The Olfactory Transcriptome and Progression of Sexual Maturation in Homing Chum Salmon Oncorhynchus keta

Abstract

Reproductive homing migration of salmonids requires accurate interaction between the reception of external olfactory cues for navigation to the spawning grounds and the regulation of sexual maturation processes. This study aimed at providing insights into the hypothesized functional link between olfactory sensing of the spawning ground and final sexual maturation. We have therefore assessed the presence and expression levels of olfactory genes by RNA sequencing (RNAseq) of the olfactory rosettes in homing chum salmon Oncorhynchus keta Walbaum from the coastal sea to 75 km upstream the rivers at the pre-spawning ground. The progression of sexual maturation along the brain-pituitary-gonadal axis was assessed through determination of plasma steroid levels by time-resolved fluoroimmunoassays (TR-FIA), pituitary gonadotropin subunit expression and salmon gonadotropin-releasing hormone (sgnrh) expression in the brain by quantitative real-time PCR. RNAseq revealed the expression of 75 known and 27 unknown salmonid olfactory genes of which 13 genes were differentially expressed between fish from the pre-spawning area and from the coastal area, suggesting an important role of these genes in homing. A clear progression towards final maturation was characterised by higher plasma 17α,20β-dihydroxy-4-pregnen-3-one (DHP) levels, increased pituitary luteinizing hormone β subunit (lhβ) expression and sgnrh expression in the post brain, and lower plasma testosterone (T) and 17β-estradiol (E2) levels. Olfactomedins and ependymin are candidates among the differentially expressed genes that may connect olfactory reception to the expression of sgnrh to regulate final maturation.

Fig 1. Sample sites of returning Chum salmon at the coastal sea of Ishikari Bay and 75 km upstream at the pre-spawning grounds of the Chitose River near Sapporo on Hokkaido, Japan.

Fig 2. Plasma steroid levels of male and female chum salmon at the coastal sea (Ishikari Bay) and at the pre-spawning ground (Chitose).

(A) 17α,20β-dihydroxy-4-pregnen-3-one (DHP) in males and females at the coastal sea (light grey) and the pre-spawning ground (dark grey). DHP levels are much higher at the pre-spawning ground than at the coastal sea, significant only for the females. (B) Testosterone (T) in males and females at the coastal sea (light grey) and the pre-spawning ground (dark grey). Both in males and females, T levels are significantly lower at the pre-spawning ground vs. the coastal sea. (C) 11-ketotestosterone (11-KT) levels in males are significantly lower at the pre-spawning ground vs. the coastal sea. (D) 17β-estradiol (E2) levels in females are significantly lower at the pre-spawning ground vs. the coastal sea.

Fig 3. Glycoprotein α (gpα), fshβ and lhβ expression in the pituitary of chum salmon.

(A) Expression of each of these target genes (normalized vs. the expression of the housekeeping gene) is shown as fold change of fish at the pre-spawning ground vs. fish at the coastal sea (set at fc 1 as indicated by the line). lhβ expression is significantly higher at fc 2.94 ± 0.40. (B) lhβ expression is significantly higher in females (f) at fc 1.85 ± 0.34 and males (m) at fc 4.77 ± 0.82.

Fig 4. Salmon gonadotropin releasing hormone (sgnrh) expression in the brain of chum salmon.

(A) Expression of sgnrh (normalized vs. the expression of the housekeeping gene) is shown as fold change of fish at the pre-spawning ground vs. fish at the coastal sea (set at fc 1 as indicated by the line). sgnrh expression is significantly higher at fc 1.62 ± 0.26. (B) sgnrh expression is higher in both sexes but significantly higher in males. pb = post-brain; fb = forebrain, f = female; m = male.