Cloning and functional verification of a porcine adipose tissue-specific promoter

Abstract

Abstract Background Fat deposition is an important economic trait in pigs. In the past decades, many genes regulating porcine fat deposition were identified by Omics technology and verified by cell biology studies. Using genetically modified pigs to investigate the function of these genes in vivo is necessary before applying in breeding. However, lack of tissue-specific promoters of pigs hinders the generation of adipose tissue-specific genetically modified pigs. Results In order to identify a porcine adipose tissue-specific promoter, we used the software Digital Differential Display (DDD) to screen 99 genes highly expressed in porcine adipose tissue. GO and KEGG enrichment analysis indicated that the 99 genes were mainly related to lipid metabolism. Q-PCR proved that LGALS12 was an adipose tissue-specific gene. Five truncated fragments of the LGALS12 promoter were cloned and the 4 kb fragment (L-4 kb) exhibited a high level of promoter activity in adipocytes and no promoter activity in non-adipocytes. Following co-transfection with adipogenic transcription factors, the promoter activity of L-4 kb was enhanced by PPARγ, C/EBPβ, and KLF15, whereas it was suppressed by KLF4. Finally, we demonstrated that L-4 kb can drive APOR gene expression to exert its function in adipocytes. Conclusions This study demonstrates that porcine LGALS12 is an adipose tissue-specific gene, and identified the 4 kb fragment of LGALS12 promoter that exhibited adipocyte-specific promoter activity. These results provide new evidence for understanding porcine fat deposition and a promoter element for adipose tissue-specific genetic modification in pigs. Highlights Identified porcine LGALS12 as an adipose tissue-specific gene. Truncated LGALS12 promoter (L-4 kb) showed adipose tissue-specific promoter activity. Identified transcription factors involved in the regulation of L-4 kb promoter activity.