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MicroRNA Detection in Prostate Tumors by Quantitative Real-time PCR qPCR
Published on: May 16, 2012
MicroRNA-185 downregulates androgen receptor expression in the LNCaP prostate carcinoma cell line
Chunyan Liu1, Zhaobo Chen1, Xiaoyan Hu1
1Department of Biochemistry and Molecular Biology, Medical School of Shandong University, Jinan, Shandong 250012, P.R. China.
Abstract:
The present study aimed to investigate whether microRNA (miR)‑185 downregulated androgen receptor expression in the LNCaP prostate carcinoma cell line. Human prostate cancer (PCa) LNCaP cells were cultured and transfected with synthetic has‑miR‑185 mimic or inhibitor. The transfected cells were subsequently evaluated with a viability assay, nuclear staining, reverse transcription quantitative polymerase chain reaction (RT‑qPCR), dual luciferase assay and western blot analysis. The results of the western blot analysis and RT‑qPCR indicated that transfection with an miR‑185 mimic markedly reduced the androgen receptor (AR) protein expression levels in LNCaP cells, whereas transfection with an miR‑185 inhibitor increased the protein expression of AR in the LNCaP cells. The results of the luciferase reporter assay demonstrated that the predicted target site in the AR 3' untranslated regions was a specific functional binding site for miR‑185, and that AR was a direct target of miR‑185. In addition, downregulation of AR by miR‑185 impaired the interaction between AR and androgen response element, and downregulated the expression of the AR target gene prostate specific antigen. Data also suggested that the downregulation of AR mediated by miR‑185, inhibited the proliferation and induced the apoptosis of the LNCaP cells. Therefore, the results of the present study suggested that miR‑185 may be a potential negative modulator of AR‑mediated signaling and may act as a tumor suppressor in prostate cancer cells.
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