Methylation of CpG Island within Second Intron Increased G- protein Coupled Receptor 120 (GPR120) mRNA Transcripts in Porcine (Sus scrofa) Subcutaneous and Visceral Adipose Tissue
Abstract：G-protein coupled receptor 120 (GPR120) is the only highly expressed G protein-coupled receptor in adipose tissues, which plays a critical role in various physiological processes. DNA methylation is an important epigenetic modification to mediate tissue-specific gene expression that mostly occurs in CpG-rich regions (CpG islands). To study the effects of CpG island methylation for the expression levels of GPR120 in porcine (Sus scrofa)subcutaneous (backfat) and visceral (greater omentum) adipose tissues, this study used qRT-PCR to detect the relative expression levels of GPR120 in different adipose tissues of Jinhua pigs (6 months and 7 years old), using CpG island searcher online software to scan the CpG island distribution across the whole GPR120 DNA sequence (5 000 bp upstream of first exon to 5 000 bp downstream of last exon), and detected the methylation status of CpG island in different adipose tissues using Sequenom MassArray methylation approach, using bisulfite-sequencing PCR (BSP) to validate the MassArray results. Results showed that compared with greater omentum, subcutaneous adipose tissue had a higher adipocyte volume both at 6 months and 7 years (P＜0.01), and the relative transcript levels of GPR120 significantly higher expressed in subcutaneous adipose tissue than that of greater omentum (P＜0.01), which was consistent with adipocyte volume differences. The DNA sequence of GPR120 had abundant GC contents, and contained five CpG islands which were predicted by bioinformatics analysis successively located within different genomic locations, that were the 5' untranslated region, first exon, intron and 3' untranslated region. The methylation status of CpG island within second intron in subcutaneous adipose tissue was significantly higher than that of greater omentum (P＜0.01) both at 6 months and 7 years, which was confirmed by the BSP approach, while the methylation levels of other 4 islands had no significant difference between subcutaneous adipose tissue and greater omentum at the two time points. There was a positively correlation between the changes in methylation levels of CpG island within second intron and gene expression. This study presented an integrated analysis of gene-wide DNA methylation patterns in different tissues, and results indicated that GPR120 had abundant CpG islands, the tissue-specific different methylation of CpG island in gene body positively associated with GPR120 expression between subcutaneous adipose tissue and greater omentum. This comprehensive analysis provides a solid basis for exploring epigenetic mechanisms of GPR120 differentially expressed in tissues.
