r89 for SF and SM samples. Alignments from the three biological replicates in each experimental group were combined (SF, top; SM; bottom). Strand orientation: −, pink; +, blue. c VNO expression levels, based on RNA-seq, for Vmn1r89. Error bars: 95% c.i. d Quantification, using two-color RNA-FISH, of the frequency of Vmn1r89-expressing VSNs relative to all mature VSNs (based on Omp expression). Error bars: s.e.m. **p < 0.01 (two-tailed t test); n = 4–6 mice (5 sections/mouse; average of 17 Vmn1r89-expressing VSNs/section). Dots represent average values for individual mice. e Representative images of Vmn1r89 expression in the VNO of the experimental mouse groups shown. f (Left) Known conspecific activators of the VRs shown in a, based on †: ref. 39, ‡: ref. 35, #: ref. 38. (Right) Quantification, based on two-color RNA-FISH, of the fraction of VSNs that co-express the VRs listed in a and Egr1 within the VNOs of individual male mice exposed to 4 females (♀ mice) or females exposed to 4 males (♂ mice) for 40 min. Vmn1r45 is a representative negative control VR. Statistical results in f are based on comparisons between the fractions of VSNs co-expressing Egr1 after exposure to female or male mice. *p < 0.05, **p < 0.01 (two-tailed t test); n = 4 VNO sections (average of eight specific VR-expressing VSNs/section); gray fill: not tested. g, h Representative images of two-color RNA-FISH analyses of Vmn1r185 co-expression with Egr1 following exposure of a male mouse to four females (♂ exposed to ♀; g) or a female mouse to four males (♀ exposed to ♂; h). Arrows: locations of representative Vmn1r185-expressing cells. SF sex-separated females, SM sex-separated males, CF sex-combined females, SM sex-combined males. Scale bars: 100 µm
