Integrated Proteotranscriptomics of the Hypothalamus Reveals Altered Regulation Associated with the FecB Mutation in the BMPR1B Gene That Affects Prolificacy in Small Tail Han Sheep

SIMPLE SUMMARY: Increasing sheep litter size is essential to improve lamb production efficiency. In Small Tail Han sheep, the large variation in litter size is determined by segregation of the bone morphogenetic receptor type 1B (BMPR1B) gene. BMPR1B is the major gene for fertility that affects the ovulation rate, and a prolific allele is FecB(B), also called the FecB mutation. The mechanism of how the hypothalamus regulates ovulation number increase in sheep with the FecB mutation is not clear. Using a systems biology workflow in DIABLO, we integrated hypothalamic transcriptomics and proteomics data to predict the main fertility-related biomarkers (gamma-aminobutyric acid type A receptor subunit alpha 1, gamma-aminobutyric acid type A receptor subunit beta 2 and FKBP prolyl isomerase 1A) in the hypothalamus that may regulate changes in the ovulation rate of FecB mutant sheep by participating in gonadotrophin-releasing hormone synthesis and secretion. These results may provide potential genetic markers for breeding multi-lambing meat sheep These results also provide new insight into the reproductive endocrine regulation of fecundity and potential genetic markers of breeding for prolificacy in meat sheep. ABSTRACT: The litter size and ovulation rate are different among ewes of different FecB genotypes in Small Tail Han sheep. These variants in reproductive phenotypes may be regulated by hormones released by the hypothalamic–pituitary–ovarian axis. However, there have been few reports on the hypothalamus regarding regulating an increase in ovulation in sheep with FecB mutation at different estrous stages. Thus, we examined the abundance of hypothalamus tissue protein profiles of six FecB mutant homozygous (BB) and six wild-type (WW) ewes at the luteal and follicular phases. We determined this abundance by tandem mass tag-based quantitative analysis and parallel reaction monitoring methods. Furthermore, an integrated proteotranscriptomic analysis was performed by the Data Integration Analysis for Biomarker discovery using the latent variable approaches for Omics studies (DIABLO) framework to examine biological processes and pathway alterations by the FecB mutant. The abundance of 154 proteins was different between the two estrous stages. Growth hormone and prolactin were particularly enriched in the neuroactive ligand–receptor interactions, the prolactin signaling pathway, and the PI3K-Akt signaling pathway which are related to hypothalamic function and reproduction. We combined proteome and transcriptome data from different estrous stages and genotypes. There is a high correlation (Pearson correlation coefficient = 0.99) between the two datasets in the first two components. We applied the traditional single-omic multivariate approach to obtain differentially abundant proteins and differentially expressed genes. The major fertility related biomarkers were selected using the two approaches mentioned above. Several key pathways (GABAergic synapse, neuroactive ligand–receptor interaction, estrogen and MAPK signaling pathways) were enriched, which are central to gonadotrophin-releasing hormone (GnRH) secretion and reproduction. A higher level of gamma-aminobutyric acid type A receptor subunit alpha1 (GABRA1) and gamma-aminobutyric acid type A receptor subunit beta2 (GABRB2) expression was observed in BB ewes as compared to WW ewes. This finding suggested that a greater production of GnRH during follicular development in BB ewes may explain the higher mature follicle number in mutant ewes. FKBP prolyl isomerase 1A (FKBP1A), which was a major feature factor in the proteome selected by DIABLO, was an important switch for activating the transforming growth factor beta (TGFβ) pathway, and its expression was higher in the WW ewes than in the BB ewes. We suggest that BB sheep maintain TGFβ pathway activity by reducing FKBP1A protein abundance. This innovative data integration in the hypothalamus may provide fresh insight into the mechanisms by which the FecB mutation affects sheep fertility, while providing novel biomarkers related to reproductive endocrinology in sheep breeding.