McConnell M J, Cunliffe H E, Chua L J, Ward T A, Eccles M R
Department of Biochemistry, University of Otago, Dunedin, New Zealand.
Oncogene. 1997 Jun 5;14(22):2689-700. doi: 10.1038/sj.onc.1201114.
PAX2 is a member of the paired box family of genes with an important role in kidney, genital tract and eye development. Another gene essential for kidney and genital tract development is the Wilms tumour gene, WT1. PAX2 and WT1 encode transcription factors expressed during fetal kidney development in patterns that overlap both spatially and temporally. The overlap of PAX2 and WT1 expression in fetal kidney prompted us to determine whether PAX2 regulates the WT1 gene. To investigate this possibility, the WT1 promoter and a series of WT1 promoter deletion fragments were cloned into a luciferase reporter vector, and used in co-transfection experiments with PAX2 expression constructs. PAX2 transactivated the WT1 promoter up to 35-fold in CHO-K1 cells, and from four- to sevenfold in 293 cells. Two regions of the WT1 promoter were required in the same promoter construct for efficient transactivation by PAX2 in CHO-K1 cells, and purified recombinant PAX2 protein was found to bind to two sites in the WT1 promoter, at -205/-230 and +377/+402. Removal of WT1 promoter sequences containing the -205/-230, or +377/+402 binding sites abolished transactivation of the WT1 promoter by PAX2 in CHO-K1 cells, and had a differential effect on transactivation of the WT1 promoter in 293 cells, depending on the PAX2 isoform used. A fragment containing the -205/-230 site alone could be transactivated by PAX2. These findings suggest that PAX2 is a tissue-specific modulator of WT1 expression, and is involved in cell growth control via WT1.
PAX2是配对盒基因家族的成员，在肾脏、生殖道和眼睛发育中起重要作用。另一个对肾脏和生殖道发育至关重要的基因是威尔姆斯瘤基因WT1。PAX2和WT1编码在胎儿肾脏发育过程中表达的转录因子，其表达模式在空间和时间上都有重叠。PAX2和WT1在胎儿肾脏中的表达重叠促使我们确定PAX2是否调节WT1基因。为了研究这种可能性，将WT1启动子和一系列WT1启动子缺失片段克隆到荧光素酶报告载体中，并用于与PAX2表达构建体的共转染实验。PAX2在CHO-K1细胞中使WT1启动子的转录激活高达35倍，在293细胞中为4至7倍。在同一启动子构建体中，WT1启动子的两个区域是PAX2在CHO-K1细胞中有效转录激活所必需的，并且发现纯化的重组PAX2蛋白与WT1启动子中的两个位点结合，分别位于-205/-230和+377/+402。去除包含-205/-230或+377/+402结合位点的WT1启动子序列消除了PAX2在CHO-K1细胞中对WT1启动子的转录激活，并且根据所使用的PAX2同工型，对293细胞中WT1启动子的转录激活有不同的影响。仅包含-205/-230位点的片段可被PAX2转录激活。这些发现表明PAX2是WT1表达的组织特异性调节因子，并通过WT1参与细胞生长控制。
