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CLC number: S828.2

On-line Access: 2017-07-05

Received: 2016-05-18

Revision Accepted: 2016-09-12

Crosschecked: 2017-06-16

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Citations:  Bibtex RefMan EndNote GB/T7714

 ORCID:

Hao Zhang

http://orcid.org/0000-0002-9455-3700

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Journal of Zhejiang University SCIENCE B 2017 Vol.18 No.7 P.626-634

http://doi.org/10.1631/jzus.B1600225


Functional characterization of the promoter of carbonyl reductase 1 gene in porcine endometrial cells


Author(s):  Ai-ling Zhang, Xian-yue Sun, Qi Yin, Jian-hua Zeng, Zhe Zhang, Jia-qi Li, Hao Zhang

Affiliation(s):  Guangdong Development Center of Applied Ecology and Ecological Engineering in Universities, Biology and Food Engineering Institute, Guangdong University of Education, Guangzhou 510310, China; more

Corresponding email(s):   zhangmeixial@163.com, zhanghao@scau.edu.cn

Key Words:  Pig, Carbonyl reductase 1 (CBR1), Promoter, NF-κ, B, Endometrium


Ai-ling Zhang, Xian-yue Sun, Qi Yin, Jian-hua Zeng, Zhe Zhang, Jia-qi Li, Hao Zhang. Functional characterization of the promoter of carbonyl reductase 1 gene in porcine endometrial cells[J]. Journal of Zhejiang University Science B, 2017, 18(7): 626-634.

@article{title="Functional characterization of the promoter of carbonyl reductase 1 gene in porcine endometrial cells",
author="Ai-ling Zhang, Xian-yue Sun, Qi Yin, Jian-hua Zeng, Zhe Zhang, Jia-qi Li, Hao Zhang",
journal="Journal of Zhejiang University Science B",
volume="18",
number="7",
pages="626-634",
year="2017",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B1600225"
}

%0 Journal Article
%T Functional characterization of the promoter of carbonyl reductase 1 gene in porcine endometrial cells
%A Ai-ling Zhang
%A Xian-yue Sun
%A Qi Yin
%A Jian-hua Zeng
%A Zhe Zhang
%A Jia-qi Li
%A Hao Zhang
%J Journal of Zhejiang University SCIENCE B
%V 18
%N 7
%P 626-634
%@ 1673-1581
%D 2017
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B1600225

TY - JOUR
T1 - Functional characterization of the promoter of carbonyl reductase 1 gene in porcine endometrial cells
A1 - Ai-ling Zhang
A1 - Xian-yue Sun
A1 - Qi Yin
A1 - Jian-hua Zeng
A1 - Zhe Zhang
A1 - Jia-qi Li
A1 - Hao Zhang
J0 - Journal of Zhejiang University Science B
VL - 18
IS - 7
SP - 626
EP - 634
%@ 1673-1581
Y1 - 2017
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B1600225


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

目的:研究CBR1基因启动子在猪子宫内膜细胞的表达调控机制。
创新点:发现CBR1基因启动子在猪子宫内膜受到炎性因子核转录因子kappa B(NF-κB)成员p65调控。p65对该启动子具有正向调节作用,但是对于CBR1基因的表达并不是必需的。
方法:通过双荧光素酶报告基因载体确定CBR1基因启动子转录活性区,通过染色质免疫沉淀(ChIP)技术确定p65能够结合CBR1基因启动子,通过超表达和干扰表达实验证实p65对CBR1基因启动子的调控作用。
结论:CBR1基因启动子−1640/−647区对于其转录活性是必需的,在−1545/−1531区存在p65的结合位点。p65在猪子宫内膜细胞中促进了CBR1基因mRNA的表达,但是干扰p65则不会造成CBR1基因mRNA表达量下降,推断p65不是CBR1基因表达的必需因素。

关键词:猪;CBR1基因;启动子;核转录因子kappa B(NF-κB);子宫内膜

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

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