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

• Published in: Journal of Zhejiang University. B. Science 2022-06, Vol.23 (6), p.502-514
• Main Authors: Zhang, Weidi, Ren, Wenzhi, Han, Dongxu, Zhao, Guokun, Wang, Haoqi, Guo, Haixiang, Zheng, Yi, Ji, Zhonghao, Gao, Wei, Yuan, Bao
• Format: Article
• Language: English
• Subjects: Animals; Biomedical and Life Sciences; Biomedicine; Cell Line, Tumor; Cell Proliferation; Cytoplasm; Enzyme-linked immunosorbent assay; Fluorescence; Fluorescence in situ hybridization; Follicle Stimulating Hormone - genetics; Follicle Stimulating Hormone - metabolism; Follicle-stimulating hormone; Follicles; Gene expression; Gene Expression Regulation, Neoplastic; Homology; Immunoprecipitation; In Situ Hybridization, Fluorescence; Localization; MicroRNAs; MicroRNAs - genetics; MicroRNAs - metabolism; miRNA; Non-coding RNA; Pituitary; Polymerase chain reaction; Rats; Research Article; Reverse transcription; Ribonucleic acid; RNA; RNA, Long Noncoding - genetics; RNA, Long Noncoding - metabolism; Smad2 protein
• ISSN: 1673-1581; 1862-1783
• DOI: 10.1631/jzus.B2101052

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.