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Lin28B plays an important role in puberty initiation in sheep. This study aimed to discuss the correlation between different growth periods and the methylation status of cytosine–guanine dinucleotide (CpG) islands in the promoter region of the Lin28B gene in the Dolang sheep's hypothalamus. In this study, the sequence of the Lin28B gene promoter region in Dolang sheep was obtained by cloning and sequencing, and methyl groups of the CpG island of the Lin28B gene promoter in the hypothalamus were detected by bisulfite sequencing PCR during the three periods of prepuberty, adolescence, and postpuberty in Dolang sheep. Lin28B expression in the Dolang sheep's hypothalamus was detected by fluorescence quantitative PCR at three stages: prepuberty, puberty, and postpuberty. In this experiment, the 2993-bp Lin28B promoter region was obtained, and it was predicted that there was a CpG island containing 15 transcription factor binding sites and 12 CpG sites, which may play a role in gene expression regulation. Overall, methylation levels increased from prepuberty to postpuberty, while Lin28B expression levels decreased, indicating that Lin28B expression was negatively correlated with promoter methylation levels. Variance analysis showed significant differences in the methylation status of CpG5, CpG7, and CpG9 between pre- and postpuberty (p < 0.05). Our data show that Lin28B expression is increased by demethylation of promoter CpG islands, with CpG5, CpG7, and CpG9 implicated as critical regulatory sites. [ABSTRACT FROM AUTHOR]

Gonadotropin-releasing hormone (GnRH) is a key factor at the onset of puberty. This decapeptide has been found in mammalian ovaries, but its regulatory mechanism in the ovary of sheep at the onset of puberty is not clear. This study investigated the coding sequence (CDS) of the GnRH gene in the ovary of Duolang sheep and the expression of GnRH mRNA in different tissues at the onset of puberty, and analyzed the effect of GnRH on ovarian granulosa cells (GCs) of Duolang sheep. The results showed that the GnRH CDS of sheep was cloned, the full length of the GnRH CDS in sheep ovary was 279 bp, and the nucleotide sequence was completely homologous to that in the hypothalamus. The expression of GnRH mRNA was highest in the hypothalamus and ovary. The expression of related hormones and receptors in GCs of Duolang sheep treated with different concentrations of GnRH for 24 h was affected. GnRH significantly inhibited LH synthesis and LHR expression in GCs. Low concentration (100 ng mL -1) had the most obvious therapeutic effect on follicle-stimulating hormone (FSH) and FSHR. Higher concentration (250 ng mL -1) significantly promoted estradiol and ER β mRNA. These findings provide strong evidence that ovarian GnRH is an important regulatory factor at the onset of puberty in sheep. [ABSTRACT FROM AUTHOR]