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目的研究甾体类激素受体AR、ER对前列腺癌L-Plastin基因的调控作用,确定甾体类激素对前列腺癌的调节作用。方法构建L-plastin启动子于含荧光素酶的pGL3质粒,利用PCR定点突变法依次构建切除ARE1、ARE2、ARE3、ERE序列的pGL3重组子,将重组子转染到前列腺癌细胞系LNCaP,检测荧光素酶强度,以了解ARE1、ARE2、ARE3、ER调控L-plastin表达能力。结果成功构建了L-plastin启动子的荧光素酶pGL3重组子以及切除ARE1、ARE2、ARE3、ERE位点的重组子,将这些重组子转染细胞后其荧光素酶强度有明显改变,切除ARE1位点后,荧光素酶活性下降1/3;切除ARE2后,荧光素酶活性与单纯切除ARE1相比下降1/2;切除ARE3后荧光素酶活性升高1倍;切除ERE后,荧光素酶活性下降约4/5。结论在L-plastin启动子转录活性中ARE1、ARE2、ARE3、ERE在前列腺癌中对其下游基因L-plastin起到调控作用,ARE1、ARE2、ERE起促进作用,ARE3起抑制作用。而且,L-plastin启动子上存在其他受甾体类激素调控的位点。
Objective To investigate the regulatory effect of steroid hormone receptors AR and ER on L-Plastin gene in prostate cancer, and to determine the regulatory effect of steroid hormones on prostate cancer. Methods The L-plastin promoter was constructed in plasmid pGL3 containing luciferase. The pGL3 recombinant with ARE1, ARE2, ARE3 and ERE sequences was constructed by PCR and the recombinant plasmid was transfected into LNCaP cells. Luciferase intensity to understand ARE1, ARE2, ARE3, ER regulation of L-plastin expression. Results The L-plastin promoter luciferase pGL3 recombinant and the excision of the ARE1, ARE2, ARE3 and ERE loci were successfully constructed. The luciferase activity of these recombinant plasmids was significantly changed after transfection of these recombinant plasmids. The ARE1 , The luciferase activity decreased by 1/3; after the removal of ARE2, the luciferase activity decreased by 1/2 compared with that of ARE1 alone; the luciferase activity was doubled after the excision of ARE3; Enzyme activity decreased by about 4/5. Conclusions ARE1, ARE2, ARE3 and ERE play roles in the regulation of L-plastin, ARE1, ARE2 and ERE in prostate cancer and ARE3 in the transcriptional activity of L-plastin. Moreover, there are other steroid hormone-regulated sites on the L-plastin promoter.