Department of Genetics, University of Texas M.D. Anderson Cancer Center, 1515 Holcombe Blvd, Unit 1010, Houston, TX 77030, USA.
Exp Cell Res. 2013 Mar 10;319(5):612-22. doi: 10.1016/j.yexcr.2012.12.021. Epub 2013 Jan 2.
Wilms tumor gene WT1 encodes a zinc finger-containing transcription factor which is required for renal development. Mutations in WT1 are observed in 20% of Wilms tumors (a pediatric kidney cancer), but the in vivo WT1 targets and associated molecular pathways involved in the etiology of Wilms tumor are still elusive. To identify WT1 targets we performed genome-wide comprehensive expression profiling using Affymetrix Gene Chip Mouse Genome 430 2.0 Arrays, comparing E13.5 mouse kidneys in which Wt1 had been somatically ablated with littermate controls. We identified Usp18 as the most differentially expressed gene in mutant kidney. Using tetracycline inducible cells we demonstrated a repressive effect of WT1 on USP18 expression. Conversely, knockdown of WT1 led to the upregulation of Usp18. Furthermore, direct binding of WT1 to the Usp18 promoter was demonstrated by ChIP assay. Overexpression of USP18 in murine and human cell lines resulted in cell proliferation. Additionally, Usp18 upregulation was observed in a mouse model of Wilms tumor. Taken together our data demonstrate that Usp18 is a transcriptional target of WT1 and suggest that increased expression of USP18 following WT1 loss contributes to Wilms tumorigenesis.
WT1 基因编码一种含锌指的转录因子，该因子是肾脏发育所必需的。WT1 突变在 20%的威尔姆斯瘤（一种儿童肾部癌症）中被观察到，但 WT1 在体内的靶标以及与威尔姆斯瘤病因相关的分子途径仍然难以捉摸。为了鉴定 WT1 的靶标，我们使用 Affymetrix Gene Chip Mouse Genome 430 2.0 Arrays 进行了全基因组综合表达谱分析，将 Wt1 在体内被体细胞灭活的 E13.5 期小鼠肾脏与同窝对照进行了比较。我们发现 Usp18 是突变肾脏中差异表达最明显的基因。通过四环素诱导细胞，我们证明了 WT1 对 USP18 表达的抑制作用。相反，WT1 的敲低导致 Usp18 的上调。此外，通过 ChIP 检测证实了 WT1 与 Usp18 启动子的直接结合。USP18 在鼠和人类细胞系中的过表达导致细胞增殖。此外，在威尔姆斯瘤的小鼠模型中观察到 Usp18 的上调。总之，我们的数据表明 Usp18 是 WT1 的转录靶标，并表明 WT1 缺失后 USP18 的表达增加有助于威尔姆斯瘤的发生。
