The transcription factor ccaat/enhancer binding protein β (C/EBPβ) and miR-27a regulate the expression of porcine Dickkopf2 (DKK2).

Mammalian folliculogenesis is a complex process through which primordial follicles develop into pre-ovulatory follicles. It is followed by ovulation, which releases mature oocytes1. After ovulation, the remaining follicular structure undergoes luteinization, and the former granulosa and thecal cells are transformed into follicular and thecal lutein cells. The complexity of folliculogenesis indicates that tightly regulated gene expression and interactions between many genes are required for successful oocyte development. Approximately 100 genes have been shown to be essential for normal folliculogenesis in mice in knock-out experiments2. WNT signaling proteins have been shown to play crucial roles in reproductive processes, including foetal development, ovarian development, gestation and mammogenesis3,4,5. Six WNT/β-catenin signaling pathway genes, including the wingless-type MMTV integration site family member 10B (WNT10B) and DKK2, have differential expression patterns in PMSG-hCG-stimulated pre-ovulatory ovarian follicles in three Chinese Taihu and three Large White cycling sows, as shown by Affymetrix Porcine GeneChip™ analyses6. DKK2 is a direct inhibitor of WNT binding to LDL receptor-related proteins 5/6 (LRP5/6), which are co-receptors of frizzled7,8. The role of DKK2 in tumourigenesis and WNT signaling has been partially described9,10,11, but no direct evidence for its functions in follicle development have yet been reported. After studying the literature, we hypothesized that DKK2 functions may be associated with embryo implantation and endometrial membrane stripping8. Recent studies have shown that miRNAs functions in cell invasion and tumourigenesis involve the WNT/β-catenin signaling pathway, including DKK2. For example, miR-21 promotes oral tongue squamous cell carcinoma (OTSCC) invasion via the WNT/β-catenin pathway by targeting DKK2 in vitro12. MiR-1260b binds to the 3′ untranslated regions (UTRs) of the DKK2, secreted frizzled-related protein 1 (sFRP1) and SMAD family member 4 (Smad4) mRNAs, thereby down-regulating their expression in renal cancer cells11. MiR-222 promotes tumourigenesis by targeting DKK2 and activating the WNT/β-catenin signaling pathway10. The objective of this study was to examine mutations and cis regulatory elements in the porcine DKK2 gene. We therefore analysed the 5′ upstream sequences and 3′ UTR of DKK2 to better understand its role.