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.2023 Jan 31;10(1):62.
doi: 10.1038/s41597-023-01967-w.

An RNA-seq time series of the medaka pituitary gland during sexual maturation

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An RNA-seq time series of the medaka pituitary gland during sexual maturation

Eirill Ager-Wick et al. Sci Data..

Abstract

Directing both organismal homeostasis and physiological adaptation, the pituitary is a key endocrine gland in all vertebrates. One of its major tasks is to coordinate sexual maturation through the production and release of hormones stimulating gonad development. In order to study its developmental dynamics in the model fish medaka (Oryzias latipes), we sampled both the pituitary and the ovaries of 68 female fish. Of these, 55 spanned the entire course of sexual maturation from prepubertal juveniles to spawning adults. An additional 13 showed either considerably faster or slower growth and development than the majority of fish. We used histological examination of the ovaries to determine a histological maturation stage, and analyzed the pituitary glands using RNA-seq optimized for low input. Taken together, these data reveal the timing of hormone production priorities, and form a comprehensive resource for the study of their regulation.

© 2023. The Author(s).

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Growth and maturation in female medaka. The 67 fish analyzed in this study range from young, small and sexually immature (pre-vitellogenic gonads) to old, large and sexually fully mature. Gonad maturation status was assessed using histology for most samples, following the stages described by Iwamatsuet al. (ref. ); for a few fish, we only obtained macroscopic estimates, and for two no estimates. We did not perform histological staging for the gonads of most of the mature, spawning fish. Closed circles indicate the 54 samples taken as part of the main time series, 11 samples flagged during the growth experiment as especially fast- or slow-growing are indicated by open circles. Two further slow-growing fish (7 and 26) are indicated by open squares – these biological outliers also show a relatively strongly deviating transcriptomic profile (see Figs. 3,4). One technical outlier sample (119), not included in any further analyses, is marked by a cross.
Fig. 2
Fig. 2
Histological confirmation of gonadal maturation. Shown are example images of the eight stages. The size of the largest observed follicle determines the maturation status (see Table 1). Bars correspond to 100 µm (panels a and b, 100 × magnification) or 250 µm (panels c–h, 40 × magnification). Samples shown here are (by fish number) (a) 109; (b) 14; (c) 24; (d) 5; (e) 4; (f) 18; (g) 63; and (h) 115.
Fig. 3
Fig. 3
Outlier detection and sample reproducibility. (a,b) Principal component analysis (PCA) of all pituitary transcriptome profiles identified three samples as potential outliers. Mature fish 119 (cross) is strongly divergent at the transcriptomic level only, and was removed from the analyses. Samples 7 and 26 are also transcriptomic outliers, however they represent unusual biological states (slow growth, see Fig. 1) and were therefore retained in the dataset. Shown are the first two components for PCA on log-transformed expression values of medium-high expressed genes using quantile (a) or scaling (b) sample normalization. (c) PCA after outlier (119) removal and quantile re-normalization, using medium-high expression genes and main time series samples only. Principal component 1 shows a clear parallel with maturation (colour scale as in Fig. 1).
Fig. 4
Fig. 4
Expression changes during development and maturation. Heatmap of the variation in expression of the most highly expressed genes in the medaka pituitary, defined as ranking in the top 10 in any of the 54 samples of the main time series. Also included are the fast and slow maturation groups, as well as the two slowly growing outlier samples. Variation is shown as standard deviation around the mean (z-score) for each gene. Genes are hierarchically clustered by expression pattern. Genes in bold encode the major pituitary protein hormones; numbers (in grey) are shorthand for Ensembl identifiers of unnamed genes (e.g. 09786 refers to gene ENSORLG00000009786). Time series samples are ordered by the first principal component of the RNA-seq profiles (see Fig. 3).
Fig. 5
Fig. 5
Alternative normalization strategies. Shown are expression levels per sample, normalized using either a quantile strategy (a) or TMM scaling (b). Violin plots show the distributions of the levels for medium-high expression genes, with median values (circles) and the lower to upper quartile range (bars) indicated. After quantile normalization, expression profiles are comparable between samples; scaling normalization does not succeed in making samples comparable.
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