Abstract: 【Objective】To interpret partially the role of FAM213B gene expression in prostaglandin synthetise and sow pregnancy through the identification of the transcription region of porcine FAM213B gene promoter and the effection of NFκB on the promoter. 【Method】 The porcine endometrium from the uterus in follicular phase was digested by collagenase and the isolated endometrial cells were cultured for the detection of the FAM213B promoter activity. Based on the mRNA and promoter sequences of FAM213B gene obtained in our former work (GenBank ID: KX444503 and 100134955), the longer 5′ regulationary sequence was amplified and sequenced. Then, seven promoter fragments with 5′ terminal deletion, containing Mlu I and Xho I sites, were linked into the Dual-luciferase Reporter vectors. Seven constructed vectors treated by endotoxin free and pRL-TK plasmid were co-transfected into endometrial cells through liposome method. The core region of transcriptional activity of the gene promoter was identified through the Dual-luciferase Reporter Assay System. The putative transcription factors binding sites were analyzed by bioinformatics, and the binding of NFκB with FAM213B promoter were detected by ChIP (Chromatin immunoprecipitation). The over-expression vectors of NFκB1and RelA and interference fragments of their own were transfected into endometrial cells. Then, the transcription activity of the promoter and mRNA expression of the gene were detected by the Dual-luciferase Reporter system and fluorescent quantitative, respectively. 【Result】 Through the PCR and sequencing, one fragment of 2 261 bp (-2178/+83) of porcine FAM213Bgene were obtained. The bioinformatics analysis showed that there were putative binding sites of CREB, CCAAT, E-box, and NFκB in the promoter. And the putative binding sites of NFκB were found in -1143/-1132 and -664/-655 regions. The result of the Dual-luciferase Reporter showed the region of P2 (-1352/+30) exhibited the strongest transcriptional activity, and it was significantly higher than that of P1 (-1760/+30) (P＜0.01), which showed there were negative regulation elements in -1760/-1352 region. And the transcriptional activity of P2 (-1352/+30) was significantly higher than that of P3(-919/+30) (P＜0.05), implying the existence of positive elements in -1352/- 919 region. The significant stronger activity of P3 (-919/+30) than P4(-604/+80) (P＜0.01) meant the existence of positive elements in -919/-604 region. No significant differences were observed between P4, P5, P6, and P7. The region of -1352/-919 was the core element of the promoter. The results of Chromatin Immunoprecipitation (ChIP) demonstrated that NFκB1 binds to one site around -1143/-1132, and RelA site around -664/-655. The co-transfection of the over expression of pcDNA3.1-NFκB1 and P2 vector into endometrial cells increased the activity of the promoter (P＜0.01), and the transfection of pcDNA3.1-NFκB1 enhanced the mRNA expression of FAM213B (P＜0.05). While the co-transfection of the over expression of pcDNA3.1-RelA and P3 vector into endometrial cells decreased the activity of the promoter (P＜0.05), and the transfection of pcDNA3.1-RelA weakened the mRNA expression of the gene. At the same time, the transfection of inference siRNA fragments of NFκB1 and RelA into endometrial cells led the contrary results for the activity of the promoter and the mRNA expression of the gene.【Conclusion】The core region of porcine FAM213B gene promoter was identified round -1352/-919. NFκB was the trancription factor of FAM213B gene. NFκB1 and RelA, two members of NFκB family, regulate the expression of FAM213B gene in endometrial cells.

Key words: pig, FAM213B gene, promoter, NFκB, endometrial cells