The Wilms' tumor gene WT1 is a common marker of progenitor cells in fetal liver

It is well known that the Wilms' tumor gene WT1 plays an important role in cell proliferation and differentiation, and in organ development. In this study, to examine the role of the WT1 gene in lineage determination, fetal liver cells from LacZ-transgenic mice, in which WT1 expression was marked by the expression of the LacZ gene driven by WT1 promoter, were FACS-sorted according to LacZ expression of high (LacZ(++)) or undetectable (LacZ(-)) levels, which paralleled endogenous WT1 expression levels. LacZ(++) fetal liver cells were enriched by hepatocyte and endothelial progenitor cells. These results indicated that WT1 expression is a common marker of both hepatocyte and endothelial progenitors. These results also implied a role of the WT1 gene in lineage determination.     

Regulation of ornithine decarboxylase gene expression by the Wilms' tumor suppressor WT1.

The importance of ornithine decarboxylase (ODC) to cell proliferation is underscored by the complex array of cell-specific mechanisms invoked to regulate its synthesis and activity. Misregulation of ODC has severe negative consequences on normal cell function, including the acquisition of tumorigenic growth properties by cells overexpressing ODC. We hypothesize that ODC gene expression is a candidate target for the anti-proliferative function of certain tumor suppressors. Here we show that the Wilms' tumor suppressor WT1 binds to multiple sites within the human ODC promoter, as determined by DNase I protection and methylation interference assays. The expression of WT1 in transfected HCT 116, NIH/3T3 and HepG2 cells represses activity of the ODC promoter controlling expression of a luciferase reporter gene. In contrast WT1 expression enhances ODC promoter activity in SV40-transfected HepG2 cells. Both the extent of modulation of ODC gene expression and the mediating WT1 binding elements are cell specific. Constructs expressing WT1 deletion mutants implicate two regions required for repressor function, as well as an intrinsic activation domain. Understanding the regulation of ODC gene expression by WT1 may provide valuable insights into the roles of both WT1 and ODC in development and tumorigenesis.     

Wilms’ Tumor Gene 1 (WT1) Silencing Inhibits Proliferation of Malignant Peripheral Nerve Sheath Tumor sNF96.2 Cell Line

Wilms’ tumor gene 1 (WT1) plays complex roles in tumorigenesis, acting as tumor suppressor gene or an oncogene depending on the cellular context. WT1 expression has been variably reported in both benign and malignant peripheral nerve sheath tumors (MPNSTs) by means of immunohistochemistry. The aim of the present study was to characterize its potential pathogenetic role in these relatively uncommon malignant tumors. Firstly, immunohistochemical analyses in MPNST sNF96.2 cell line showed strong WT1 staining in nuclear and perinuclear areas of neoplastic cells. Thus, we investigated the effects of silencing WT1 by RNA interference. Through Western Blot analysis and proliferation assay we found that WT1 knockdown leads to the reduction of cell growth in a time- and dose-dependent manner. siWT1 inhibited proliferation of sNF96.2 cell lines likely by influencing cell cycle progression through a decrease in the protein levels of cyclin D1 and inhibition of Akt phosphorylation compared to the control cells. These results indicate that WT1 knockdown attenuates the biological behavior of MPNST cells by decreasing Akt activity, demonstrating that WT1 is involved in the development and progression of MPNSTs. Thus, WT1 is suggested to serve as a potential therapeutic target for MPNSTs.     

Significant reduction of WT1 gene expression,possibly due to epigenetic alteration in Wilms' tumor

WT1 at 11p13 is a tumor suppressor gene, an aberration of which causes Wilms' tumor (WT). Since WT1 expression is reduced in a certain proportion of WTs and its mutation is found only in 10-20% of WTs, we examined WT1 gene silencing due to epigenetic alteration in a total of 22 WTs. WT1 expression was significantly reduced in half of WTs without any mutation in the WT1 gene itself, suggesting that the reduction of expression was possibly epigenetic. We found promoter hypermethylation in one WT with loss of heterozygosity (LOH) and showed that promoter methylation reduced reporter gene activity by a reporter assay. These data suggested that methylation was an epigenetic mechanism leading to WT1 silencing and that the expression-reduced allele by hypermethylation combined with LOH was consistent with the revised two-hit model. In addition, as the beta-catenin mutation is frequently associated with the WT1 mutation, the association of WT1 silencing with the beta-catenin mutation was also investigated. beta-catenin mutated in only one WT without WT1 silencing, suggesting that the beta-catenin mutation was not associated with the reduction of WT1 expression.     

The human sex-determining gene SRY is a direct target of WT1

The product of the Wilms' tumor gene, WT1, is essential for male sex determination and differentiation in mammals. In addition to causing Wilms' tumor, mutations in WT1 often cause two distinct but overlapping urogenital defects in men, Denys-Drash syndrome and Frasier syndrome. In this study we investigated the regulation of the sex determination gene SRY by WT1. Our results showed that WT1 up-regulates the SRY gene through the proximal early growth response gene-1-like DNA-binding sequences in the core promoter. Mutant WT1 proteins in Denys-Drash syndrome patients were unable to activate this promoter. These mutants did not act in a dominant negative manner, as expected over the wild-type WT1 in this promoter. We also found that WT1 could transactivate the endogenous SRY gene. These observations, together with the overlapping expression patterns of WT1 and SRY in human gonads, led us to propose that WT1 regulates SRY in the initial sex determination process in humans and activates a cascade of genes ultimately leading to the complete organogenesis of the testis.     

Downregulation of Wilms' tumor 1 protein inhibits breast cancer proliferation

High levels of Wilms' Tumor 1 (WT1) mRNA have been correlated with poor prognosis in breast cancer patients. However, the function of WT1 protein in breast cancer is not known. We observed that the levels of WT1 protein correlated with the proliferation of breast cancer cells. When the proliferation of breast cancer cells was stimulated by 17beta-estradiol, WT1 protein expression increased. But when the proliferation of breast cancer cells was inhibited by tamoxifen or all-trans retinoic acid (ATRA), WT1 protein expression decreased. We hypothesize that WT1 protein plays a role in regulating breast cancer cell proliferation. Using liposome-incorporated WT1 antisense oligodeoxynucleotides, we found that downregulation of WT1 protein expression led to breast cancer growth inhibition and reduced cyclin D1 protein levels. These results indicate that WT1 protein contributes to breast cancer progression by promoting breast cancer cell proliferation.