miR-489-3p靶向负调控BDNF并抑制PC12细胞增殖

miR-489-3p negatively regulates BDNF expression and inhibits PC12 cell proliferation

目的:探究微小RNA-489-3p(miR-489-3p)对神经生长因子家族重要成员脑源性神经营养因子(BDNF)的转录后调控及对PC12细胞增殖能力的影响。方法:通过生物信息预测和双萤光素酶报告基因实验检测miR-489-3p与BDNF的结合能力。PC12细胞分别转染mimic control/miR-489-3p mimic和inhibitor control/miR-489-3p inhibitor,分别采用RT-qPCR、Western blot、CCK-8和EdU实验检测miR-489-3p和BDNF的表达及细胞增殖能力的变化。结果:生物信息分析和双萤光素酶报告基因实验结果显示,miR-489-3p能够结合BDNF的3′-UTR。转染miR-489-3p mimic可以显著提高PC12细胞的miR-489-3p水平,而BDNF的蛋白表达显著下调,细胞增殖能力显著降低(P<0.05);转染miR-489-3p inhibitor可以显著降低PC12细胞的miR-489-3p水平,而BDNF的蛋白表达显著上调,并且显著提高了细胞的增殖能力(P<0.05)。过表达或抑制miR-489-3p对BDNF的mRNA表达并无显著影响。结论:miR-489-3p可以在转录后水平靶向负调控BDNF的表达。miR-489-3p抑制PC12细胞的增殖。

AIM: To explore the post-transcriptionally regulatory effect of microRNA-489-3 p(miR-489-3 p) on brain-derived neurotrophic factor(BDNF) expression, and its effect on the proliferation of PC12 cells. METHODS: The binding capacity between miR-489-3 p and BDNF was detected by bioinformatic prediction and dual-luciferase reporter assay. The PC12 cells were transfected with mimic control/miR-489-3 p mimic or inhibitor control/anti-miR-489-3 p inhibitor. RT-qPCR, Western blot, CCK-8 assay and EdU staining were used to detect the expression of miR-489-3 p and BDNF and the cell proliferation. RESULTS: The results of bioinformatic analysis and dual-luciferase reporter assay indicated that miR-489-3 p bound to the 3′-UTR of BDNF. miR-489-3 p was significantly increased by transfection of miR-489-3 p mimic, while the protein expression of BDNF was significantly down-regulated, and the cell proliferation was significantly decreased(P<0.05). The miR-489-3 p level was significantly reduced by transfection with miR-489-3 p inhibitor, while the protein expression of BDNF was significantly up-regulated and the proliferation of the PC12 cells was significantly improved(P<0.05). However, the mRNA expression of BDNF was not significantly affected by over-expression or inhibition of miR-489-3 p. CONCLUSION: miR-489-3 p negatively regulates the expression of BDNF at the post-transcriptional level and inhibits the proliferation of PC12 cells, thus providing a new molecular mechanism for neurologic diseases caused by down-regulation of BDNF expression.