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On-line Access: 2022-06-08

Received: 2021-12-13

Revision Accepted: 2022-02-04

Crosschecked: 2022-06-08

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Citations:  Bibtex RefMan EndNote GB/T7714

 ORCID:

Weidi ZHANG

https://orcid.org/0000-0002-6047-4916

Wenzhi REN

https://orcid.org/0000-0001-7052-4967

Wei GAO

https://orcid.org/0000-0001-8093-0555

Bao YUAN

https://orcid.org/0000-0003-3490-0755

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Journal of Zhejiang University SCIENCE B 2022 Vol.23 No.6 P.502-514

http://doi.org/10.1631/jzus.B2101052


LncRNA-m18as1 competitively binds with miR-18a-5p to regulate follicle-stimulating hormone secretion through the Smad2/3 pathway in rat primary pituitary cells


Author(s):  Weidi ZHANG, Wenzhi REN, Dongxu HAN, Guokun ZHAO, Haoqi WANG, Haixiang GUO, Yi ZHENG, Zhonghao JI, Wei GAO, Bao YUAN

Affiliation(s):  Department of Laboratory Animals, College of Animal Sciences, Jilin University, Changchun 130062, China; more

Corresponding email(s):   gaowei81@jlu.edu.cn, yuan_bao@jlu.edu.cn

Key Words:  Long noncoding RNA (lncRNA), MicroRNA (miRNA), Competitive endogenous RNA (ceRNA), Follicle-stimulating hormone (FSH), Mothers against decapentaplegic homolog 2/3 (Smad2/3)


Weidi ZHANG, Wenzhi REN, Dongxu HAN, Guokun ZHAO, Haoqi WANG, Haixiang GUO, Yi ZHENG, Zhonghao JI, Wei GAO, Bao YUAN. LncRNA-m18as1 competitively binds with miR-18a-5p to regulate follicle-stimulating hormone secretion through the Smad2/3 pathway in rat primary pituitary cells[J]. Journal of Zhejiang University Science B, 2022, 23(6): 502-514.

@article{title="LncRNA-m18as1 competitively binds with miR-18a-5p to regulate follicle-stimulating hormone secretion through the Smad2/3 pathway in rat primary pituitary cells",
author="Weidi ZHANG, Wenzhi REN, Dongxu HAN, Guokun ZHAO, Haoqi WANG, Haixiang GUO, Yi ZHENG, Zhonghao JI, Wei GAO, Bao YUAN",
journal="Journal of Zhejiang University Science B",
volume="23",
number="6",
pages="502-514",
year="2022",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B2101052"
}

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%T LncRNA-m18as1 competitively binds with miR-18a-5p to regulate follicle-stimulating hormone secretion through the Smad2/3 pathway in rat primary pituitary cells
%A Weidi ZHANG
%A Wenzhi REN
%A Dongxu HAN
%A Guokun ZHAO
%A Haoqi WANG
%A Haixiang GUO
%A Yi ZHENG
%A Zhonghao JI
%A Wei GAO
%A Bao YUAN
%J Journal of Zhejiang University SCIENCE B
%V 23
%N 6
%P 502-514
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%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B2101052

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T1 - LncRNA-m18as1 competitively binds with miR-18a-5p to regulate follicle-stimulating hormone secretion through the Smad2/3 pathway in rat primary pituitary cells
A1 - Weidi ZHANG
A1 - Wenzhi REN
A1 - Dongxu HAN
A1 - Guokun ZHAO
A1 - Haoqi WANG
A1 - Haixiang GUO
A1 - Yi ZHENG
A1 - Zhonghao JI
A1 - Wei GAO
A1 - Bao YUAN
J0 - Journal of Zhejiang University Science B
VL - 23
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SP - 502
EP - 514
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PB - Zhejiang University Press & Springer
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DOI - 10.1631/jzus.B2101052


