Characterization and transcriptional regulation analysis of the porcine TNFAIP8L2 gene

Abstract

Tumor necrosis factor alpha-induced protein 8-like 2 (TNFAIP8L2) is a new member of the tumor necrosis factor-alpha-induced protein 8 (TNFAIP8) family that functions as an important factor in the maintenance of immune homeostasis. In this study, we cloned the cDNA sequences and analyzed the genomic structure of porcine TNFAIP8L2. RH mapping using the IMpRH panel showed that this gene was closely linked to microsatellite marker SW512 in pig chromosome 4. Subcellular localization analysis showed GFP-TNFAIP8L2 fusion protein distributed in nucleus as well as cytoplasm including mitochondria and endoplasmic reticulum. Real-time PCR analysis revealed that porcine TNFAIP8L2 was more highly expressed in spleen than other tissues. To understand its characterization of transcriptional regulation, we cloned approximately 2 kb of 5'-regulatory region upstream to the porcine TNFAIP8L2 translational start site and generated sequential deletion constructs evaluated in dual-luciferase reporter assay. The results demonstrated that its core promoter is 435 base pairs (bp) upstream to the transcription initiation site. Then, site-directed mutation experiment combined with electrophoretic mobility shift assay (EMSA) indicated that M-CAT binding factor (MCBF) and activator protein 1 (AP-1) were important transcription factors for porcine TNFAIP8L2. These findings provide an important basis for further understanding of porcine TNFAIP8L2 regulation and function in swine.