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Abstract: Prostaglandins (PGs) play a critical role in porcine reproduction, of which prostaglandin E2 (PGE2) and prostaglandin F2b1; (PGF2b1;) exert antiluteolytic and luteolysis actions, respectively. As a rate-limiting enzyme, bonyl reductase 1 (CbR1)%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>carbonyl reductase 1 (CbR1) catalyzes the conversion of PGE2 to PGF2b1;. A high ratio of PGE2:PGF2b1; is beneficial to the establishment and maintenance of porcine pregnancy. PG is essential for the establishment of pregnancy which resembles the proinflammatory response and nuclear factor bA;b (bA%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>NF-bA;b) is involved in the process. bioinformatic analysis has shown that bA%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>NF-bA;b is a possible factor bound to two cis-regulatory elements in CbR1 b%5D=abstract&ck%5b%5D=keyword'>b>promoter. In this study, we cloned the 2997 bp (−2875/+122) of the b%5D=abstract&ck%5b%5D=keyword'>b>promoter, and constructed six 5'-deleted dual-luciferase reporter recombinant vectors. In endometrial cells, the region of P2 (−1640/+7) exhibited the greatest transcriptional activity at driving luciferase expression, but not significantly different from that of P1 (−2089/+7). The activity of P1, P2, and P3 (−1019/+7) was highly significantly higher than that of others (P<0.01), suggesting that two positive regulatory elements were likely present in the regions of −1640/−1019 and −1019/−647. The results also showed that the −1640/−647 region was indispensable for the b%5D=abstract&ck%5b%5D=keyword'>b>promoter. The results of chromatin immunoprecipitation (ChIP) demonstrated that the bA%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>NF-bA;b subunit p65 binds to one site around −1545/−1531. Using four reference genes, we found that the over-expression of p65 enhanced the expression of CbR1 (P<0.05) in porcine endometrial epithelial cells, while knockdown of the p65 did not down-regulate the CbR1 expression. These results indicated that bA%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>NF-bA;b (p65) could bind to the special element of CbR1 gene b%5D=abstract&ck%5b%5D=keyword'>b>promoter in porcine endometrial epithelial cells in vitro. The binding site of bA%29&ck%5b%5D=abstract&ck%5b%5D=keyword'>b>NF-bA;b was a positive regulator for the CbR1 gene b%5D=abstract&ck%5b%5D=keyword'>b>promoter, but was not necessary for the basic expression.

CBR1基因启动子在猪子宫内膜细胞的功能研究

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