Profiling of direct mRNA targets of miR-130a, miR-203 and miR-205 in prostate cancer cell line LNCaP

Micro RNAs (miRNAs) miR-130a, miR-203 and miR-205 are jointly downregulated in prostate cancer and act as repressors of AR-signaling. MiRNAs are small non-coding RNAs that regulate the expression of specific mRNA targets mainly by translational repression, mRNA deadenylation or cleavage. Reconstitution of these lost miRNAs in the LNCaP PCa cell line cause morphology changes, growth arrest, and apoptosis, increasing when the miRNAs were co-expressed. This series identifies direct targets of miR-130a, miR-203, and miR-205 by AGO2-RNA co-immunoprecipitation as described by (Beitzinger et al. 2007) upon miRNA reconstitution in LNCaP cells and analyzing AGO2-bound mRNAs using Affymetrix Genechips. Relative levels of AGO2 bound versus total RNA expression were compared between miRNA reconstituted and miR-scr transfected samples. Three arrays each for AGO2-bound RNA upon reconstitution of miR-130a, miR-203, miR-205, a scramble miRNA, and three arrays each for total RNA upon reconstitution of miR-130a, miR-203, miR-205, a scramble miRNA.

微小RNA（microRNAs, miRNAs）miR-130a、miR-203与miR-205在前列腺癌中共同下调，并作为雄激素受体（Androgen Receptor, AR）信号通路的抑制因子发挥功能。miRNAs是一类小型非编码RNA，主要通过翻译抑制、mRNA脱腺苷化或裂解作用调控特定靶mRNA的表达。将这些缺失的miRNAs重新导入LNCaP前列腺癌（prostate cancer, PCa）细胞系后，可引发细胞形态改变、生长停滞与凋亡，且共表达这些miRNAs时上述效应会增强。本数据集通过AGO2（Argonaute 2）-RNA免疫共沉淀（AGO2-RNA co-immunoprecipitation）技术，参照Beitzinger等人2007年的研究方法，在LNCaP细胞中重新导入miRNAs后，利用Affymetrix基因芯片（Affymetrix Genechips）分析AGO2结合的mRNA，从而鉴定miR-130a、miR-203与miR-205的直接靶标。研究比较了miRNAs重新导入组与乱序对照miRNA（miR-scr）转染组中，AGO2结合RNA与总RNA表达的相对水平。针对miR-130a、miR-203、miR-205及乱序对照miRNA的重新导入，AGO2结合RNA样本各进行3次芯片检测；针对上述各组的总RNA样本，同样各进行3次芯片检测。