GMP & T Cell Therapy Unit, German Cancer Research Center (DKFZ), 210, Im Neuenheimer Feld 280, D-69120, Heidelberg, Germany.
Faculty of Biosciences, University Heidelberg, Heidelberg, Germany.
Mol Cell Biochem. 2023 Feb;478(2):305-315. doi: 10.1007/s11010-022-04507-z. Epub 2022 Jul 2.
The transcription factor SOX9 represents an important mediator of breast cancer progression. miRNAs are small non-coding RNAs inhibiting translation of target genes upon interaction with the 3'-UTR region of respective mRNA molecules. Deregulated miRNA expression is involved in hallmarks of cancer like sustained proliferation and inhibition of apoptosis. Here, we investigated the miRNA-mediated regulation of SOX9 expression in two breast cancer cell lines, thereby providing further insights into cellular mechanisms driving breast cancer progression. The modulating effects of miR-134-3p, miR-224-3p, and miR-6859-3p on SOX9 expression were analyzed by qPCR and Western blot in human MDA-MB-231 breast cancer cells. Direct binding of the above-mentioned miRNAs to the SOX9 3'-UTR was assessed by luciferase reporter assays and site-directed mutagenesis. Expression levels of the investigated miRNAs in tumor samples versus healthy tissues were analyzed in silico using publicly available databases. Transfection of miR-134-3p, miR-224-3p, or miR-6859-3p reduced SOX9 expression on mRNA and protein level. Reporter assays proved direct binding of miR-134-3p and miR-224-3p to the SOX9 3'-UTR in MDA-MB-231 and MCF-7 cells. Expression analysis performed in silico revealed reduced expression of both miRNAs in breast cancer tissues. We describe three novel miRNAs targeting SOX9 in human breast cancer cell lines. Among them miR-134-2p and miR-224-3p might act as tumor suppressors, whose down-regulation induces elevated SOX9 levels thereby promoting breast cancer progression.
转录因子 SOX9 是乳腺癌进展的重要介质。miRNA 是小的非编码 RNA，在与相应 mRNA 分子的 3'-UTR 区域相互作用时抑制靶基因的翻译。miRNA 表达失调参与癌症的标志，如持续增殖和凋亡抑制。在这里，我们研究了两种乳腺癌细胞系中 SOX9 表达的 miRNA 介导调节，从而为推动乳腺癌进展的细胞机制提供了进一步的见解。通过 qPCR 和 Western blot 在人 MDA-MB-231 乳腺癌细胞中分析了 miR-134-3p、miR-224-3p 和 miR-6859-3p 对 SOX9 表达的调节作用。通过荧光素酶报告基因测定和定点突变评估了上述 miRNA 与 SOX9 3'-UTR 的直接结合。使用公共可用数据库在计算机上分析了肿瘤样本与健康组织中研究的 miRNA 的表达水平。miR-134-3p、miR-224-3p 或 miR-6859-3p 的转染降低了 MDA-MB-231 和 MCF-7 细胞中 SOX9 的 mRNA 和蛋白水平表达。报告基因测定证明了 miR-134-3p 和 miR-224-3p 在 MDA-MB-231 和 MCF-7 细胞中直接结合 SOX9 3'-UTR。计算机上的表达分析显示这两种 miRNA 在乳腺癌组织中的表达降低。我们在人乳腺癌细胞系中描述了三种靶向 SOX9 的新 miRNA。其中 miR-134-2p 和 miR-224-3p 可能作为肿瘤抑制因子，其下调诱导 SOX9 水平升高，从而促进乳腺癌进展。
