Scibetta Angelo G, Santangelo Samantha, Coleman Julia, Hall Debbie, Chaplin Tracy, Copier John, Catchpole Steve, Burchell Joy, Taylor-Papadimitriou Joyce
Breast Cancer Biology Group, King's College London School of Medicine, 3rd Floor, Thomas Guy House, Guy's Hospital, London SE1 9RT, United Kingdom.
Mol Cell Biol. 2007 Oct;27(20):7220-35. doi: 10.1128/MCB.00274-07. Epub 2007 Aug 20.
The PLU-1/JARID1B nuclear protein, which is upregulated in breast cancers, belongs to the ARID family of DNA binding proteins and has strong transcriptional repression activity. To identify the target genes regulated by PLU-1/JARID1B, we overexpressed or silenced the human PLU-1/JARID1B gene in human mammary epithelial cells by using adenovirus and RNA interference systems, respectively, and then applied microarray analysis to identify candidate genes. A total of 100 genes showed inversely correlated differential expression in the two systems. Most of the candidate genes were downregulated by the overexpression of PLU-1/JARID1B, including the MT genes, the tumor suppressor gene BRCA1, and genes involved in the regulation of the M phase of the mitotic cell cycle. Chromatin immunoprecipitation assays confirmed that the metallothionein 1H (MT1H), -1F, and -1X genes are direct transcriptional targets of PLU-1/JARID1B in vivo. Furthermore, the level of trimethyl H3K4 of the MT1H promoter was increased following silencing of PLU-1/JARID1B. Both the PLU-1/JARID1B protein and the ARID domain selectively bound CG-rich DNA. The GCACA/C motif, which is abundant in metallothionein promoters, was identified as a consensus binding sequence of the PLU-1/JARID1B ARID domain. As expected from the microarray data, cells overexpressing PLU-1/JARID1B have an impaired G(2)/M checkpoint. Our study provides insight into the molecular function of the breast cancer-associated transcriptional repressor PLU-1/JARID1B.
在乳腺癌中上调的PLU-1/JARID1B核蛋白属于DNA结合蛋白的ARID家族，具有很强的转录抑制活性。为了鉴定受PLU-1/JARID1B调控的靶基因，我们分别使用腺病毒和RNA干扰系统在人乳腺上皮细胞中过表达或沉默人PLU-1/JARID1B基因，然后应用微阵列分析来鉴定候选基因。共有100个基因在这两个系统中呈现出反向相关的差异表达。大多数候选基因在PLU-1/JARID1B过表达时被下调，包括金属硫蛋白（MT）基因、肿瘤抑制基因BRCA1以及参与有丝分裂细胞周期M期调控的基因。染色质免疫沉淀试验证实，金属硫蛋白1H（MT1H）、-1F和-1X基因是PLU-1/JARID1B在体内的直接转录靶标。此外，PLU-1/JARID1B沉默后，MT1H启动子的三甲基化H3K4水平升高。PLU-1/JARID1B蛋白和ARID结构域均选择性地结合富含CG的DNA。在金属硫蛋白启动子中大量存在的GCACA/C基序被鉴定为PLU-1/JARID1B ARID结构域的共有结合序列。正如微阵列数据所预期的那样，过表达PLU-1/JARID1B的细胞G(2)/M期检查点受损。我们的研究为乳腺癌相关转录抑制因子PLU-1/JARID1B的分子功能提供了深入了解。
