CCAAT/enhancer binding protein-beta negatively regulates the expression of glycerol-3-phosphate dehydrogenase 1 in pig PK-15 cells

Soluble glycerol-3-phosphate dehydrogenase 1 (GPD1, EC 1.1.1.8) plays important roles in the synthesis of triacylglycerol and in the glycerol-3-phosphate shutter. Though GPD1 is expressed in most adult tissues, little is known about the regulation of its expression. In this study, we analyzed the characters, organization and core region of the promoter of pig GPD1 gene by in silico analysis and activity detection of deletion mutants. We also identified and testified the negative regulation effect of C/EBP β on pig GPD1 gene by Chromatin immunoprecipitation (ChIP) assay and over-expression experiments in cultured pig kidney cells. Compared to that of human, pig GPD1 gene promoter has three conserved regions and one deletion region. In silico analysis indicated that pig GPD1 promoter was TATA-less with at least 3 CpG islands of over 200 bp in length and over 60% in GC content. The activity detection of deletion mutants suggested that the essential elements required for the optimal promoter activity scatter in the promoter region, while the core promoter region was from -422 bp to -1 bp. Chromatin immunoprecipitation (ChIP) assay results indicated that C/EBP β had plenty of binding sites in pig GPD1 promoter with the common cis-element (5’- TKNNGCAAK -3’). The over-expression examination of C/EBP β showed that the expression of GPD1 was negatively regulated by C/EBP β in pig kidney cells. Overall, our study revealed that the pig GPD1 promoter is a TATA-less promoter, and in promoter region, the binding sites of C/EBP β share common motif of (5’-TKNNGCAAK -3’). We also showed that pig GPD1 gene is regulated negatively by C/EBP β in cultured kidney cells.