Continuous artificial selection in production-oriented breeding has left selective signatures and genomic variability in domesticated sheep. In this study, we performed whole-genome sequencing of 10 Bamei mutton sheep with different litter sizes from the same population. In this population, reproductive traits have undergone intensive selection via the breeding strategy. In this research, numerous mutations were selected in the population after screening, but the rates of heterozygosity did not differ between the two populations. When a neighbor-joining tree was constructed using SNPs selected based on FST score, these two groups were differentiated into two genetic clusters. Most quantitative traits are known to respond quickly to artificial selection, and this population subjected to selection of litter size might have evolved in opposite directions with soft sweeps (28, 29).
Selective sweeps can identify important genomic regions that have been swept in the recent past. Some genes associated with complex, economically important traits have been identified with the help of selected sweep analysis by whole-genome sequencing. We used the XP-EHH test to detect alleles near fixation within a Bamei mutton sheep population. Upon undergoing recent selection, the selected allele generally reaches a high frequency or fixation in one group, but remains polymorphic in the whole population (30). In this research, we discovered 155 regions selected for in the polytocous population, which was greater than the 43 regions screened in the monotocous population. Using fixed window selection of FST and XP-EHH, 221 candidate genes were found to be associated with the prolificacy trait. In a previous study using a segregated flock based on QTL and GWAS mapping, some mutations (FecB, FecX, and FecG) were identified to affect ovulation in sheep (11). However, here we did not identify any mutations previously reported to increase the number of ovulations in sheep. The litter size per breeding ewe is not only influenced by ovulation, but also affected by a number of factors including fertilization rate and pregnancy loss (in the embryonic and fetal development period). Recently, the LEPR gene, estrogen receptor 1 (ESR1) gene, and prolactin (PRL) gene were found to be associated with fecundity, as revealed by a selective sweep analysis in European commercial and semi-feral breeds and a Chinese indigenous breed distributed in different ecoregions (23, 31, 32).
JUN, ITPR3, and PLCB2 in the selected region were enriched in GnRH, oxytocin, and estrogen signaling pathway associated with the complex process from estrus to lambing. This process is organized by complex communication among the hypothalamus, pituitary, ovary, and uterus. Gonadotrophin releasing hormone (GnRH), secreted by the hypothalamus, regulates the synthesis and release of gonadotrophins, follicle-stimulating hormone (FSH), and luteinizing hormone (LH) from the pituitary (33). FSH and LH support the growth and maturation of follicles. Estradiol and progesterone secreted by the corpus luteum inhibit GnRH release by feedback modulation. During the follicular phase, following luteolysis, estradiol reaches a critical threshold and stimulates the preovulatory gonadotrophin surge, appearing to help the ovulation of mature follicles (11).
PLCB is involved in a wide range of signals of reproductive processes, being regulated by many hormones (FSH, LH, GnRH, oxytocin). PLCB2 as a member of the PLCB subfamily is activated by G-protein-linked receptors and can hydrolyze phosphatidylinositol 4,5-bisphosphate to form inositol-1,3,4-trisphosphate (IP3) and diacylglycerol (DAG), which stimulate Ca2+ release and protein kinase C activity, respectively (34). ITPR3 is a member of the IP3 receptor family. IP3, as an intracellular secondary messenger, mobilizes Ca2+ from endoplasmic reticulum stores, which transduces several calcium-dependent cascades. IP3 receptor downregulation induced by GnRH can suppress the secretion of LH/FSH (35, 36).
JUN is one of three JUN members [JUN (c-JUN), JUNB, and JUND] constituting the AP-1 family of heterodimeric transcription factors by combining with four FOS members [FOS (c-Fos), FOSB, FRA1, and FRA2]. GnRH can stimulate the expression of JUN by rapidly and transiently binding to the AP1 site in the FSHB promoter and then stimulating FSHB transcription (36). Conditional knockout of JUN in mice resulted in subfertility in both sexes, such as impaired spermatogenesis in males, diminished corpora lutea in females, and lower gonadal steroid (GnRH and LH) levels (37).
The nonsynonymous mutation p.S936A at KDM4B was discovered and confirmed in monotocous and polytocous sheep; it may be the reason for the signal of a selective sweep at the KDM4B locus. KDM4B belonging to the KDM4/JMJD2 family of histone demethylases contains a JmjN domain, JmjC domain, tandem plant homeodomains (PHD), and tandem Tudor domains. KDM4B catalyzes the demethylation of H3K9me3 and H3K9me2 at or near regulated promoters to promote expression of the downstream pathway induced by multiple different extracellular stimuli (38). KDM4B plays a central role in regulating the ER signaling cascade by controlling expression of the ER and FOXA1 genes. These two important genes can maintain the estrogen-dependent phenotype (39). In the developing ovarian follicle, granulosa cells are the main producers of estrogen. KDM4B is expressed in granulosa cells at early stages of folliculogenesis and its level is correlated with pregnancy failure in IVF patients (40). KDM4B expression in the uterus is also associated with recurrent pregnancy loss in women (41). The identified intersection between steroid hormones and KDM4B in the ovary and uterus sheds new light on the regulation of reproduction.