Abstract: 
Long noncoding RNAs (lncRNAs) are expressed in different species and different tissues, and perform different functions, but little is known about their involvement in the synthesis or secretion of follicle-stimulating hormone (FSH). In general, we have revealed lncRNA‍‒‍microRNA (miRNA)‍‒‍‍messenger RNA (mRNA) interactions that may play important roles in rat primary pituitary cells. In this study, a new lncRNA was identified for the first time. First, we analyzed the gene expression of lncRNA-m18as1 in different tissues and different stages by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and observed the localization of lncRNA-m18as1 with fluorescence in situ hybridization, which indicated that this lncRNA was distributed mainly in the cytoplasm. Next, we used RT-qPCR and enzyme-linked immunosorbent assay (ELISA) to analyze the regulation of FSH synthesis and secretion after overexpression or knockdown of lncRNA-m18as1 and found that lncRNA-m18as1 was positively correlated with FSH synthesis and secretion. In addition, mothers against decapentaplegic homolog 2 (Smad2) was highly expressed in our sequencing results. We also screened miR-18a-5p from our sequencing results as a miRNA that may bind to lncRNA-m18as1 and Smad2. We used RNA immunoprecipitation-qPCR (RIP-qPCR) and/or dual luciferase assays to confirm that lncRNA-m18as1 interacted with miR-18a-5p and miR-18a-5p interacted with Smad2. Fluorescence in situ hybridization (FISH) showed that lncRNA-m18as1 and miR-18a-5p were localized mainly in the cytoplasm. Finally, we determined the relationship among lncRNA-m18as1, miR-18a-5p, and the Smad2/3 pathway. Overall, we found that lncRNA-m18as1 acts as a molecular sponge of miR-18a-5p to regulate the synthesis and secretion of FSH through the Smad2/3 pathway.

LncRNA-m18as1竞争性结合miR-18a-5p通过Smad2/3通路调控大鼠原代腺垂体细胞FSH分泌

张卫迪1,任文陟2,韩东旭1,赵国坤1,王皓琪1,郭海祥1,郑毅1,冀中豪1,高巍1,袁宝2
1吉林大学动物科学学院,实验动物系,中国长春市,130062
2吉林大学吉林省模型动物工程研究中心,中国长春市,130062
目的:探索长链非编码RNA(lncRNA)-微小RNA(miRNA)-信使RNA(mRNA)网络对促卵泡素(FSH)合成与分泌的作用,为调控FSH的分子机制提供理论基础。
创新点:筛选出了一个新的lncRNA作为研究对象;首次鉴定并分析了该lncRNA对FSH合成与分泌的调控作用;确定了该lncRNA调控FSH合成与分泌的机制。
方法:我们通过逆转录定量聚合酶链反应(RT-qPCR)筛选出了一个新的lncRNA,并根据功能将其最终命名为lncRNA-m18as1。经过敲降或过表达lncRNA-m18as1后,我们采用RT-qPCR与酶联免疫吸附剂测定(ELISA)分析了lncRNA-m18as1对FshβmRNA以及FSH分泌的调控作用。我们预测并确定了lncRNA-m18as1发挥作用的lncRNAm18as1/miR-18a-5p/Smad2轴。我们使用RNA结合蛋白免疫沉淀测定-逆转录定量聚合酶链反应(RIP-qPCR)和/或双荧光素酶报告分析方法分析了miR-18a-5p与lncRNA-m18as1、Smad2的靶向关系。此外,我们使用RT-qPCR和蛋白质印迹法(western blot)分析了lncRNA-m18as1和miR-18a-5p在轴中对向下游因子的调控作用,同时通过荧光原位杂交技术(FISH)观察了lncRNA-m18as1以及miR-18a-5p在细胞核与细胞质中的分布。
结论:研究结果表明在大鼠腺垂体中高表达且阶段性表达的lncRNA-m18as1促进FshβmRNA的表达与FSH的分泌。进一步的分子机制研究表明lncRNA-m18as1竞争性结合miR-18a-5p调控Smad2蛋白的表达,进而调控FSH的合成与分泌。

关键词:长链非编码RNA(lncRNA);微小RNA(miRNA);竞争性内源RNA(ceRNA);促卵泡素(FSH);Smad家族成员2/3(Smad2/3)

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

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