Lysophosphatidic acid receptor-2 (LPAR2) is a G protein-coupled receptor [1]. G-protein-coupled receptors (GPCRs) are a family of membrane receptors with a conserved pattern of seven transmembrane alpha helices that transmit extracellular signals into cells [2,3]. To date, six LPA receptors (LPAR1 to LPAR6) have been identified, all of which can connect to downstream cellular messaging networks [4,5]. Triggered by extracellular ligand binding, GPCRs activate isomeric G proteins within the cell, thereby inducing downstream signaling cascades. GPCR ligands include amines, peptides, nucleic acids, and lipids that induce various cellular responses, including survival, proliferation, differentiation, and migration [6]. LPARs can activate different downstream signaling pathways through different distributions in organs and tissues and bind to different heterotrimeric G proteins, resulting in gene regulation and LPA-induced changes in cell function [7]. These molecules play an important role in regulating various physiological phenomena. Recent studies have shown that LPAR2 is involved in the reproductive physiological processes of various animals. LPAR2 is the main receptor responsible for the effects of LPA in ovarian endometriotic cysts [8]. A similar abundance of LPAR1-4 transcripts was found in bovine cumulus cells, where LPAR2 was expressed at higher levels than the other three LPARs [9]. Furthermore, LPAR2 transcripts were found in trophectoderm extracts of sheep embryos during early pregnancy [10]. The above studies indicate that LPAR2 might play an important role in the reproductive physiology of various mammals.

The litter size is an important economic trait in sheep breeding that is regulated by many factors and is a complex quantitative trait. Many studies have shown that molecular marker-assisted selection is an important technical means to accelerate the breeding process of livestock [11,12]. The major genes controlling litter size have been successively identified in many sheep breeds worldwide [13]. In two Egyptian sheep breeds, genetic polymorphism in growth differentiation factor 9 (GDF9) was found to be significantly associated with litter size in both breeds via marker-assisted selection analysis [14]. Several polymorphisms of IGF1 were found in specific loci in three breeds of sheep, small-tailed Han sheep, Hu sheep, and Dorset sheep, and are important for the selection and breeding of small-tailed Han sheep [15]. With the maturation of high-throughput sequencing technology, molecular marker-assisted selection has been widely used in sheep breeds worldwide. Therefore, identifying the genes with the main effect and finding SNP sites significantly related to reproductive phenotypes are the focus of our research on sheep reproduction. Exploring the molecular mechanism of related genes and the interaction network with other key factors can accelerate the process of sheep breeding [12]. Ovarian granulosa cells are an important part of the follicle and play an important role in the female reproductive process [16]. Granulosa cells provide nutritional support for oocyte development and maturation through gap junctions and paracrine factors [17]. Granulosa cell damage was reported to affect hormone and cytokine secretion and reduce oocyte quality and embryo developmental potential [18]. In mammals, LPA induces its effects through the best-known and most characteristic types of high-affinity G protein-coupled receptors, including LPAR1/EDG2, LPAR2/EDG4, and LPAR3/EDG7 [19,20]. Due to the heterogeneity of receptor subtypes, expression patterns, and effector pathways, the roles of LPA are diverse and extensive, as LPA regulates many biological processes, including cell-cell interactions [21], tumorigenesis [22], cell proliferation, differentiation [23,24], and migration [25]. In bovine ovaries, LPA stimulates E2 synthesis and the MAPK/ERK pathways in bovine follicular granulosa cells via LPAR2 [26,27]. In addition, the highest mRNA levels of LPAR2 were found in COCs isolated from bovine ovarian follicles, which may indicate that LPAR2 plays the most important role in LPAR2-mediated signaling in bovine ovarian follicles [9]. In an ovarian transcriptome analysis of white Muscovy ducks with low egg production, LPAR2 was identified as a potential candidate gene associated with egg production [28]. These findings suggest that LPAR2 may play a key role in the development of GCs in sheep, which in turn affects ovulation and litter size.

The 3′UTR of eukaryotic mRNA plays an important role in the post-transcriptional regulation of gene expression, including the regulation of mRNA transport, mRNA stability, translation efficiency, and subcellular localization [29]. Increasing evidence has shown that polymorphisms in miRNA binding sites can affect the binding of miRNAs to target genes, resulting in alterations in target gene expression and phenotype [30]. In this study, we investigated the potential mechanism by which the SNP c.*701C > T of LPAR2 affects the number of lambs. The results identified the SNP c.*701C > T in the 3′UTR of LPAR2, and this mutation led to specific binding of miR-939-5p. Further studies revealed that miR-939-5p regulated the apoptosis of sheep granulosa cells with the CC genotype of LPAR2.