Expression data from cell lines forced expressed PGC7/Stella

Global DNA hypomethylation and DNA hypermethylation of promoter regions—including tumor suppressor genes—are frequently detected in human cancers. Although many studies have suggested a contribution to carcinogenesis, it is still unclear whether the aberrant DNA hypomethylation observed in tumors is a consequence or a cause of cancer. We found that overexpression of Stella (also known as PGC7, Dppa3), a maternal factor required for the maintenance of DNA methylation in early embryos, induced global DNA hypomethylation and transformation in NIH3T3 cells. This hypomethylation was due to the binding of Stella to Np95 (also known as Uhrf1, ICBP90) and the subsequent impairment of Dnmt1 localization. In addition, enforced expression of Stella enhanced the metastatic ability of B16 melanoma cells through the induction of metastasis-related genes by inducing DNA hypomethylation of their promoter regions. Such DNA hypomethylation itself causes cellular transformation and metastatic ability. These data provide new insight into the function of global DNA hypomethylation in carcinogenesis. We used microarrays to detail the global programme of gene expression by PGC7/Stella overexpression. RNA was extracted from NIH3T3 or B16F10 murine cell lines overexpressed PGC7/Stella and was hybridized on Affymetrix microarrays. We compared gene expression levels between control and PGC7/Stella-overexpressed cells. Microarray analysis was performed in NIH3T3 cells including two independent Stella-expressing NIH3T3 clones and a mixture of Stella-expressing NIH3T3 clones and in B16-F10 cells including three independent Stella-expressing B16-F10 clones.

在人类癌症中，常可检测到全基因组DNA低甲基化以及启动子区域（包含抑癌基因区域）的DNA高甲基化。尽管诸多研究已提示其在肿瘤发生发展中发挥作用，但肿瘤中出现的异常DNA低甲基化究竟是癌症的诱因还是结果，目前仍不明确。本研究发现，早期胚胎DNA甲基化维持所必需的母源因子Stella（亦称PGC7、Dppa3）在NIH3T3细胞中过表达时，可诱导全基因组DNA低甲基化与细胞转化。该低甲基化现象源于Stella与Np95（亦称Uhrf1、ICBP90）的结合，以及后续Dnmt1定位功能的受损。此外，Stella的强制过表达可通过诱导转移相关基因启动子区域的DNA低甲基化，上调这些基因的表达，从而增强B16黑色素瘤细胞的转移能力。此类DNA低甲基化本身即可引发细胞转化与转移能力的获得。上述研究结果为阐明全基因组DNA低甲基化在肿瘤发生中的作用机制提供了新的视角。本研究通过微阵列技术，详细解析了PGC7/Stella过表达所调控的全基因组基因表达谱。我们从过表达PGC7/Stella的NIH3T3或B16F10小鼠细胞系中提取总RNA，将其与Affymetrix微阵列芯片进行杂交。我们对对照组与PGC7/Stella过表达细胞的基因表达水平进行了比较分析。本研究的微阵列分析涵盖两组样本：NIH3T3细胞组，包含2株独立的Stella表达阳性NIH3T3克隆以及1株Stella表达阳性NIH3T3克隆的混合样本；B16-F10细胞组，包含3株独立的Stella表达阳性B16-F10克隆。